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Downregulation of p21-activated kinase-1 inhibits the growth of gastric cancer cells involving cyclin B1
International Journal of Cancer
Abstract
p21-Activated kinase 1 (Pak1), a serine/threonine kinase, has been implicated in cytoskeletal remodelling, cell motility, apoptosis and transformation. However, the role of Pak1 in gastric cancer remains unclear. In this study, we detected Pak1 expression in gastric cancer tissues from 40 patients by western blot. Overexpression of Pak1 was associated with progression, metastasis and prognosis of gastric cancer. In addition, we found that knockdown of Pak1 expression significantly inhibited anchorage-dependent and anchorage-independent growth in gastric cancer cells, and markedly inhibited gastric cancer cell xenograft tumor growth. In further study, data showed that Pak1 could regulate the expression of cyclin B1 at the mRNA and protein levels, and impact the subcellular distribution and the promoter activity of cyclin B1. Results from deletion and mutant analysis supplied a new NF-kappaB binding sites at position -321 of cyclin B1 promoter, and indicated that Pak1 regulated the transcription of cyclin B1 in gastric cancer through NF-kappaB. In conclusion, Pak1 may be a potential prognostic marker and therapeutic target in gastric cancer.
... 15 Our previous studies and those of others have shown that Pak1 signaling has a profound effect on gastric cancer. 2 MicroRNAs (miRNAs) are non-coding small RNA molecules that regulate gene expression at the post-transcriptional level by binding to the 3 0 UTR of their target mRNAs and repress protein production by destabilizing mRNA and silencing translation. 16 Many miRNAs have been shown to play crucial roles at a number of steps that confer tumor metastasis, including EMT, anoikis, angiogenesis, invasion, and migration. ...
... First, we performed miRNA chip analysis (CapitalBio) to identify differentially expressed miRNAs in Pak1-knockdown gastric cancer cells (BGC823; i.e., stable knockdown cell clone). 2 A notable candidate gene we found was miR-132. Its level was significantly higher in Pak1knockdown cells than in control cells ( Figure S1). ...
... Our previous studies consistently pointed to a crucial role for Pak1 in gastric cancer progression and metastasis. 2 Recently, several studies have shown that Pak1 is an important target gene of the miRNA signaling pathway. In breast cancer cells, the expression and function of Pak1 is regulated by miR-7. ...
... In addition, PAK1 plays key roles in nuclear signaling and activation of the JNK/SAPK and p38MAPK pathways [4,5]. Although it has been suggested that PAK1 influences the prognosis of various cancer types [3,[6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21], current knowledge of the contribution of PAK1 to cancer prognosis remains limited. ...
... In the present study, we used a statistical approach to systematically investigate the association between PAK1 and the prognosis of solid tumors. Over the past decade, a series of studies have focused on the relationship between PAK1 expression and solid cancer prognosis [3,[6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21], but the results of those individual studies were not conclusive. We therefore performed a meta-analysis using a relatively large sample from 15 studies (3068 patients) with the aim of conclusively determining the relationship between PAK1 and prognosis in patients with solid tumors. ...
... Through searching and selection, a final list of 17 studies [3,[6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21] was collected for qualitative synthesis ( Figure 1). The participants in the studies spanned different ethnicities (11 studies of Asians and 6 studies of Caucasians) and cancer types (3 studies of breast cancer, 2 colorectal cancer, 2 gastric cancer, 2 head and neck cancer, 2 ovarian cancer, 1 gastroesophageal junction adenocarcinoma, 1 glioblastoma, 1 hepatocellular carcinoma, 1 pancreatic cancer, 1 renal cell carcinoma, and 1 urothelial carcinoma of the upper urinary tract). ...
p21 protein (Cdc42/Rac)-activated kinase 1 (PAK1) expression appears to be predictive of prognosis in various solid tumors, though the evidence is not yet conclusive. We therefore performed a meta-analysis to explore the relationship between PAK1 and prognosis in patients with solid tumors. Relevant publications were searched in several widely used databases, and 15 studies (3068 patients) were included in the meta-analysis. Pooled hazard ratios (HRs) and 95% confidence intervals (CIs) were calculated to evaluate the strength of the association between PAK1 and prognosis. Associations between PAK1 expression and prognosis were observed for overall survival (HR = 2.81, 95% CI = 1.07-7.39) and disease-specific survival (HR = 2.15, 95% CI = 1.47-3.16). No such association was detected for time to tumor progression (HR = 1.78, 95% CI = 0.99-3.21).Our meta-analysis thus indicates that PAK1 expression may be a predictive marker of overall survival and disease-specific survival in patients with solid tumors.
... Amplification of Pak1 has been found in breast cancer, kidney cancer, liver cancer and stomach cancer. Moreover, Pak1 has been reported to induce proliferation, motility and invasion of these cancer cells (13)(14)(15)(16). Pak1 is believed to be involved in several cell signaling pathways, such as mitogenactivated protein kinases (MAPK) and nuclear factor-κB (NF-κB) (3,17). ...
... Overexpression of Pak1 has been reported to increase cell proliferation in breast cancer by up-regulating the expression of cyclin D1 (20) and in gastric cancer by up-regulating the expression of cyclin B1 (16). However, the role of Pak1 in the progression of CRC remained elusive. ...
... Given that Pak1 is also activated by several other physiological signals, such as guanine nucleotide factor pix (28), filamin A (29), adaptor protein Nck (30) and phosphatidylinositol 3-kinase/Akt (10), activation of Pak1 in CRC cells may be induced through Cdc42/Rac1-independent mechanisms. Although Pak1 has been reported to be involved in cell proliferation (7,14,16), we observed little difference in the cell proliferation rates between Pak1 stable cells and control cells, suggesting that Pak1 may play different roles in the development of CRC. In conclusion, we have shown that Pak1 plays an important role in cancer metastasis. ...
... The PAK1 is a serine/threonine protein kinase which is stimulated by active Rac1 and Cdc42-GTPases [3][4][5], and has been found to be key regulator of cancer-cell signaling networks [6]. There has been mounting evidence that PAK1 is tightly related to the progression of cancer and may become a promising diagnostic and therapeutic target for cancer [7][8][9][10][11][12]. Therefore, it is worthwhile to study the novel binding partners of PAK1. ...
... Emerging data suggest that PAK1 is overexpressed in human cancer [7,8], including gastric cancer [9][10][11]. The oncogenic activity of PAK1 is quite certain. ...
... Stable overexpressed cell lines were seeded in 60-mm plates to perform the flow cytometry and cell counts assays as described in our previous paper [11,24]. For colony formation assay, 500 cells were plated in six-well plates to assess the proliferation potential of cells and incubated at 37°C in a 5% CO 2 incubator. ...
To date, microrchidia (MORC) family CW-type zinc-finger 2 (MORC2), has been found to be involved in p21-activated kinase1 (PAK1) pathway to maintain genomic integrity. Here, we explore its novel role in cancer. We demonstrate that PAK1-mediated MORC2 phosphorylation promotes cell cycle progression, defective phosphorylation of MORC2-S677A results in attenuated cell proliferation and tumorigenicity of gastric cancer cells, which is significantly enhanced in overexpression of phospho-mimic MORC2-S677E form, suggesting the importance of MORC2 phosphorylation in tumorigenesis. More importantly, phosphorylation of MORC2 correlates positively with PAK1 expression in clinical gastric cancer. Furthermore, high expression of PAK1 and phosphorylation of MORC2 appear to be associated with poor prognosis of clinical gastric cancer. Collectively, these findings revealed a novel function of MORC2 phosphorylation in promoting gastric cell proliferation in vitro and tumorigenesis in vivo, suggesting that blocking PAK1-mediated MORC2 phosphorylation might be a potential therapeutic strategy for gastric tumorigenesis.
... PAK activation affects NF-κB, 16 . RAS activation initiates PAK1, RAF/mitogen activated protein kinase (MAPK) and phospoinositide 3-kinase (PI3K)/AKT signalling. ...
... 141 PAK1 knockdown in a GC xenograft model reduced tumour size. 16 Stable and transient knockdown of PAK1 within GC cell lines (BGC823 and ACS) induced G2 cell cycle arrest, reduced proliferation and inhibited anchorage independent cell growth. 16 Overexpression of the catalytically active (CA-PAK1) mutant enhanced the expression of cyclin B1, a critical component of the G2/M transition of the cell cycle, whereas PAK1 inhibition decreased the transcription of cyclin B1. ...
... 16 Stable and transient knockdown of PAK1 within GC cell lines (BGC823 and ACS) induced G2 cell cycle arrest, reduced proliferation and inhibited anchorage independent cell growth. 16 Overexpression of the catalytically active (CA-PAK1) mutant enhanced the expression of cyclin B1, a critical component of the G2/M transition of the cell cycle, whereas PAK1 inhibition decreased the transcription of cyclin B1. PAK1 deficient clones had reduced activity of NF-κB, and pharmacological inhibition of NF-κB reduced the activity of cyclin B1. ...
P-21 activated kinases (PAKs) are effectors of Rac1/Cdc42 which coordinate signals from the cell membrane to the nucleus. Activation of PAKs drive important signalling pathways including mitogen activated protein kinase, phospoinositide 3-kinase (PI3K/AKT), NF-κB and Wnt/β-catenin. Intestinal PAK1 expression increases with inflammation and malignant transformation, although the biological relevance of PAKs in the development and progression of GI disease is only incompletely understood. This review highlights the importance of altered PAK activation within GI inflammation, emphasises its effect on oncogenic signalling and discusses PAKs as therapeutic targets of chemoprevention.
... Although PAK1 has been studied in gastrointestinal cancers, the role of PAK1 in GEJ adenocarcinoma remains missing [21,24,25,31,33,58]. Our analysis using IHC, immunoblot analysis, and data mining indicates that PAK1 is highly differentially over-expressed in GEJ tumor tissues compared with noncancerous tissues. ...
... Our analysis using IHC, immunoblot analysis, and data mining indicates that PAK1 is highly differentially over-expressed in GEJ tumor tissues compared with noncancerous tissues. Similarly to our study, PAK1 protein overexpression and copy number gain were found in more than 40% of gastric cancer tissues [25,59], and advanced gastric cancer tissues express higher PAK1 levels than do matched noncancerous adjacent mucosa [58]. PAK1 was also reported to be amplified in circulating esophageal squamous cell carcinoma cells [60]. ...
... [I+II]) in gastric cancer [58]. Data derived from both IHC of specimens and analysis of silicon data set did not show an association between PAK1 and PCNA, a cell proliferation marker, in GEJ adenocarcinoma. ...
Gastroesophageal junction (GEJ) adenocarcinoma is a lethal cancer with rising incidence, yet the molecular biomarkers that have strong prognostic impact and also hold great therapeutic promise remain elusive. We used a data mining approach and identified the p21 protein-activated kinase 1 (PAK1), an oncogene and drugable protein kinase, to be among the most promising targets for GEJ adenocarcinoma. Immunoblot analysis and data mining demonstrated that PAK1 protein and mRNA were upregulated in cancer tissues compared to the noncancerous tissues. Immunohistochemistry revealed PAK1 overexpression in 72.6% of primary GEJ adenocarcinomas (n = 113). A step-wise increase in PAK1 levels was noted from paired normal epithelium, to atypical hyperplasia and adenocarcinoma. PAK1 overexpression in tumor was associated with lymph node (LN) metastasis (P<0.001), advanced tumor stage (P<0.001), large tumor size (P = 0.006), residual surgical margin (P = 0.033), and unfavorable overall survival (P<0.001). Multivariate analysis showed PAK1 overexpression is an independent high-risk prognostic predictor (P<0.001). Collectively, PAK1 is overexpressed during tumorigenic progression and its upregulation correlates with malignant properties mainly relevant to invasion and metastasis. PAK1 expression could serve as a prognostic predictor that holds therapeutic promise for GEJ adenocarcinoma.
... PAK1 has been associated with gastric cancer cell migration, invasion [163], and hematogenous metastasis [164]. In gastric cancer patients, high PAK1 expression was associated with advanced-stage tumors, distant metastasis, and reduced survival [165,166]. Recent studies also indicate PAK1 activation by several long non-coding RNAs and miR-NAs in the progression and EMT of gastric cancers [167,168]. In a mouse model of intestinal cancer, PAK1 has been demonstrated to initiate tumor development [169]. ...
... [ [163][164][165][166][167][168][169][170] Pancreatic Ductal Carcinoma PAK1 activation is linked to autophagy and immune evasion. Co-targeting PAK1 sensitizes cancer cells to chemotherapeutics in vitro. ...
Metastatic prostate cancer (mPCa) has limited therapeutic options and a high mortality rate. The p21-activated kinase (PAK) family of proteins is important in cell survival, proliferation, and motility in physiology, and pathologies such as infectious, inflammatory, vascular, and neurological diseases as well as cancers. Group-I PAKs (PAK1, PAK2, and PAK3) are involved in the regulation of actin dynamics and thus are integral for cell morphology, adhesion to the extracellular matrix, and cell motility. They also play prominent roles in cell survival and proliferation. These properties make group-I PAKs a potentially important target for cancer therapy. In contrast to normal prostate and prostatic epithelial cells, group-I PAKs are highly expressed in mPCA and PCa tissue. Importantly, the expression of group-I PAKs is proportional to the Gleason score of the patients. While several compounds have been identified that target group-I PAKs and these are active in cells and mice, and while some inhibitors have entered human trials, as of yet, none have been FDA-approved. Probable reasons for this lack of translation include issues related to selectivity, specificity, stability, and efficacy resulting in side effects and/or lack of efficacy. In the current review, we describe the pathophysiology and current treatment guidelines of PCa, present group-I PAKs as a potential druggable target to treat mPCa patients, and discuss the various ATP-competitive and allosteric inhibitors of PAKs. We also discuss the development and testing of a nanotechnology-based therapeutic formulation of group-I PAK inhibitors and its significant potential advantages as a novel, selective, stable, and efficacious mPCa therapeutic over other PCa therapeutics in the pipeline.
... The lncRNA LUCAT1 is probably linked to PAK1 via hsa-miR-377-3p, which is a potential ceRNA regulatory relationship. According to previous studies, PAK1 is involved in the development and progression of GC [19,20]. Therefore, we selected the lncRNA LUCAT1 to validate its relationship with GC. ...
... S1). According to previous studies, PAK1is involved in the development and progression of GC [19,20], suggesting that LUCAT1 may participate in GC tumorigenesis. ...
Gastric cancer(GC) remains the fourth-leading malignancy worldwide and has a high mortality rate.Accumulating evidence reveals that long noncoding RNAs (lncRNAs) play essential roles in tumorigenesis and metastasis and can be used as potential biomarkers for diagnosis and prognosis. The current study sought to define the lncRNA LUCAT1 and verify its malignant biological behaviors in GC. We conducted bioinformatic analysis to screen differentially expressed lncRNAs between GC tissue and paracancerous tissue. Gene expression profiles were downloaded from the National Center of Biotechnology Information Gene Expression Omnibus(GEO). Real-time quantitative polymerase chain reaction (RT-qPCR) was carried out to verify LUCAT1 expression in both GC tissue and paracancerous tissue. Furthermore, the associations between LUCAT1 and clinical features were analyzed. In addition, the malignant behaviors of LUCAT1 in GC were investigated by knocking down LUCAT1 expression in the SGC7901 and AGS cell lines. The results indicated that LUCAT1 expression was obviously upregulated in GC samples compared with paracancerous tissue samples. Moreover, the expression pattern of LUCAT1 showed close correlations with tumor diameter (P<0.001), differentiation grade (P=0.026), and lymphnode metastasis(LNM)status (P=0. 020). In vitro, shRNA-mediated knockdown of LUCAT1 expression inhibited proliferation, migration, and invasion and led to S-phase cell cycle arrest and apoptosisin GC cells. Thus, the lncRNA LUCAT1 may be used as a potential biomarker for early signs of LNM in GC and may play a crucial role in the development of GC.
... Pak1, a serine/threonine kinase, plays critical roles in cytoskeletal remodeling, cell motility, apoptosis, and transformation 1,2 . It effects many distinct signal transduction pathways. ...
... Table 1-Association between the expression of miR-132 with clinicopathologic features in patients with gastric cancer 1 Median of relative expression, with 25th-75th percentile in parenthesis. 2 Mann-Whitney test (for 2 groups) or Kruskal Wallis test (for >2 groups). 3 Hepatic metastasis, pulmonary metastasis, bone metastasis and brain metastasis occurrence before operation or after operation within five years. ...
We, along with others, have shown previously that P21-activated kinase 1Pak1) played a pivotal role in gastric cancer progression and metastasis. However, whether Pak1 could control gastric cancer metastasis by regulating miRNAs has never been explored. Here, we report a novel mechanism of Pak1 on tumor metastasis. A detailed examination revealed that Pak1 interacted with and phosphorylated the serine 62 residue of ATF2, then blocked its translocation into the nucleus. We also confirmed ATF2 bound to the promoter of miR-132 and tightly regulated its transcription, and thus explained the regulatory mechanism of miR-132 by Pak1. Besides, miR-132 significantly reduced cell adhesion, migration and invasion of gastric cancer cells in vitro, and significantly prevented tumor metastasis in vivo. miR-132 specifically inhibited hematogenous metastasis, but not lymph node or implantation metastases. In order to further delineate the effects of Pak1/ATF2/miR-132 cascade on gastric cancer progression, we identified several targets of miR-132 using bioinformatics Targetscan algorithm. Notably, miR-132 reduced the expression of CD44, fibronectin (FN), and such inhibition enabled lymphocytes to home in on gastric cancer cells and inducing tumor apoptosis. Taken together, our studies establish a novel cell-signaling pathway and opens new possibilities for therapeutic intervention of gastric cancer.
... PKN1 encodes a protein kinase C super family member, regulating cell migration and gene expression [24]. This protein is associated with ovarian cancer [21], colorectal cancer [22], prostate cancer [23,24], breast cancer [25], and gastric cancer [26]. PPP1R9A encodes neurabin, which is involved in the synapse formation and function. ...
... Thus, we examined the association between these three genes and lung adenocarcinoma further in the following sections. Ovarian cancer [21], colorectal cancer [22], prostate cancer [23,24], breast cancer [25], gastric cancer [26] PPP1R9A protein phosphatase 1, regulatory subunit 9A ...
Epigenetic mechanisms such as DNA methylation or histone modifications are essential for the regulation of gene expression and development of tissues. Alteration of epigenetic modifications can be used as an epigenetic biomarker for diagnosis and as promising targets for epigenetic therapy. A recent study explored cancer-cell specific epigenetic biomarkers by examining different types of epigenetic modifications simultaneously. However, it was based on microarrays and reported biomarkers that were also present in normal cells at a low frequency. Here, we first analyzed multi-omics data (including ChIP-Seq data of six types of histone modifications: H3K27ac, H3K4me1, H3K9me3, H3K36me3, H3K27me3, and H3K4me3) obtained from 26 lung adenocarcinoma cell lines and a normal cell line. We identified six genes with both H3K27ac and H3K4me3 histone modifications in their promoter regions, which were not present in the normal cell line, but present in ≥85% (22 out of 26) and ≤96% (25 out of 26) of the lung adenocarcinoma cell lines. Of these genes, NUP210 (encoding a main component of the nuclear pore complex) was the only gene in which the two modifications were not detected in another normal cell line. RNA-Seq analysis revealed that NUP210 was aberrantly overexpressed among the 26 lung adenocarcinoma cell lines, although the frequency of NUP210 overexpression was lower (19.3%) in 57 lung adenocarcinoma tissue samples studied and stored in another database. This study provides a basis to discover epigenetic biomarkers highly specific to a certain cancer, based on multi-omics data at the cell population level.
... In breast cancer, nuclear expression of PAK1 in conjunction with phosphorylation of estrogen receptor on the PAK1 site (ser305) predicts resistance to tamoxifen therapy, while the cytoplasmic levels of PAK1 correlate with recurrence rate and mortality [109,110]. Similarly, in patients with gastric cancer higher levels of PAK1 were associated with advanced tumor stages, metastasis and reduced survival [111]. There are also numerous reports of elevated PAK1 activity in cell lines, although in most cases such reports cannot rule out that the changes had been selected or caused by in vitro cultivation of the cells. ...
... [110,119,[224][225][226][227][228] Colorectal carcinoma Elevated levels of protein; gene amplification mCGH; aCGH; FISH; IHC. [230] Gastric carcinoma Elevated levels of protein WB [111] Glioblastoma elevated levels of phosphorylated protein IHC [231] Hepatocellular carcinoma Elevated levels of RNA and protein qPCR,WB, IHC [232] NK and NK-like T cell lymphomas Gene amplification mCGH; aCGH [233] Ovarian carcinoma Elevated levels of RNA, total and phosphorylated protein; gene amplification. ...
... Pak1 regulates the mRNA and protein levels of Cyclin B1 for the promotion of progression and metastasis of gastric cancer as its knockdown resulted in the inhibition of the growth of gastric cancer cells and xenograft tumors. The mechanism of Cyclin B1 regulation via Pak1 involves the recruitment of NF-kB to the Cyclin B1 promoter site [184]. Cyclin B1 levels are also regulated post-transcriptionally via miR-663, which is a micro-RNA contributing to the suppression of the growth of gastric cancer cells [185]. ...
Gastric cancer predominantly occurs in adenocarcinoma form and is characterized by uncontrolled growth and metastases of gastric epithelial cells. The growth of gastric cells is regulated by the action of several major cell cycle regulators including Cyclins and Cyclin-dependent kinases (CDKs), which act sequentially to modulate the life cycle of a living cell. It has been reported that inadequate or over-activity of these molecules leads to disturbances in cell cycle dynamics, which consequently results in gastric cancer development. Manny studies have reported the key roles of Cyclins and CDKs in the development and progression of the disease in either in vitro cell culture studies or in vivo models. We aimed to compile the evidence of molecules acting as regulators of both Cyclins and CDKs, i.e., upstream regulators either activating or inhibiting Cyclins and CDKs. The review entails an introduction to gastric cancer, along with an overview of the involvement of cell cycle regulation and focused on the regulation of various Cyclins and CDKs in gastric cancer. It can act as an extensive resource for developing new hypotheses for future studies.
... The regulation of PAKs on gastric cancer progression is positive. The major target proteins and signaling pathways of PAK1 are introduced to regulate gastric cancer cell proliferation (MORC2 [110], Smad2/3 [111]), migration, metastasis and invasion (ATF2/miR-132 [112], RUFY3 [113]) and cell cycle processes (CyclinD1 [114], Cyclin B [115]). However, PAK4 mainly focuses on the regulation of the migration, metastasis and invasion (via LIMK1/Cofilin/microfilament [92], SCG10/microtubule [116], DGCR6L [117], Coro1C [118]) of gastric cancer cells. ...
Gastrointestinal tumors are the most common tumors, and they are leading cause of cancer deaths worldwide, but their mechanisms are still unclear, which need to be clarified to discover therapeutic targets. p21-activating kinase (PAK), a serine/threonine kinase that is downstream of Rho GTPase, plays an important role in cellular signaling networks. According to the structural characteristics and activation mechanisms of them, PAKs are divided into two groups, both of which are involved in the biological processes that are critical to cells, including proliferation, migration, survival, transformation and metabolism. The biological functions of PAKs depend on a large number of interacting proteins and the signaling pathways they participate in. The role of PAKs in tumors is manifested in their abnormality and the consequential changes in the signaling pathways. Once they are overexpressed or overactivated, PAKs lead to tumorigenesis or a malignant phenotype, especially in tumor invasion and metastasis. Recently, the involvement of PAKs in cellular plasticity, stemness and the tumor microenvironment have attracted attention. Here, we summarize the biological characteristics and key signaling pathways of PAKs, and further analyze their mechanisms in gastrointestinal tumors and others, which will reveal new therapeutic targets and a theoretical basis for the clinical treatment of gastrointestinal cancer.
... HLF-mediated miR-132 inhibited glioma cell invasion and radio resistance via TTK inhibition [94]. P21-activated kinase 1 (Pak1) is a serine/threonine kinase that has key functions in cell migration, apoptosis, and neoplastic transformation [95,96]. It regulates various cellular processes such as tumor cell invasion, drug resistance, angiogenesis, and EMT [97]. ...
Cancer as one of the leading causes of human deaths has always been one of the main health challenges in the world. Despite recent advances in therapeutic and diagnostic methods, there is still a high mortality rate among cancer patients. Late diagnosis is one of the main reasons for the high ratio of cancer related deaths. Therefore, it is required to introduce novel early detection methods. Various molecular mechanisms are associated with the tumor progression and metastasis. MicroRNAs (miRNAs) are a class of non-coding RNAs (ncRNAs) family that has important functions in regulation of the cellular processes such as cell proliferation, apoptosis, and tumor progression. Moreover, they have higher stability in body fluids compared with mRNAs which can be introduced as non-invasive diagnostic markers in cancer patients. MiR-132 has important functions as tumor suppressor or oncogene in different cancers. In the present review, we have summarized all of the studies which have been reported the role of miR-132 during tumor progressions. We categorized the miR-132 target genes based on their cell and molecular functions. Although, it has been reported that the miR-132 mainly functions as a tumor suppressor, it has also oncogenic functions especially in pancreatic tumors. MiR-132 mainly exerts its roles during tumor progressions by regulation of the transcription factors and signaling pathways. Present review clarifies the tumor specific molecular mechanisms of miR-132 to introduce that as an efficient non-invasive diagnostic marker in various cancers.
... p21activated kinase 1(Pak 1) is a key oncogenic signaling pathway, frequently involved in cellular processes and human malignancies [104]. KSHV-GPCR may drive Pak1, as an upstream stimulus of NF-κB activity during the cellular transformation in KS tumors [105]. A recent research revealed the crucial effect of cell adhesion molecule 1 (CADM1; involved in cell signaling and tumorigenesis) in survival of PEL cells and KSHV-induced tumorigenesis through chronic activation of NF-κB [101]. ...
Among the DNA tumor viruses Epstein-Barr virus (EBV) and Kaposi sarcoma herpesvirus (KSHV), account for a considerable percentage of virus-associated cancers. Deregulation of transcription factors signaling pathways is one of the most significant oncogenic characteristics of EBV and KSHV. NF-κB is a transcription factor that play a remarkable role in oncogenesis because of its function as a master regulator of a spectrum of genes involved in physiological and pathophysiological process. Constitutive activation of NF-κB is a frequent and well-described event in many human malignancies. Compelling evidence represent EBV and KSHV are capable of targeting different components of NF-κB cascade. Here, we summarized recent findings to clarify the precise relationship between dysregulation of NF-κB and EBV and KSHV-related malignancies. This essay also emphasizes on contribution of various viral products in developing cancer through alteration of NF-κB signaling pathway.
... PLK1 is an important cell cycle regulator that can enable mitotic entry by phosphorylating cdc25 and cyclinB1 [150,151]. Interestingly, PAK1 can also regulate cyclinB1 activity via transcription and expression enhancing through NF-KB activation in gastric cancer [152]. Moreover, the oncogenic transcription factor MYC, another crucial downstream effector of PAK1, can regulate several cyclin related kinases (CDK4/6, cyclin D, CDK2, and cyclin E) to promote cell cycle progression [153]. ...
The p21-Activated kinase 1 (PAK1), a member of serine-threonine kinases family, was initially identified as an interactor of the Rho GTPases RAC1 and CDC42, which affect a wide range of processes associated with cell motility, survival, metabolism, cell cycle, proliferation, transformation, stress, inflammation, and gene expression. Recently, the PAK1 has emerged as a potential therapeutic target in cancer due to its role in many oncogenic signaling pathways. Many PAK1 inhibitors have been developed as potential preclinical agents for cancer therapy. Here, we provide an overview of essential roles that PAK1 plays in cancer, including its structure and autoactivation mechanism, its crucial function from onset to progression to metastasis, metabolism, immune escape and even drug resistance in cancer; endogenous regulators; and cancer-related pathways. We also summarize the reported PAK1 small-molecule inhibitors based on their structure types and their potential application in cancer. In addition, we provide overviews on current progress and future challenges of PAK1 in cancer, hoping to provide new ideas for the diagnosis and treatment of cancer.
... Immunohistochemistry was performed as described previously [20]. The immunohistochemical results were judged by the histological score (HSCORE) [21]. ...
MicroRNAs (miRNAs) are known to be associated with certain cancers, including osteosarcoma. We examined osteosarcoma tissues and cell lines, and found that most expressed lower levels of miR-548d-3p than adjacent tissues and normal cell lines. KRAS was identified as a potential target gene of miR-548d-3p. In osteosarcoma cells, miR-548d-3p exerted tumor-suppressive effects by downregulating KRAS. Functional assays revealed that miR-548d-3p mimics dramatically reduced cell growth and migration in vitro. These results suggest that miR-548d-3p mimics could be applied for osteosarcoma treatment.
... The TNM classification was undertaken according to the standard criteria of the 7th TNM staging system. Immunohistochemistry has been described previously, 20 and immunohistochemical results were judged by HSCORE (histological score). 21 ...
MORC2 (microrchidia family CW‐type zinc finger 2) is a newly identified chromatin remodeling protein that functions in diverse biological processes including gene transcription. NDRG1 is a metastasis suppressor and a prognostic biomarker for colorectal cancer (CRC). However, the relationship between MORC2 and NDRG1 transcriptional regulation and the roles of MORC2 in CRC remain elusive. Here, we showed that MORC2 down‐regulated NDRG1 mRNA, protein levels and promoter activity in CRC cells. We also found that MORC2 bound to the –446~–213bp region of the NDRG1 promoter. Mechanistically, histone deacetylase SIRT1 was involved in NDRG1 transcriptional regulation. MORC2 was able to interact with SIRT1 and inhibit NDRG1 promoter activity cumulatively with SIRT1. MORC2 overexpression led to a decreasement of H3Ac and H4Ac of the NDRG1 promoter. Importantly, we demonstrated that NDRG1 was essential in MORC2‐mediated promotion of CRC cell migration and invasion in vitro, as well as lung metastasis of CRC cells in vivo. Moreover, MORC2 expression correlated negatively with NDRG1 expression in CRC patients. High expression of MORC2 was significantly associated with lymph node metastasis (p=0.019) and poor pTNM stage (p=0.02) and the expression of MORC2 correlated with poor prognosis in colon cancer patients. Our findings thus contribute to the knowledge of the regulatory mechanism of MORC2 in down‐regulating NDRG1, and suggest MORC2 as a potential therapeutic target for CRC.
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... Immunohistochemistry. Immunohistochemistry of cancer tissues as well as noncancerous tissues has been performed as described previously 43 and immunohistochemical results were evaluated by HSCORE (histological score). 44 Statistics. ...
The p21-activated kinase 4 (PAK4) is overexpressed in different cancers and promotes proliferation of cancer cells. Reprogramming of glucose metabolism is found in most cancer cells which in turn supports rapid proliferation. However, the relationship between PAK4 and glucose metabolism in cancer cells has not been explored. In this study, we reported that PAK4 promoted glucose intake, NADPH production and lipid biosynthesis, leading to an increased proliferation of colon cancer cells. Mechanistically, PAK4 interacted with glucose-6-phosphate dehydrogenase (G6PD), a rate-limiting enzyme of the pentose phosphate pathway and increased G6PD activity via enhancing Mdm2-mediated p53 ubiquitination degradation. In addition, we demonstrated a close positive correlation between PAK4 and G6PD expression in colon cancer specimens. Furthermore, expression of PAK4 or G6PD was positively correlated with an aggressive phenotype of clinical colon cancer. These findings revealed a novel glucose metabolism-related mechanism of PAK4 in promoting colon cancer cell growth, suggesting that PAK4 and/or G6PD blockage might be a potential therapeutic strategy for colon cancer.
... The NF-B and AP-1 signaling pathway regulates cyclin B1 expression, which is key to the control of G 2 /M transition (61,62). Both the NF-B and AP-1 binding sites have been identi- fied in the cyclin B1 promoter region (62,63). NF-B-dependent transcription is necessary for G 2 /M progression (64). ...
Glioblastoma is the most common malignant brain tumor and has a poor prognosis. Tachykinin receptor neurokinin-1 (NK1R) is a promising target in glioblastoma therapy because of its overexpression in human glioblastoma. NK1R agonists promote glioblastoma cell growth, while NK1R antagonists efficiently inhibit cell growth both in vitro and in vivo. However, the molecular mechanisms involved in these effects are incompletely understood. Beta-arrestin (ARRB) is scaffold protein for signaling transducer. Here, we show that the ARRB1-mediated signaling pathway is essential for NK1-mediated glioblastoma cell proliferation. ARRB1 knockdown significantly inhibited NK1-mediated glioblastoma cell proliferation and induced G2/M phase cell-cycle arrest. ARRB1-knockdown cells showed remarkable downregulation of CDC25C/CDK1/cyclin B1 activity. We also demonstrated that ARRB1 mediated prolonged phosphorylation of ERK1/2 and Akt in glioblastoma cells induced by -NK1R activation. ERK1/2 and Akt phosphorylation were involved in regulating CDC25C/CDK1/cyclin B1 activity. The lack of long-term ERK1/2 and Akt activation in ARRB1-knockdown cells was at least partly responsible for the delayed cell cycle progression and proliferation. Moreover, we found that ARRB1-mediated ERK1/2 and Akt phosphorylation regulated the transcriptional activity of both NF-?B and AP-1, which were involved in cyclin B1 expression. ARRB1 deficiency increased the sensitivity of glioblastoma cells to the treatment of NK1R antagonists. Taken together, our results suggest that ARRB1 plays an essential role in NK1R-mediated cell proliferation and the G2/M transition in glioblastoma cells. Interference with ARRB1-mediated signaling via NK1R may have potential significance for therapeutic strategies targeting glioblastoma.
... In the present study, overexpression of PAK-1 is found to be an independent predictor of disease recurrence. Comparably, Carter [29] and [26][27][28]. ...
Background:
Colorectal carcinoma (CRC) is a significant cause of major morbidity and mortality. PAK-1 is a protein that regulates cytoskeletal dynamics and cell motility. The purpose of the present study is to investigate the relationship between PAK-1 immunoexpression and CRC progression and its validity as an independent prognostic factor.
Patients and methods:
Paraffin blocks of 103 primary CRCs and 37 nodal metastases were retrieved and tissue microarrays were constructed. Immunohistochemistry was performed using anti-PAK-1 antibody. Immunostaining was scored and results were analysed in relation to clinicopathological parameters.
Results:
PAK-1 was overexpressed in primary CRC (P<0.001). No difference between low and high expression in nodal metastasis (P=0.139). There was no difference between PAK-1 immunoexpression in primary and nodal metastasis (P=0.275). High PAK-1 immunoexpression was associated with disease recurrence (P=0.03). However, there was no association with most clinicopathological parameters. PAK-1 overexpression was detected as an independent predictor of disease recurrence (P=0.05). No association was found between PAK-1 immunoexpression and disease free survival (log-rank =1.287, P=0.257).
Conclusion:
PAK-1 overexpression may be involved in CRC progression and could be considered an independent predictor of disease recurrence. Further in vivo and in vitro molecular studies are needed to investigate the role of PAK-1 in colorectal carcinogenesis.
... PAK1 downregulation inhibits the proliferation of gastric cancer cells by decreasing the expression of cyclins D1 and B1 [77,78] . PAK4 also plays essential roles in gastric cancer tumorigenesis and progression. ...
P21-activated kinases (PAKs) are central players in various oncogenic signaling pathways. The six PAK family members are classified into group I (PAK1-3) and group II (PAK4-6). Focus is currently shifting from group I PAKs to group II PAKs. Group II PAKs play important roles in many fundamental cellular processes, some of which have particular significance in the development and progression of cancer. Because of their important functions, group II PAKs have become popular potential drug target candidates. However, few group II PAKs inhibitors have been reported, and most do not exhibit satisfactory kinase selectivity and "drug-like" properties. Isoform- and kinase-selective PAK inhibitors remain to be developed. This review describes the biological activities of group II PAKs, the importance of group II PAKs in the development and progression of gastrointestinal cancer, and smallmolecule inhibitors of group II PAKs for the treatment of cancer.
... Stable overexpressing MORC2 SGC-7901 cell lines were seeded in 60-mm plates to perform the flow cytometry as described in our previous paper [27,29]. For colony formation assay, 500 cells were plated in six-well plates to assess the proliferation potential of cells and incubated at 37°C in a 5% CO 2 incubator. ...
Microrchidia (MORC) family CW-type zinc-finger 2 (MORC2) regulates chromatin remodeling during the DNA-damage response, represses gene transcription, promotes lipogenesis. Here, we found that MORC2 down-regulated p21 by recruiting HDAC1 to the p21 promoter, in a p53-independent manner. MORC2-mediated down-regulation of p21 in turn promoted cell cycle progression in gastric cancer cells. Furthermore, MORC2 expression correlated negatively with p21 expression in gastric tumors in patients. We suggest that MORC2 may be a potential therapeutic target in cancer.
... In the present study, of the 23 lymph node metastasis patients, 18 showed PAK1 overexpression. Liu et al. [38] observed that, higher level of PAK1 in gastric cancer was associated with metastatic tumor. Likewise, overexpression of PAK1 was associated with lymph node metastasis in gastroesophageal junction cancer [19]. ...
Primary esophageal small cell carcinoma (PESCC) is a rare, but fatal subtype of esophageal carcinoma. No effective therapeutic regimen for it. P21-activated kinase 1 (PAK1) is known to function as an integrator and an indispensable node of major growth factor signaling and the molecular therapy targeting PAK1 has been clinical in pipeline. We thus set to examine the expression and clinical impact of PAK1 in PESCC. The expression of PAK1 was detected in a semi-quantitative manner by performing immunohistochemistry. PAK1 was overexpressed in 22 of 34 PESCC tumors, but in only 2 of 18 adjacent non-cancerous tissues. Overexpression of PAK1 was significantly associated with tumor location (p = 0.011), lymph node metastasis (p = 0.026) and patient survival (p = 0.032). We also investigated the association of PAK1 with DNA damage, a driven cause for malignancy progression. γH2AX, a DNA damage marker, was detectable in 18 of 24 (75.0%) cases, and PAK1 expression was associated with γH2AX (p = 0.027). Together, PAK1 is important in metastasis and progression of PESCC. The contribution of PAK1 to clinical outcomes may be involved in its regulating DNA damage pathway. Further studies are worth determining the potentials of PAK1 as prognostic indicator and therapeutic target for PESCC.
... 24,25 For example, elevated PAK1 expression is correlated with cancer progression and lymph node metastases in gastric cancer tissues. 26,27 Therefore, it is worthwhile to study the novel binding partners of PAK1. ...
Actin protrusion at the cell periphery is central to the formation of invadopodia during tumor cell migration and invasion. Although RUFY3 (RUN and FYVE domain containing 3)/SINGAR1 (single axon-related1)/RIPX (Rap2 interacting protein X) has an important role in neuronal development, its pathophysiologic role and relevance to cancer are still largely unknown. The purpose of this study was to elucidate the molecular mechanisms by which RUFY3 involves in gastric cancer cell migration and invasion. Here, our data show that overexpression of RUFY3 leads to the formation of F-actin-enriched protrusive structures at the cell periphery and induces gastric cancer cell migration. Furthermore, P21-activated kinase-1 (PAK1) interacts with RUFY3, and promotes RUFY3 expression and RUFY3-induced gastric cancer cell migration; inhibition of PAK1 attenuates RUFY3-induced SGC-7901 cell migration and invasion. Importantly, we found that the inhibitory effect of cell migration and invasion is significantly enhanced by knockdown of both PAK1 and RUFY3 compared with knockdown of RUFY3 alone or PAK1 alone. Strikingly, we found significant upregulation of RUFY3 in gastric cancer samples with invasive carcinoma at pathologic TNM III and TNM IV stages, compared with their non-tumor counterparts. Moreover, an obvious positive correlation was observed between the protein expression of RUFY3 and PAK1 in 40 pairs of gastric cancer samples. Therefore, these findings provide important evidence that PAK1 can positively regulate RUFY3 expression, which contribute to the metastatic potential of gastric cancer cells, maybe blocking PAK1-RUFY3 signaling would become a potential metastasis therapeutic strategy for gastric cancer.
... Downregulation of PAK1 expression reduced gastric cancer cell migration and invasion [16]. In another study of gastric cancer, Liu et al. found that overexpression of PAK1 was associated with progression, metastasis and prognosis, likely by regulating the transcription of cyclin B1 through nuclear factor-κB (NF-κB) [17]. Overexpression of PAK1 was also found in colorectal cancer, and PAK1 expression was significantly increased in adenomas, invasive carcinomas, and lymph node metastases compared to normal colon [18]. ...
Background
P21-activated protein kinase 1 (PAK1), a main downstream effector of small Rho GTPases, is overexpressed in many malignancies. PAK1 overexpression is associated with poor prognosis in some tumor types, including breast cancer, gastric cancer, and colorectal cancer. However, the expression and clinical relevance of PAK1 expression in human pancreatic cancer remains unknown.
Methods
The present study investigated the clinical and prognostic significance of PAK1 expression in pancreatic carcinoma. We examined and scored the expression of PAK1 by immunohistochemistry in 72 primary pancreatic carcinoma samples and 20 liver metastatic samples. The relationships between PAK1 and clinicopathological parameters and prognosis in primary and metastatic pancreatic cancer were analyzed.
Results
Among the total 92 cases, primary pancreatic cancer samples had a significantly higher rate (38/72, 52.8%) of high PAK1 expression than liver metastatic samples (5/20, 25.0%) (P = 0.028). Among the 72 primary pancreatic cancer patients, high PAK1 expression was associated with younger age (P = 0.038) and moderately or well differentiated tumor (P = 0.007). Moreover, a positive relationship was found between high PAK1 expression and overall survival (OS) (P < 0.005). Patients with high PAK1 expression had a better OS than those with low PAK1 expression. Univariate and multivariate analysis by Cox regression including PAK1 and other prognostic pathological markers demonstrated high PAK1 immunostaining as a prognostic factor for survival in pancreatic cancer patients (P < 0.005).
Conclusions
We report for the first time that PAK1 is a novel prognostic marker for pathologically confirmed human pancreatic cancer. Reduced expression of PAK1 correlates with poor histological differentiation in pancreatic cancer.
... PKN1, also known as PAK1, is a serine/threonine kinase implicated in formation of mammary gland tumors and premalignant lesions in animal models, although with a long latency. 1 It has been reported that overexpression of PKN1 correlates with aggressive ovarian, 2 colorectal, 3 and prostate cancers. 4 Other findings suggested that PKN1 has a role in the development of invasive phenotypes of breast 5 and gastric 6 cancer cells. PKN1 contains three highly conserved regions: 1) a unique regulatory domain in the amino terminus; 2) a catalytic domain homologous to PKC in carboxyl terminal region; and 3) a so-called D-region located between the regulatory and catalytic domains. ...
Background
The response of cells to TGFβ and EGF is mediated by a network of various intracellular regulators. The signaling crosstalk between different regulators is of key importance for tumorigenesis. The crosstalk may explain the modulation of cellular responses to the same regulator by another signaling molecule. As PKN1 – a serine/threonine kinase implicated in tumorigenesis – was identified as potential crosstalk node for TGFβ and EGF signaling, the cellular functions that may be affected by PKN1 in a crosstalk of TGFβ and EGF were explored.
Methods
To investigate the contribution of PKN1 to TGFβ and EGF signaling, transiently PKN1-transfected HEC-1-A endometrial cancer cells were generated and subjected to treatment with TGFβ1, EGF, and their combination. Proliferation, apoptosis, invasion, wound healing, and migration assays were performed. The impact of PKN1 on the expression and phosphorylation of intracellular proteins was monitored by immunoblotting.
Results
It was demonstrated that PKN1 modulated the responses of HEC-A-1 endometrial cancer cells to TGFβ1 and EGF. PKN1 had an inhibitory effect on the stimulation of cell migration, and PKN1 kinase activity was required for the inhibitory effect of TGFβ and EGF on cell proliferation and invasiveness. It was observed that phosphorylation of Smad2, FAK, and Erk1/2 correlated with responses of the cells to TGFβ1 and EGF.
Conclusion
PKN1 modulates TGFβ- and EGF-dependent regulation of cell proliferation, migration, and invasiveness, and therefore is a component of the network signaling downstream of TGFβ and EGF.
... It has been reported that overexpression of PKN1 correlated with aggressive ovarian (Galgano et al, 2009), colorectal (Carter et al, 2004), and prostate cancers (Metzger et al, 2003). Other findings suggested that PKN1 has a role in the development of invasive phenotypes of breast (Adam et al, 2000) and gastric (Liu et al, 2009) cancer cells. Despite the recent advances in knowledge regarding the role of PKN1 and MST1 in tumorigenesis, the involvement of these proteins in the development and progression of endometrial cancer has been as of yet unexplored. ...
Background:
The individual features of tumours are often disregarded in cohort studies. As these features may represent a source for individualised cancer treatment, it is important to develop a novel approach for their assessment.
Methods:
We used proteomics, systems biology, and immunohistochemistry to explore protein expression in human endometrial tumours, to identify deregulated regulatory mechanisms, and to validate observed changes in protein expression using tissue microarrays.
Results:
Compared with the evaluation of common tumour features, the evaluation of individual tumour features gave a more comprehensive and detailed overview of the regulatory processes in endometrial tumours. Systemic analysis of the individual proteome profiles showed that endometrial tumours employed different proteins to regulate similar functions. Comparison of our data with publicly available data sets of molecular profiling of human endometrial tumours confirmed that individual tumour features are not simply irrelevant individual variations, but are indeed important in endometrial tumorigenesis. Validation through tissue microarray investigation of MST1 and PKN1 proteins confirmed the usefulness of this approach, and suggested that MST1 and PKN1 may be considered as predictive biomarkers of endometrial cancer.
Conclusion:
We show that individualised profiling of endometrial tumours may deliver better insights into a tumour's physiology, thereby giving a better prediction of tumour development. Individual tumour features may also be used to tailor cancer treatment.
... In breast cancer, nuclear expression of PAK1, in conjunction with phosphorylation of the estrogen receptor on the PAK1 site (serine 305), predicts resistance to tamoxifen therapy, and the cytoplasmic levels of PAK1 correlate with the recurrence rate and mortality [35,37]. Similarly, higher levels of PAK1 were associated with advanced tumor stage, metastasis and reduced survival in patients with gastric cancer [38,39]. There are also numerous reports of elevated PAK1 activity in cell lines, although in most of these cases such reports cannot rule out the possibility that the changes have been selected for or caused by in vitro culture. ...
Histone modification enzymes regulate gene expression by altering the accessibility of promoters to transcription factors. We sought to determine whether the genes encoding histone modification enzymes are dysregulated in pediatric acute lymphoblastic leukemia (ALL). A real-time PCR array was designed, tested and used to profile the expression of 85 genes encoding histone modification enzymes in bone marrow mononuclear cells from 30 pediatric ALL patients and 20 normal controls. The expression profile of histone-modifying genes was significantly different between normal karyotype B cell pediatric ALL and normal controls. Eleven genes were upregulated in pediatric ALL, including the histone deacetylases HDAC2 and PAK1, and seven genes were downregulated, including PRMT2 and the putative tumor suppressor EP300. Future studies will seek to determine whether these genes serve as biomarkers of pediatric ALL. Ingenuity Pathway Analysis revealed that Gene Expression and Organ Morphology was the highest rated network, with 13 focus molecules (significance score = 35). Ingenuity Pathway Analysis also indicated that curcumin and miR-34 are upstream regulators of histone-modifying enzymes; future studies will seek to validate these results and examine the role of curcumin and miR-34 in leukemia. This study provides new clues into the molecular mechanisms of pediatric ALL.
... Knockdown of PAK4 reduced the proliferation and survival of ovarian cancer cells (20), pancreatic cancer 8988T cells (18), JEG3, and JAR choriocarcinoma cells (33) and increased the levels of apoptotic proteins in gastric cancer cell lines (34). In comparison, knockdown of PAK1 inhibited the anchorage-dependent (14) and -independent (35) growth of breast cancer cells, the proliferation of NSCLC cancers in vitro and in vivo (14), colon cancer in vitro (27) and in vivo (28), and the growth of gastric cancer cells anchorage-dependently and -independently as well as tumor xenografts (36). In previous studies, gene amplification or increased protein expression of PAK4 or PAK1 has been suggested to correlate with dependence on this pathway for tumor progression and tumor cell proliferation. ...
The p21-activated kinase (PAK) serine / threonine kinases are important effectors of the small GTPases Rac and Cdc42, and play significant roles in controlling cell growth, motility and transformation. Knockdown of PAK4 or PAK1 inhibited the proliferation of mutant KRAS or BRAF colon cancer cells in vitro. Dependence on PAK4 or PAK1 protein for colon cancer cell proliferation was independent of PAK4 or PAK1 protein expression levels. Mutant KRAS HCT116 colorectal cells were the most sensitive to PAK4 or PAK1 knockdown resulting in the potent inhibition of anchorage-dependent and -independent proliferation as well as the formation and proliferation of HCT116 colon cancer spheroids. This inhibition of proliferation did not correlate with inhibition of RAF/MEK/ERK or PI3K/AKT signaling. In HCT116 cells, knockdown of PAK4 or PAK1 caused changes to the actin cytoskeleton resulting in reduced basal spread and cell elongation and increased cell rounding. These cytoskeletal rearrangements appeared to be independent of LIMK/cofilin/paxillin phosphorylation. PAK4 or PAK1 knockdown initially induced growth arrest in HCT116 cells followed by cell death at later time points. Inhibition of the anti-apoptotic proteins Bcl-2 and Bcl-XL with the pharmacological inhibitor ABT-737 increased effector caspase activation and apoptosis, and reduced cell survival with PAK4 or PAK1 knockdown. These results support a role for the p21-activated kinases in the proliferation of mutant KRAS driven colorectal carcinoma cells via pathways not involving RAF/MEK/ERK and PI3K/AKT signaling.
... Both PAK1 and PAK4 are overexpressed in gastric cancer and play important roles in its metastasis [80][81][82]. In human gastric cancer cell lines, PAK1 regulates the expression and activity of cyclins D1 and B1 via NF-κB, and inhibition of PAK1 suppresses proliferation and migration by decreasing the expression of cyclins D1 and B1 [83,84]. In gastric cancer patients, overexpression of PAK1 is associated with advanced and metastatic tumour stages, and increased PAK1 activity is related to reduced survival [81]. ...
p21-activated kinases (PAKs) were initially identified as effector proteins downstream from GTPases of the Rho family. To date, six members of the PAK family have been discovered in mammalian cells. PAKs play important roles in growth factor signalling, cytoskeletal remodelling, gene transcription, cell proliferation and oncogenic transformation. A large body of research has demonstrated that PAKs are up-regulated in several human cancers, and that their over-expression is linked to tumour progression and resistance to therapy. Structural and biochemical studies have revealed the mechanisms involved in PAK signalling, and opened the way to the development of PAK-targeted therapies for cancer treatment. Here we summarize recent findings from biological and clinical research on the role of PAKs in gastrointestinal cancer, and discuss the current status of PAK-targeted anticancer therapies.
... Although it has been reported that Pak1 overexpression might induce gastric cancer cell proliferation by regulating cyclin B1 expression (19), the studies about the role of Pak1 in invasion and metastasis of gastric cancer were still absent. In our study, both Pak1 overexpression and activity in gastric cancer were observed to significantly correlate with more aggressive tumor behavior in terms of depth of invasion, lymph node metastasis, distant metastasis, tumor stages, tumor grades, and shorter disease-free survival. ...
P21-activated protein kinase (Pak1), a main downstream effector of small Rho GTPases, plays an important role in the regulation of cell morphogenesis, motility, mitosis and angiogenesis. However, the role of Pak1 in gastric cancer metastasis remains unclear. Here, we showed that Pak1 is overexpressed in gastric cancer tissues from 74 patients by immunohistochemistry. Overexpression of Pak1 was associated with metastasis and prognosis of gastric cancer. In addition, overexpression of Pak1 increased gastric cancer cell motility and invasion, whereas downregulation of Pak1 expression reduced gastric cancer cell migration and invasion. In further study, data showed that activated Pak1 inhibited stress fiber and focal adhesion complex formation in gastric cancer cells and led to the formation of motile phenotypes. Importantly, activated Pak1 elicited phosphorylation of the ERK and JNK-dependent pathway in gastric cancer cell lines. In conclusion, our results suggest that Pak1 is overexpressed in gastric cancer and plays an important role in the metastasis of gastric cancer. The mechanism by which Pak1 induces cancer metastasis may involve activation of ERK and JNK.
... PAK, a mammalian homologs of Ser/Thr protein kinase, is the effector of bradykinin peptide receptors pathway, which has been implicated in a variety of intracellular signaling events, including cell cytoskeleton rearrangement, proliferation, cell-cycle progression, cell migration and angiogenesis (Tang et al., 2000;Fryer and Field, 2005;Galan Moya et al., 2009). Overexpression and activation of PAK has been identified in the development of cancers (Kumar and Vadlamudi, 2002;Liu et al., 2009). Recently, targeting PAKs have been proposed as a potential therapeutic strategy for gastric cancer Li et al., 2010). ...
The overexpression of IRX1 gene correlates with the growth arrest in gastric cancer. Furthermore, overexpression of IRX1 gene suppresses peritoneal spreading and long distance metastasis. To explore the precise mechanisms, we investigated whether restoring IRX1 expression affects the angiogenesis or vasculogenic mimicry (VM). Human umbilical vein endothelial cells (HUVECs) and chick embryo and SGC-7901 gastric cancer cells were used for angiogenesis and VM analysis. Small interfering RNA was used for analyzing the function of BDKRB2, a downstream target gene of IRX1. As results, the remarkable suppression on peritoneal spreading and pulmonary metastasis of SGC-7901 cells by IRX1 transfectant correlates to reduced angiogenesis as well as VM formation. Using the supernatant from SGC-7901/IRX1 cells, we found a strong inhibiting effect on angiogenesis both in vitro and in chick embryo. SGC-7901/IRX1 cells revealed strong inhibiting effect on VM formation too. By gene-specific RNA interference for BDKRB2, or its effector PAK1, we got an effective inhibition on tube formation, cell proliferation, cell migration and invasion in vitro. In conclusion, enforcing IRX1 expression effectively suppresses peritoneal spreading and pulmonary metastasis via anti-angiogenesis and anti-VM mechanisms, in addition to previously found cell growth and invasion. BDKRB2 and its downstream effector might be potential targets for anti-cancer strategy.
... Based on our data, we speculated that ECRG1 and ECRG4 might co-regulate p21 expression to control cell cycle progression in ESCC. It is well known that p21 upregulation is able to block the cell cycle at G1 phase [29,30]. So the p21 overexpression induced by ECRG4 and ECRG1 co-expression could be the possible molecular mechanism for cell cycle G1 phase block and growth suppression in ESCC. ...
The esophageal carcinoma related gene 4 (ECRG4) was initially identified and cloned from human normal esophageal epithelium in our laboratory (GenBank accession no.AF325503). ECRG4 has been described as a novel tumor suppressor gene associated with prognosis in esophageal squamous cell carcinoma (ESCC).
In this study, binding affinity assay in vitro and co-immunoprecipitation experiment in vivo were utilized to verify the physical interaction between ECRG4 and transmembrane protease, serine 11A (TMPRSS11A, also known as ECRG1, GenBank accession no. AF 071882). Then, p21 protein expression, cell cycle and cell proliferation regulations were examined after ECRG4 and ECRG1 co-transfection in ESCC cells.
We revealed for the first time that ECRG4 interacted directly with ECRG1 to inhibit cancer cell proliferation and induce cell cycle G1 phase block in ESCC. Binding affinity and co-immunoprecipitation assays demonstrated that ECRG4 interacted directly with ECRG1 in ESCC cells. Furthermore, the ECRG4 and ECRG1 co-expression remarkably upregulatd p21 protein level by Western blot (P < 0.001), induced cell cycle G1 phase block by flow cytometric analysis (P < 0.001) and suppressed cell proliferation by MTT and BrdU assay (both P < 0.01) in ESCC cells.
ECRG4 interacts directly with ECRG1 to upregulate p21 protein expression, induce cell cycle G1 phase block and inhibit cancer cells proliferation in ESCC.
... In the present research, we discovered for the first time that ECRG4 inhibited cancer cells proliferation and induced cell cycle G1 phase block by up-regulating p21 expression level through p53 mediated pathway in ESCC. It is well known that p21, the critical cyclin-dependent kinase inhibitor, is able to block the cell cycle at G1 phase [25,26]. So the p21 expression upregulation could be the molecular mechanism for the ECRG4-induced cell cycle G1 phase block in ESCC. ...
The esophageal cancer related gene 4 (ECRG4) was initially identified and cloned in our laboratory from human normal esophageal epithelium (GenBank accession no.AF325503). ECRG4 was a new tumor suppressor gene in esophageal squamous cell carcinoma (ESCC) associated with prognosis. In this study, we investigated the novel tumor-suppressing function of ECRG4 in cancer cell migration, invasion, adhesion and cell cycle regulation in ESCC.
Transwell and Boyden chamber experiments were utilized to examined the effects of ECRG4 expression on ESCC cells migration, invasion and adhesion. And flow cytometric analysis was used to observe the impact of ECRG4 expression on cell cycle regulation. Finally, the expression levels of cell cycle regulating proteins p53 and p21 in human ESCC cells transfected with ECRG4 gene were evaluated by Western blotting.
The restoration of ECRG4 expression in ESCC cells inhibited cancer cells migration and invasion (P < 0.05), which did not affect cell adhesion capacity (P > 0.05). Furthermore, ECRG4 could cause cell cycle G1 phase arrest in ESCC (P < 0.05), through inducing the increased expression of p53 and p21 proteins.
ECRG4 is a candidate tumor suppressor gene which suppressed tumor cells migration and invasion without affecting cell adhesion ability in ESCC. Furthermore, ECRG4 might cause cell cycle G1 phase block possibly through inducing the increased expression of p53 and p21 proteins in ESCC.
Microrchidia CW-type zinc finger 2 (MORC2) is a member of the MORC superfamily of nuclear proteins. Growing evidence has shown that MORC2 not only participates in gene transcription and chromatin remodeling but also plays a key in human disease and tumor development by regulating the expression of downstream oncogenes or tumor suppressors. The present review provides an updated overview of MORC2 in the aspect of cancer hallmark and therapeutic resistance and summarizes its upstream regulators and downstream target genes. This systematic review may provide a favorable theoretical basis for emerging players of MORC2 in tumor development and new insight into the potential clinical application of basic science discoveries in the future.
Background
p21-activated kinase 1 (PAK1) plays a fundamental role in promoting the development and progression of several cancers and is a potential therapeutic target. However, the biological function and underlying mechanism of PAK1 in esophageal squamous cell carcinoma (ESCC) remain unclear.
Methods
The expression of PAK1 was detected in both ESCC cell lines and clinical samples. Cell growth was measured by MTT, focus formation and soft agar assays. Cell migration and invasion were detected by wound healing and transwell assays. Animal models of subcutaneous tumourigenicity and tail vein metastasis were performed to determine the inhibitory effect of pharmacological inhibitor IPA-3 on tumor growth and metastasis of ESCC cells.
Results
We found that PAK1 was frequently overexpressed in ESCC. Ectopic expression of PAK1 promoted cellular growth, colony formation and anchorage-independent growth. Overexpressing PAK1 also enhanced migration, invasion and the expression of MMP-2 and MMP-9 in ESCC cells. In contrast, silencing PAK1 by lentiviral knockdown or a specific inhibitor IPA-3 resulted in a contrary effect. Subsequent investigations revealed that Raf1/MEK1/ERK signaling pathway was involved in PAK1-mediated effect. Enhanced expression of Raf1 attenuated the inhibitory functions of PAK1 shRNA. Whereas blocking of Raf1 by shRNA or specific inhibition of MEK1 by U0126 antagonized the oncogenetic effect of PAK1 on ESCC cells. More importantly, Pharmacological inhibition of PAK1 by IPA-3 significantly suppressed tumor growth and lung metastasis of ESCC cells in vivo.
Conclusions
These data support that PAK1 is an ideal target for the development of potential therapeutic drugs for ESCC patients even with metastasis.
Electronic supplementary material
The online version of this article (10.1186/s12964-019-0343-5) contains supplementary material, which is available to authorized users.
Background:
Human T-cell leukemia virus type 1 (HTLV-1) causes adult T-cell leukemia (ATL). Viral Tax protein plays a major role in ATL development. Pim family of serine/threonine kinases is composed of Pim-1, -2, and -3. The potential of Pim family as a target in ATL was analyzed.
Methods:
RT-PCR and Western blotting were used to determine the expression of Pim kinases, Tax, and intracellular signal molecules. Knockdown of Pim-3 and RelA was performed using small interfering RNA. The effects on cell proliferation, viability, cell cycle, and apoptosis were analyzed by WST-8, propidium iodide, and APO2.7 assay. NF-κB DNA binding activity was investigated by electrophoretic mobility shift assay.
Results:
Pim-3 expression was restricted to HTLV-1-infected T-cell lines. Tax induced Pim-3 expression through NF-κB. Knockdown of Pim-3 showed growth inhibition of HTLV-1-infected T cells. NJC97-NH, a novel inhibitor of the Pim-1/3 kinases, inhibited cell viability. NJC97-NH induced G2/M cell cycle arrest associated with downregulation of cyclin A and cyclin B1 expression, as well as apoptosis accompanied with downregulation of XIAP and Mcl-1 expression through inhibition of NF-κB pathway, mediated through decrease in IκBα and RelA phosphorylation.
Conclusion:
Pim-3 is a potentially suitable target for the development of novel therapeutic agents against ATL. This article is protected by copyright. All rights reserved.
Since the initial recognition of a mechanistic role of p21-activated kinase 1 (PAK1) in breast cancer invasion, PAK1 has emerged as one of the widely overexpressed or hyperactivated kinases in human cancer at-large, allowing the PAK family to make in-roads in cancer biology, tumorigenesis, and cancer therapeutics. Much of our current understanding of the PAK family in cancer progression relates to a central role of the PAK family in the integration of cancer-promoting signals from cell membrane receptors as well as function as a key nexus-modifier of complex, cytoplasmic signaling network. Another core aspect of PAK signaling that highlights its importance in cancer progression is through PAK's central role in the cross talk with signaling and interacting proteins, as well as PAK's position as a key player in the phosphorylation of effector substrates to engage downstream components that ultimately leads to the development cancerous phenotypes. Here we provide a comprehensive review of the recent advances in PAK cancer research and its downstream substrates in the context of invasion, nuclear signaling and localization, gene expression, and DNA damage response. We discuss how a deeper understanding of PAK1's pathobiology over the years has widened research interest to the PAK family and human cancer, and positioning the PAK family as a promising cancer therapeutic target either alone or in combination with other therapies. With many landmark findings and leaps in the progress of PAK cancer research since the infancy of this field nearly 20 years ago, we also discuss postulated advances in the coming decade as the PAK family continues to shape the future of oncobiology.
Objective: To investigate the expression and clinical significance of Iroquois homebox gene IRX1 in human hepatocellular carcinoma (HCC) cells. Methods: A total of 82 hepatocellular carcinoma tissue samples were collected, of which 26 were highly differentiated tissues at Edmondson stage I , 25 were moderately differentiated tissues at Edmondson stages I - II and II , 31 were lowly differentiated tissues at Edmondson stage III , and determined for expression of IRX1 by real-time fluorescent quantitative PCR (Q-PCR) and Western blot, and for location of IRX1 and clinical character by immunohistochemistry, using 11 normal or non-tumor tissue samples as control. Results: Both the expression levels of IRX1 gene and protein were higher in hepatocellular carcinoma tissue than in control tissue (P < 0.05), which were related to the differentiation level of carcinoma. The lower the differentiation level of hepatocellular carcinoma tissue was, the higher the expression level of IRX1 was (P < 0.05). The positive rate of IRX1 in hepatocellular carcinoma tissue was significantly higher than that in non-tumor control tissues, which increased with the increased stage and decreased differentiation level of carcinoma, while showed no significant relationship to HBV infection or accompanied liver cirrhosis (P > 0.05). Of the highly differentiated carcinoma tissues positive for IRX1, 68.2% were positive in nuclei, 18.2% in cytoplasm, and 13.6% in both nuclei and cytoplasm. However, of the lowly differentiated carcinoma tissues positive for IRX1, 65.5% were positive in both nuclei and cytoplasm. Conclusion: IRX1 might involve in the regulation of onset and progress of HCC, of which the expression was related to the differentiation of carcinoma, indicating that IRX1 might be used as an indicator for evaluation of prognosis and differentiation level as well as targeted molecular dierapy of HCC.
LMO1, a nuclear transcription coregulator, is implicated in the pathogenesis of T-cell acute lymphoblastic leukemia and neuroblastoma. However, the role of LMO1 in human prostate cancer (PCa) is still unknown. Androgen receptor (AR) plays a critical role in the progression of prostate cancer. The activation of AR signaling pathway could be modulated by AR cofactors. In the present study, we discovered that LMO1 could bind to AR and co-localize with AR in the nucleus. In addition, the expression of LMO1 in human PCa tissues was significantly higher than that in benign prostate hyperplasia (BPH) tissues. Moreover, LMO1 appeared to be a novel coactivator to enhance AR transcriptional activities, followed by the elevation of expression of P21 and PSA, downstream targets of AR. Taken together, LMO1 appears to be a coactivator of AR involved in the progression of prostate cancer, and could be a promising molecular target for treating prostate cancer.
p21-activated kinases (PAKs) are a family of serine/threonine protein kinases comprised of six isoform (PAK1-6), all of which are direct targets of the small GTPases Rac and Cdc42. PAKs have recently been shown to regulate various cellular activities, including cell motility, survival and proliferation, the organization and function of cytoskeleton and extracellular matrix, transcription and translation. PAKs are overexpressed or hyperactivated in several human tumor, such as breast cancer, gastric cancer, ovarian cancer etc., which makes them an attractive new therapeutic targets. Thus, there has been considerable interest in the development of inhibitors to the PAKs, as biological markers and leads for the development of therapeutics.
Cell division cycle 42 (CDC42), an important member of the Ras homolog (Rho) family, plays key role in regulating multiple cellular processes such as cell cycle progression, migration, cell cytoskeleton organization, cell fate determination and differentiation. Among the downstream effectors of CDC42, P21-activated kinases (PAKs) obtains the most attention. Although a large body of evidence indicate that CDC42/PAKs pathway plays important role in tumor growth, invasion and metastasis, the mechanism of their negative regulation remains unclear. Here, we identified CDC42, a PAKs activating factor, was a target of miR-133. Ectopic overexpression of miRNAs not only downregulated CDC42 expression and PAKs activation, but also inhibited cancer cell proliferation and migration. We also found that miR-133 was down-regulated in 180 pairs gastric cancer tissues. miR-133 expression was negatively associated with tumor size, invasion depth and peripheral organ metastasis. Besides, dysfunction of miR-133 was an independent prognosis factor for overall survival. Our findings could provide new insights into the molecuar mechanisms of gastric carcinogenesis, and may help facilitating development of CDC42/PAK-based therapies for human cancer.
The alteration of p21-activated kinase 4 (PAK4) and transforming growth factor-beta (TGF-β) signaling effector Smad2/3 was detected in several types of tumors, which acts as oncogenic factor and tumor suppressor, but the relationship between these events has not been explored. Here, we demonstrate that PAK4 interacts with and modulates phosphorylation of Smad2/3 via both kinase-dependent and kinase-independent mechanisms, which attenuate Smad2/3 axis transactivation and TGF-β-mediated growth inhibition in gastric cancer cells. First, PAK4 interaction with Smad2/3, which is independent of PAK4 kinase activity, blocks TGF-β1-induced phosphorylation of Smad2 Ser465/467 or Smad3 Ser423/425 and the consequent activation. In addition, PAK4 phosphorylates Smad2 on Ser465, leading to the degradation of Smad2 through ubiquitin-proteasome-dependent pathway under hepatocyte growth factor (HGF) stimulation. Interestingly, PAK4 expression correlates negatively with phospho-Ser465/467 Smad2 but positively with phospho-Ser465 Smad2 in gastric cancer tissues. Furthermore, the expressions of HGF, phospho-Ser474 PAK4 and phospho-Ser465 Smad2 are markedly increased in gastric cancer tissues, and the expression of Smad2 is decreased in gastric cancer tissues. Our results document an oncogenic role of PAK4 in repression of Smad2/3 transactivation that involved in tumorigenesis, and suggest PAK4 as a potential therapeutic target for gastric cancer.Oncogene advance online publication, 12 August 2013; doi:10.1038/onc.2013.300.
Sparstolonin B (SsnB) is a novel bioactive compound isolated from Sparganium stoloniferum, an herb historically used in Traditional Chinese Medicine as an anti-tumor agent. Angiogenesis, the process of new capillary formation from existing blood vessels, is dysregulated in many pathological disorders, including diabetic retinopathy, tumor growth, and atherosclerosis. In functional assays, SsnB inhibited endothelial cell tube formation (Matrigel method) and cell migration (Transwell method) in a dose-dependent manner. Microarray experiments with human umbilical vein endothelial cells (HUVECs) and human coronary artery endothelial cells (HCAECs) demonstrated differential expression of several hundred genes in response to SsnB exposure (916 and 356 genes, respectively, with fold change 2, p < 0.05, unpaired t-test). Microarray data from both cell types showed significant overlap, including genes associated with cell proliferation and cell cycle. Flow cytometric cell cycle analysis of HUVECs treated with SsnB showed an increase of cells in the G1 phase and a decrease of cells in the S phase. Cyclin E2 (CCNE2) and Cell division cycle 6 (CDC6) are regulatory proteins that control cell cycle progression through the G1/S checkpoint. Both CCNE2 and CDC6 were downregulated in the microarray data. Real Time quantitative PCR confirmed that gene expression of CCNE2 and CDC6 in HUVECs was downregulated after SsnB exposure, to 64% and 35% of controls, respectively. The data suggest that SsnB may exert its anti-angiogenic properties in part by downregulating CCNE2 and CDC6, halting progression through the G1/S checkpoint. In the chick chorioallantoic membrane (CAM) assay, SsnB caused significant reduction in capillary length and branching number relative to the vehicle control group. Overall, SsnB caused a significant reduction in angiogenesis (ANOVA, p < 0.05), demonstrating its ex vivo efficacy.
2-Arylamino-4-aryl-pyrimidines were found to be potent inhibitors of PAK1 kinase. The synthesis and SAR are described. The incorporation of a bromide at the 5-position of the pyrimidine core and in combination with a 1,2-dimethylpiperazine pendant domain yielded a lead compound with potent PAK1 inhibition and anti-proliferative activity in various colon cancer cell lines.
Amplification at 11q13-q14 is a common event in cancers from multiple anatomical sites. This complex amplicon has multiple cores and several genes have been put forward as potential "drivers." In this review, based on the technical advancements of the last decade, which resulted in methods allowing for a deeper genomic and functional genomic characterization of amplicons and their drivers, we discuss the current definitions of amplicons and driver genes, the clinical and biological significance of the 11q13-q14 amplicon in distinct types of human cancer, its coamplification partners, and the roles of various genes located within the amplicon as potential "drivers." Finally, we appraise the available data for novel therapies targeting genes mapping to this amplicon. © 2012 Wiley Periodicals, Inc.
The p21-activated kinases (PAKs) are one of the first direct kinase targets of Ras-related small GTPases to be discovered and have emerged as central players in growth factor signaling networks that regulate morphogenetic processes. In some situations, PAKs control cell proliferation, but their wider role involves establishing cell polarity and promoting cellular plasticity via changes in the actin cytoskeleton. PAKs have been shown to impact on three important areas of human health, namely, cancer, brain function, and virus infection. We review the mechanisms and targets of PAKs in these contexts and provide an overview of the ways in which inhibitors might act to arrest tumor growth, combat virus infection, and promote cell apoptosis. Although PAKs are most abundant in the brain, there are few details of how they might be operating in this context. The advent of new and more selective PAK inhibitors promises new avenues of treatment and allows us to probe in greater detail the importance of PAK biology.
P21-activated protein kinase1 (PAK1), a main downstream effector of small Rho GTPases, Rac1, and Cdc42, plays an important role in the regulation of cell morphogenesis, motility, mitosis, and angiogenesis. Despite its importance, the molecular mechanisms of PAK1 that contributed to colorectal carcinogenesis remain unclear. Our immunohistochemistry showed that PAK1 expression was increased with colorectal cancer (CRC) progression through the adenoma to carcinoma sequence. Furthermore, our results suggested a relationship between PAK1 nuclear localization and the Dukes staging. In the present study, we showed that PAK1 knockdown decreased proliferation and delayed the G1/S cell-cycle transition, and increased apoptosis in vivo and in vitro. In addition, PAK1 knock-down downregulated c-Jun amino terminal kinases (JNK) activity and the levels of cyclinD1, CDK4/6. Inhibition of the JNK activity by chemical inhibitor (SP600125) significantly reduced the effects of PAK1 on CRC proliferation via accumulation of phosphatase and tensin homolog deleted on chromosome 10 (PTEN). In conclusion, our results demonstrate that knockdown of PAK1 could enhance the chemosensitivity of CRCs to 5-fluorouracil through G1 arrest. The mechanism by which PAK1 induced cancer growth might involve activation of JNK as well as downregulation of PTEN. Targeting PAK1 may represent a novel treatment strategy for developing novel chemotherapeutic agents.
The incidence and mortality of gastric cancer have fallen dramatically in US and elsewhere over the past several decades. Nonetheless, gastric cancer remains a major public health issue as the fourth most common cancer and the second leading cause of cancer death worldwide. Demographic trends differ by tumor location and histology. While there has been a marked decline in distal, intestinal type gastric cancers, the incidence of proximal, diffuse type adenocarcinomas of the gastric cardia has been increasing, particularly in the Western countries. Incidence by tumor sub-site also varies widely based on geographic location, race, and socioeconomic status. Distal gastric cancer predominates in developing countries, among blacks, and in lower socioeconomic groups, whereas proximal tumors are more common in developed countries, among whites, and in higher socio-economic classes. Diverging trends in the incidence of gastric cancer by tumor location suggest that they may represent two diseases with different etiologies. The main risk factors for distal gastric cancer include Helicobacter pylori (H pylori) infection and dietary factors, whereas gastroesophageal reflux disease and obesity play important roles in the development of proximal stomach cancer. The purpose of this review is to examine the epidemiology and risk factors of gastric cancer, and to discuss strategies for primary prevention.
In this study, we show that phosphorylated 3-phosphoinositide-dependent kinase 1 (PDK1) phosphorylates p21-activated kinase
1 (PAK1) in the presence of sphingosine. We identify threonine 423, a conserved threonine in the activation loop of kinase
subdomain VIII, as the PDK1 phosphorylation site on PAK1. Threonine 423 is a previously identified PAK1 autophosphorylation
site that lies within a PAK consensus phosphorylation sequence. After pretreatment with phosphatases, autophosphorylation
of PAK1 occurred at all major sites except threonine 423. A phosphothreonine 423-specific antibody detected phosphorylation
of recombinant, catalytically inactive PAK1 after incubation with wild-type PAK1, indicating phosphorylation of threonine
423 occurs by an intermolecular mechanism. The biological significance of PDK1 phosphorylation of PAK1 at threonine 423 in vitro is supported by the observation that these two proteins interact in vivo and that PDK1-phosphorylated PAK1 has an increased activity toward substrate. An increase of phosphorylation of catalytically
inactive PAK1 was observed in COS-7 cells expressing wild-type, but not catalytically inactive, PDK1 upon elevation of intracellular
sphingosine levels. PDK1 phosphorylation of PAK1 was not blocked by pretreatment with wortmannin or when PDK1 was mutated
to prevent phosphatidylinositol binding, indicating this process is independent of phosphatidylinositol 3-kinase activity.
The data presented here provide evidence for a novel mechanism for PAK1 regulation and activation.
Bad is a critical regulatory component of the intrinsic cell death machinery that exerts its death-promoting effect upon heterodimerization
with the antiapoptotic proteins Bcl-2 and Bcl-xL. Growth factors promote cell survival through phosphorylation of Bad, resulting in its dissociation from Bcl-2 and Bcl-xL and its association with 14-3-3τ. Survival of interleukin 3 (IL-3)-dependent FL5.12 lymphoid progenitor cells is attenuated
upon treatment with the Rho GTPase-inactivating toxin B fromClostridium difficile. p21-activated kinase 1 (PAK1) is activated by IL-3 in FL5.12 cells, and this activation is reduced by the phosphatidylinositol
3-kinase inhibitor LY294002. Overexpression of a constitutively active PAK mutant (PAK1-T423E) promoted cell survival of FL5.12
and NIH 3T3 cells, while overexpression of the autoinhibitory domain of PAK (amino acids 83 to 149) enhanced apoptosis. PAK
phosphorylates Bad in vitro and in vivo on Ser112 and Ser136, resulting in a markedly reduced interaction between Bad and
Bcl-2 or Bcl-xL and the increased association of Bad with 14-3-3τ. Our findings indicate that PAK inhibits the proapoptotic effects of Bad
by direct phosphorylation and that PAK may play an important role in cell survival pathways.
We have examined the consequences of overexpression of the IkappaBalpha and IkappaBbeta inhibitory proteins on the regulation of NF-kappaB-dependent beta interferon (IFN-beta) gene transcription in human cells after Sendai virus infection. In transient coexpression studies or in cell lines engineered to express different forms of IkappaB under tetracycline-inducible control, the IFN-beta promoter (-281 to +19) linked to the chloramphenicol acetyltransferase reporter gene was differentially inhibited in response to virus infection. IkappaBalpha exhibited a strong inhibitory effect on virus-induced IFN-beta expression, whereas IkappaBbeta exerted an inhibitory effect only at a high concentration. Despite activation of the IkappaB kinase complex by Sendai virus infection, overexpression of the double-point-mutated (S32A/S36A) dominant repressors of IkappaBalpha (TD-IkappaBalpha) completely blocked IFN-beta gene activation by Sendai virus. Endogenous IFN-beta RNA production was also inhibited in Tet-inducible TD-IkappaBalpha-expressing cells. Inhibition of IFN-beta expression directly correlated with a reduction in the binding of NF-kappaB (p50-RelA) complex to PRDII after Sendai virus infection in IkappaBalpha-expressing cells, whereas IFN-beta expression and NF-kappaB binding were only slightly reduced in IkappaBbeta-expressing cells. These experiments demonstrate a major role for IkappaBalpha in the regulation of NF-kappaB-induced IFN-beta gene activation and a minor role for IkappaBbeta in the activation process.
The p21-activated kinase (PAK1) is a serine-threonine protein kinase that is activated by binding to the Rho family small
G proteins Rac and Cdc42hs. Both Rac and Cdc42hs have been shown to regulate the activity of the transcription factor NFκB.
Here we show that expression of active Ras, Raf-1, or Rac1 in fibroblasts stimulates NFκB in a PAK1-dependent manner and that
expression of active PAK1 can stimulate NFκB on its own. Similarly, in macrophages activation of NFκB as well as transcription
from the tumor necrosis factor α promoter depends on PAK1. In these cells lipopolysaccharide is a potent activator of PAK1
kinase activity. We also demonstrate that expression of active PAK1 stimulates the nuclear translocation of the p65 subunit
of NFκB but does not activate the inhibitor of κB kinases α or β. These data demonstrate that PAK1 is a crucial signaling
molecule involved in NFκB activation by multiple stimuli.
Stimulation of growth factor signaling has been implicated in the development of invasive phenotypes and the activation of p21-activated kinase (Pak1) in human breast cancer cells (Adam, L., Vadlamudi, R., Kondapaka, S. B., Chernoff, J., Mendelsohn, J., and Kumar, R. (1998) J. Biol. Chem. 273, 28238-28246; Adam, L., Vadlamudi, R., Mandal, M., Chernoff, J., and Kumar, R. (2000) J. Biol. Chem. 275, 12041-12050). To study the role of Pak1 in the regulation of motility and growth of breast epithelial cells, we developed human epithelial MCF-7 clones that overexpressed the kinase-active T423E Pak1 mutant under an inducible tetracycline promoter or that stably expressed the kinase-active H83L,H86L Pak1 mutant, which is deficient in small GTPase binding sites. The expression of both T423E and H83L,H86L Pak1 mutants in breast epithelial cells was accompanied by increased cell motility without any apparent effect on the growth rate of cells. The T423E Pak1 mutant was primarily localized to filopodia, and the H83L,H86L Pak1 mutant was primarily localized to ruffles. Cells expressing T423E Pak1 exhibited a regulatable stimulation of mitogen-activated protein kinase and Jun N-terminal kinase activities. The expression of kinase-active Pak1 mutants significantly stimulated anchorage-independent growth of cells in soft agar in a preferential mitogen-activated protein kinase-sensitive manner. In addition, regulatable expression of kinase-active Pak1 resulted in an abnormal organization of mitotic spindles characterized by appearance of multiple spindle orientations. We also provide evidence to suggest a close correlation between the status of Pak1 kinase activity and base-line invasiveness of human breast cancer cells and breast tumor grades. This study is the first demonstration of Pak1 regulation of anchorage-independent growth, potential Pak1 regulation of invasiveness, and abnormal organization of mitotic spindles of human epithelial breast cancer cells.
Activation of the canonical mitogen-activated protein kinase (MAPK) cascade by soluble mitogens is blocked in non-adherent cells. It is also blocked in cells in which the cAMP-dependent protein kinase (PKA) is activated. Here we show that inhibition of PKA allows anchorage-independent stimulation of the MAPK cascade by growth factors. This effect is transient, and its duration correlates with sustained tyrosine phosphorylation of paxillin and focal-adhesion kinase (FAK) in non-adherent cells. The effect is sensitive to cytochalasin D, implicating the actin cytoskeleton as an important factor in mediating this anchorage-independent signalling. Interestingly, constitutively active p21-activated kinase (PAK) also allows anchorage-independent MAPK signalling. Furthermore, PKA negatively regulates PAK in vivo, and whereas the induction of anchorage-independent signaling resulting from PKA suppression is blocked by dominant negative PAK, it is markedly prolonged by constitutively active PAK. These observations indicate that PKA and PAK are important regulators of anchorage-dependent signal transduction.
Helicobacter pylori, the causative agent of several human gastric diseases, induces activation of the immediate early response transcription
factor nuclear factor κB (NF-κB), which subsequently triggers release of proinflammatory cytokines in colonized epithelial
cells. Here we report that inH. pylori infection p21-activated kinase 1 (PAK1) activates NF-κB. Activated PAK1 associates with NF-κB-inducing kinase, which upon
activation directs the activity of IκB kinases to IκBα. Our results indicate that in epithelial cells PAK1 participates in
a unique pathway that links H. pylori-dependent effector molecules to the activation of NF-κB and the induction of the innate immune response.
The p21-activated kinases (PAKs), in common with many kinases, undergo multiple autophosphorylation events upon interaction
with appropriate activators. The Cdc42-induced phosphorylation of PAK serves in part to dissociate the kinase from its partners
PIX and Nck. Here we investigate in detail how autophosphorylation events affect the catalytic activity of PAK by altering
the autophosphorylation sites in both α- and βPAK. Bothin vivo and in vitro analyses demonstrate that, although most phosphorylation events in the PAK N-terminal regulatory domain play no direct role
in activation, a phosphorylation of αPAK serine 144 or βPAK serine 139, which lie in the kinase inhibitory domain, significantly
contribute to activation. By contrast, sphingosine-mediated activation is independent of this residue, indicating a different
mode of activation. Thus two autophosphorylation sites direct activation while three others control association with focal
complexes via PIX and Nck.
Stimulation of p21-activated kinase-1 (Pak1) signaling promotes motility, invasiveness, anchorage-independent growth and abnormal mitotic assembly in human breast cancer cells. Here, we provide new evidence that, before the onset of mitosis, activated Pak1 is specifically localized with the chromosomes during prophase and on the centrosomes in metaphase and moves to the contraction ring during cytokinesis. To identify mitosis-specific substrates of Pak1, we screened a synchronized G2-M expression library by using a glutathione transferase Pak1 solid-phase-based kinase reaction. This analysis identified histone H3 as a substrate of Pak1 both in vitro and in vivo, and it specifically interacted with Pak1 but not Pak2 or Pak3. Site-directed mutagenesis indicated that Pak1 phosphorylates histone H3 on Ser10. Expressions of the wild-type, or catalytically active, Pak1 caused it to appear at the poles corresponding to mitotic centrosomes in a variety of mammalian cells. Together, these results suggest for the first time that Pak1 interacts with and phosphorylates histone H3 and may thus influence the Pak1-histone H3 pathway, which in turn may influence mitotic events in breast cancer cells.
p21-activated kinase 1 (Pak1) has been shown recently to induce hyperplasia in the mammary epithelium, a phenotype also manifested by overexpression of cyclin D1, a known indicator of the proliferative stage. Here we investigated the role of the Pak1 pathway in the expression of cyclin D1 using tissue culture models and transgenic mice expressing activated Pak1 in mammary glands. We found that hyperplastic mammary glands from catalytically active Pak1 transgenic mice exhibit a 5- to 7-fold increased expression of cyclin D1 as compared with stage-matched wild-type mice. In addition, Pak1 levels were elevated in human breast tumors and also correlated well with increased cyclin D1 expression. Increased expression of Pak1 in breast cancer cells stimulated cyclin D1 promoter activity, elevated levels of cyclin D1 mRNA, protein, and nuclear accumulation of cyclin D1. Conversely, Pak1 inhibition by an auto-inhibitory peptide (amino acids 83-149) or Pak1 knockdown by short interference RNA markedly reduced the expression of cyclin D1, suggesting a requirement of a functional Pak1 pathway for optimal expression of cyclin D1. Results from deletion and mutant analysis indicate that Pak1 regulates cyclin D1 transcription by means of an NF-kappaB-dependent pathway. Together, these findings suggest a model wherein Pak1 regulation of cyclin D1 expression might involve an NF-kappaB-dependent pathway and that hyperplasia in the mammary glands of Pak1-TG mice may be associated, at least in part, with the up-regulation of cyclin D1, and that Pak1 is up-regulated in human breast tumors.
The p21-activated kinase-1 (Pak-1) promotes cell motility and invasiveness. Pak-1 is activated by the Rac, Rho, and Cdc42 small GTPases in response to a variety of stimuli including ras and phosphatidylinositol 3'-kinase/AKT pathway activation. Because Pak-1 plays a central role in regulating cell motility and invasiveness, we sought to determine whether Pak-1 may be involved in the malignant progression of colorectal carcinoma.
Pak-1 expression was examined by immunohistochemistry in archived tissues from normal human colons, tubular and tubulovillous adenomas, invasive adenocarcinomas (stages I-III/IV), and lymph node metastases (184 total specimens from 38 patients). Specific cytoplasmic immunostaining was evaluated for overall intensity and uniformity to derive a combined histoscore (stain intensity x percentage of epithelium stained).
Pak-1 expression was increased significantly with colorectal cancer progression from normal tissue to lymph node metastases (P < 0.0001). Furthermore, Pak-1 expression was increased significantly in adenomas, invasive carcinomas, and lymph node metastases compared with normal colon (P < 0.0001). Strikingly, Pak-1 expression was significantly higher in lymph node metastases than in invasive cancers, adenomas, or normal colon (P < 0.0001). Moreover, in patients with multiple lesions representing different stages of disease, Pak-1 expression was increased specifically in the most advanced lesions.
This study demonstrates that Pak-1 expression is increased significantly with malignant progression of human colorectal carcinoma. These data, along with numerous functional studies demonstrating a central role for Pak-1 activity in tumor invasiveness and motility, implicate Pak-1 as an exciting target for therapy of colorectal carcinoma.
The incidence and mortality of gastric cancer have fallen dramatically in US and elsewhere over the past several decades. Nonetheless, gastric cancer remains a major public health issue as the fourth most common cancer and the second leading cause of cancer death worldwide. Demographic trends differ by tumor location and histology. While there has been a marked decline in distal, intestinal type gastric cancers, the incidence of proximal, diffuse type adenocarcinomas of the gastric cardia has been increasing, particularly in the Western countries. Incidence by tumor sub-site also varies widely based on geographic location, race, and socio-economic status. Distal gastric cancer predominates in developing countries, among blacks, and in lower socio-economic groups, whereas proximal tumors are more common in developed countries, among whites, and in higher socio-economic classes. Diverging trends in the incidence of gastric cancer by tumor location suggest that they may represent two diseases with different etiologies. The main risk factors for distal gastric cancer include Helicobacter pylori (H pylori) infection and dietary factors, whereas gastroesophageal reflux disease and obesity play important roles in the development of proximal stomach cancer. The purpose of this review is to examine the epidemiology and risk factors of gastric cancer, and to discuss strategies for primary prevention.
Hepatocellular carcinoma (HCC) is one of the major malignancies in the world. The prognosis of HCC is poor, due to frequent intrahepatic metastasis and tumor recurrence. P21-activated protein kinase (Pak1), a main downstream effector of small Rho GTPases, Rac1 and Cdc42, plays an important role in the regulation of cell morphogenesis, motility, mitosis, and angiogenesis. Here, we show that Pak1 gene was overexpressed in human HCCs. Overexpression of Pak1 in human HCCs was associated with more aggressive tumor behavior in terms of more metastatic phenotype and more advanced tumor stages. In addition, HCC cell line stably expressing Pak1 displayed increased cell motility rates and, conversely, knockdown of endogenous Pak1 expression by small interfering RNA reduced the migration rates of HCC cells. In an established metastatic HCC cell line, we found that Pak1 was overexpressed compared with its primary HCC cell line and this overexpression was associated with higher cell motility. Importantly, we found that c-Jun NH(2)-terminal kinase (JNK) was activated in HCC cell lines overexpressing Pak1. Inhibition of the JNK activity by chemical inhibitor significantly reduced the migration rates of HCC cells via attenuation of paxillin phosphorylation at Ser(178). In conclusion, our results document that Pak1 is overexpressed in HCCs and plays an important role in the metastasis of HCC. The mechanism by which Pak1 induces cancer metastasis may involve activation of JNK and phosphorylation of paxillin.
Elucidation of mechanisms regulating cell cycle progression is of fundamental importance for cell and cancer biology. Although several genes and signaling pathways are implicated in G1-S regulation, less is known regarding the mechanisms controlling cell cycle progression through G2 and M phases. We report that extracellular signal-regulated kinase 5 (ERK5), a member of the mitogen-activated protein kinases, is activated at G2-M and required for timely mitotic entry. Stimulation of ERK5 activated nuclear factor kappaB (NFkappaB) through ribosomal S6 kinase 2 (RSK2)-mediated phosphorylation and degradation of IkappaB. Furthermore, selective inhibition of NFkappaB at G2-M phases substantially delayed mitotic entry and inhibited transcription of G2-M-specific genes, including cyclin B1, cyclin B2, Plk-1, and cdc25B. Moreover, inhibition of NFkappaB at G2-M diminished mitosis induced by constitutive activation of ERK5, providing a direct link between ERK5, NFkappaB, and regulation of G2-M progression. We conclude that a novel ERK5-NFkappaB signaling pathway plays a key role in regulation of the G2-M progression.
The serine-threonine kinase PAK1 is activated by small GTPase-dependent and -independent mechanisms and promotes cell survival.
However, the role of tyrosyl phosphorylation in the regulation of PAK1 function is poorly understood. In this study, we have
shown that the prolactin-activated tyrosine kinase JAK2 phosphorylates PAK1 in vivo. Wild type, but not kinase-dead, JAK2 directly phosphorylates PAK1 in cells and in an in vitro kinase assay. PAK1 tyrosines 153, 201, and 285 were identified as sites of JAK2 tyrosyl phosphorylation by mass spectrometry
and two-dimensional peptide mapping. Mutation of PAK1 tyrosines 153, 201, and 285 to phenylalanines individually or in combination
implicated these PAK1 tyrosines in the regulation of PAK1 kinase activity. Tyrosyl phosphorylation by JAK2 significantly increases
PAK1 kinase activity, whereas similar phosphorylation of the PAK1 Y153F,Y201F,Y285F mutant has no effect on PAK1 activity.
Tyrosyl phosphorylation of wild type PAK1 decreases apoptosis induced by serum deprivation and staurosporine treatment and
increases cell motility. In contrast, these parameters are unaltered in the PAK1 Y153F,Y201F,Y285F mutant. Our findings indicate
that JAK2 phosphorylates PAK1 at these specific tyrosines and that this phosphorylation plays an important role in cell survival
and motility.
Kaposi's sarcoma-associated herpes virus (KSHV) contributes to the pathogenesis of Kaposi's sarcoma and primary effusion lymphomas. KSHV encodes a G protein-coupled receptor (KSHV-GPCR) that signals constitutively and transforms NIH3T3 cells. Here, we show that KSHV-GPCR transformation requires activation of the small G protein Rac1 and its effector, the p21-activated kinase 1 (Pak1). Either transient or sustained expression of KSHV-GPCR activated both Rac1 and Pak1. Furthermore, expression of dominant-negative mutants of Rac (RacN17) or Pak1 (PakR299, Pak-PID) inhibited KSHV-GPCR-induced focus formation and growth in soft agar. We also demonstrate that signaling from Pak1 to nuclear factor-kappaB (NFkappaB) is required for cell transformation induced by KSHV-GPCR. KSHV-GPCR induced transcriptional activation by NFkappaB. This process is inhibited by the PAK-PID, whereas reciprocally, expression of constitutively active Pak1 (PakL107F) activated NFkappaB comparably to KSHV-GPCR. The Pak-PID and RacN17 inhibited the KSHV-GPCR-induced phosphorylation of inhibitor of kappaB kinase-beta and inhibitor of kappaB-alpha, implying that it is Pak1-dependent phosphorylation and subsequent destruction of the inhibitor of kappaB proteins that allows NFkappaB activation. Finally, experiments with the KSHV-GPCR inverse agonist interferon-gamma-inducible protein-10, the Galpha(i) inhibitor pertussis toxin, and an inhibitor of phosphatidylinositol 3'-kinase, wortmannin, indicate that signaling through the Galpha(i) pathway and phosphatidylinositol 3'-kinase contributes to the cell transformation and NFkappaB activation induced by the KSHV-GPCR.
P21-activated kinases (Paks), a family of serine/threonine kinases, are effectors of the Rho GTPases Cdc42 and Rac1. Mammalian Pak1 and Pak homologs in simple eukaryotes are implicated in controlling G(2)/M transition and/or mitosis. Another serine/threonine kinase, polo-like kinase 1 (Plk1), is an important regulator of mitotic events, such as centrosome maturation, mitotic entry, spindle formation, sister chromatid cohesion and cytokinesis. Plk1 phosphorylation is thought to be one of the critical regulatory events leading to these Plk1-mediated functions. We show here that Pak1 is required for cell proliferation, mitotic progression and Plk1 activity in HeLa cells. Gain or loss of Pak function directly impacted phosphorylation and activity of Plk1. Phosphorylation of Plk1 on Ser 49 is important for metaphase-associated events. Inhibition of Pak activity leads to delay in G(2)/M progression and abnormal spindle formation, mirroring some attributes of Plk1 deregulation. Our results reveal a role for Pak in regulating Plk1 activity and mitotic progression, and connect Pak to the complex protein interaction network enabling cell division.
Cancer of the stomach is one of the most commonly diagnosed malignancies and remains an important cause of mortality world wide. This type of cancer is not uniformly distributed among populations but shows a marked variation in both incidence and mortality. Although gastric cancer is declining in many parts of the world, the reasons for this decline are not well understood and its etiology remains unclear. Several factors are suspected to play a role in gastric carcinogenesis, including the effects of diet, exogenous chemicals, intragastric synthesis of carcinogens, genetic factors, infectious agents and pathological conditions in the stomach (such as gastritis). A new look at the results of epidemiological and experimental studies is important for the establishment of strategies for control. Since cancer of the stomach has a very poor prognosis in its more advanced stages, such a control program must have its main focus on primary prevention. This review describes our knowledge about cancer of the stomach regarding epidemiology, pathogenesis and prevention.
FoxM1 (previously named WIN, HFH-11 or Trident) is a Forkhead box (Fox) transcription factor widely expressed in proliferating cells. Various findings, including a recent analysis of FoxM1 knockout mice, suggest that FoxM1 is required for normal S-M coupling during cell cycle progression. To study the regulatory role of FoxM1 and its downstream regulatory targets, three stably transfected HeLa lines that display doxycycline (dox)-inducible FoxM1 expression were established. Over-expression of FoxM1 by dox induction facilitates growth recovery from serum starvation. Quantitation of cyclin B1 and D1 levels using flow cytometric, Western and Northern analyses reveals that elevated FoxM1 levels lead to stimulation of cyclin B1 but not cyclin D1 expression. Transient reporter assays in the dox-inducible lines and upon co-transfection with a constitutive FoxM1 expression plasmid suggest that FoxM1 can activate the cyclin B1 promoter.
Gastric cancer is one of the leading causes of cancer mortality in the world. Gastric adenocarcinomas account for more than 95% of gastric tumors, whereas gastrointestinal stromal tumors (GISTs) are the most common neoplasms of the rare gastric mesenchymal tumors. Although the incidence of mid-distal gastric adenocarcinomas is decreasing, the incidence of gastroesophageal junctional tumors and Barrett's adenocarcinomas is increasing for unknown reasons. The majority of gastric tumors are sporadic in nature. However, there are rare, inherited gastric cancer predisposition traits, such as germline p53 (Li-Fraumeni syndrome) as well as E-cadherin (CDH1) alterations in familial diffuse gastric cancers. Gastric cancer has been observed to be part of the spectrum of neoplasms associated with germline mismatch repair gene (MMR) alterations that give rise to the hereditary nonpolyposis colorectal cancer (HNPCC) entity. Comparative genomic hybridization analyses have identified several amplifications and losses of DNA copy numbers in gastric cancers. Loss of heterozygosity (LOH) studies have shown several chromosomal loci with significant allelic loss, thus indicating the possibility of harboring a tumor suppressor gene important in gastric tumorigenesis. Microsatellite instability (MIS) and associated alteration of the TGF-bIIR, IGFRII, BAX, E2F-4, hMSH3, and hMSH6 genes are found in a subset of gastric carcinomas. Cell adhesion molecule abnormalities such as those involving CDH1 may play an important role in diffuse-type gastric cancer development. Although, multiple somatic alterations have been described in gastric carcinomas at the molecular level, the significance of these changes in gastric tumorigenesis remains to be established in most instances. The critical molecular alterations in gastric cancers that may lead to advances in our armamentarium to combat this lethal disease remain to be fully characterized.
The serine/threonine kinase p21-activated kinase 1 (Pak1) controls the actin cytoskeletal and ruffle formation through mechanisms that are independent of GTPase activity. Here we identify filamin FLNa as a Pak1-interacting protein through a yeast two-hybrid screen using the amino terminus of Pak1 as a bait. FLNa is stimulated by physiological signalling molecules to undergo phosphorylation by Pak1 and to interact and colocalize with endogenous Pak1 in membrane ruffles. The ruffle-forming activity of Pak1 is functional in FLNa-expressing cells but not in FLNa-deficient cells. In FLNa, the Pak1-binding site involves tandem repeat 23 in the carboxyl terminus and phosphorylation takes place on serine 2152. The FLNa-binding site in Pak1 is localized between amino acids 52 and 132 in the conserved Cdc42/Rac-interacting (CRIB) domain; accordingly, FLNa binding to the CRIB domain stimulates Pak1 kinase activity. Our results indicate that FLNa may be essential for Pak1-induced cytoskeletal reorganization and that the two-way regulatory interaction between Pak1 and FLNa may contribute to the local stimulation of Pak1 activity and its targets in cytoskeletal structures.
Stimulation of p21-activated kinase-1 (Pak1) induces cytoskeleton reorganization and signaling pathways in mammary cancer cells. Here, we show that inhibition of Pak1 kinase activity by a dominant-negative fragment or by short interference RNA markedly reduced the estrogen receptor-alpha (ER) transactivation functions. To understand the role of Pak1 in mammary glands, we developed a murine model expressing constitutively active Thr423 glutamic acid Pak1 driven by the beta-lactoglobulin promoter. We show that mammary glands from these mice developed widespread hyperplasia associated with apocrine metaplasia and lobuloalveolar hyperdevelopment during lactation. Mammary tissues with active Pak1 also exhibited an increased activation of mitogen-activated protein kinase and stimulated transactivation functions of the ER and expression of endogenous ER target genes. Furthermore, Pak1 directly phosphorylated the activation function-2 domain of the ER at the N-terminal residue Ser305, and its mutation to Ala (S305A) abolished the Pak1-mediated phosphorylation and transactivation functions of the ER, while its mutation to glutamic acid (S305E) promoted transactivation activity of ER. These findings reveal a novel role for the Pak1-ER pathway in promoting hyperplasia in mammary epithelium.
The p21 activated kinases (Paks), an evolutionarily conserved family of serine/threonine kinases, are important for a variety of cellular functions including cell morphogenesis, motility, survival, mitosis, and angiogenesis. Paks are widely expressed in numerous tissues and are activated by growth factors and extracellular signals through GTPase-dependent and -independent mechanisms. Overexpression of Paks in epithelial cancer cells has been shown to increase migration potential, increase anchorage independent growth, and cause abnormalities in mitosis. Dysregulation of Paks has been reported in several human tumors and neurodegenerative diseases. A growing list of novel Pak interacting proteins has opened up exciting avenues of investigation by which to understand the functions of Paks in tumorigenesis. In this review, we will summarize the current knowledge of the Paks family with respect to emerging cellular functions and possible contributions to cancer.
Amplification of chromosomal regions leads to an increase of DNA copy numbers and expression of oncogenes in many human tumors. The identification of tumor-specific oncogene targets has potential diagnostic and therapeutic implications. To identify distinct spectra of oncogenic alterations in ovarian carcinoma, metaphase comparative genomic hybridization (mCGH), array CGH (aCGH), and ovarian tumor tissue microarrays were used in this study. Twenty-six primary ovarian carcinomas and three ovarian carcinoma cell lines were analyzed by mCGH. Frequent chromosomal overrepresentation was observed on 2q (31%), 3q (38%), 5p (38%), 8q (52%), 11q (21%), 12p (21%), 17q (21%), and 20q (52%). The role of oncogenes residing in gained chromosomal loci was determined by aCGH with 59 genetic loci commonly amplified in human tumors. DNA copy number gains were most frequently observed for PIK3CA on 3q (66%), PAK1 on 11q (59%), KRAS2 on 12p (55%), and STK15 on 20q (55%). The 11q13-q14 amplicon, represented by six oncogenes (CCND1, FGF4, FGF3, EMS1, GARP, and PAK1) revealed preferential gene copy number gains of PAK1, which is located at 11q13.5-q14. Amplification and protein expression status of both PAK1 and CCND1 were further examined by fluorescence in situ hybridization and immunohistochemistry using a tissue microarray consisting of 268 primary ovarian tumors. PAK1 copy number gains were observed in 30% of the ovarian carcinomas and PAK1 protein was expressed in 85% of the tumors. PAK1 gains were associated with high grade (P < 0.05). In contrast, CCND1 gene alterations and protein expression were less frequent (10.6% and 25%, respectively), suggesting that the critical oncogene target of amplicon 11q13-14 lies distal to CCND1. This study demonstrates that aCGH facilitates further characterization of oncogene candidates residing in amplicons defined by mCGH.
Previously, we showed PAK-PIX-GIT targets and regulates focal adhesions; here, we uncover a different function for the complex at the centrosome. Active PAK1 is particularly evident in mitosis and phosphorylates the centrosomal adaptor GIT1 on serine 517. Interestingly, direct centrosome targeting activates the kinase via a process not requiring Rho GTPases; excision of the centrosome prevents this activation. Once activated, PAK1 dissociates from PIX/GIT but can bind to and phosphorylate the important centrosomal kinase Aurora-A. PAK1 promotes phosphorylation of Aurora-A on Thr288 and Ser342, which are key sites for kinase activation in mitosis. In vivo PAK activation causes an accumulation of activated Aurora-A; conversely, when betaPIX is depleted or PAK is inhibited, there is a delay in centrosome maturation. These observations may underlie reported effects of active PAK on cells, including histone H3 phosphorylation, alterations in centrosome number, and progression through mitosis.
The pivotal role of kinases in signal transduction and cellular regulation has lent them considerable appeal as pharmacological targets across a broad spectrum of cancers. p21-activated kinases (Paks) are serine/threonine kinases that function as downstream nodes for various oncogenic signalling pathways. Paks are well-known regulators of cytoskeletal remodelling and cell motility, but have recently also been shown to promote cell proliferation, regulate apoptosis and accelerate mitotic abnormalities, which results in tumour formation and cell invasiveness. Alterations in Pak expression have been detected in human tumours, which makes them an attractive new therapeutic target.
Kaposi's sarcoma (KS) is the most common neoplasm in untreated AIDS patients and accounts for significant morbidity and mortality worldwide. We have recently reported that Notch signaling (which plays an important role in cell proliferation, apoptosis, and oncogenesis) is constitutively activated in KS tumor cells. Blockade of this activity using gamma-secretase inhibitors resulted in apoptosis of SLK cells, a KS tumor cell line; however, this apoptosis was preceded by a prolonged G(2)-M cell cycle arrest. This result led us to hypothesize that the cells were undergoing mitotic catastrophe, an abnormal mitosis that leads to eventual cell death. Here, we show that Notch inhibition in KS tumor cells using gamma-secretase inhibitors or Notch-1 small interfering RNA resulted in G(2)-M cell cycle arrest and mitotic catastrophe characterized by the presence of micronucleated cells and an increased mitotic index. Interestingly, Notch inhibition led to a sustained increase in nuclear cyclin B1, a novel observation suggesting that Notch signaling can modulate expression of this critical cell cycle protein. Further analysis showed the induction of cyclin B1 was due, at least in part, to increased nuclear factor-kappaB (NF-kappaB) activity, which was also required for the G(2)-M growth arrest after Notch inhibition. Taken together, these studies suggest that Notch inhibition can initiate aberrant mitosis by inducing NF-kappaB activity that inappropriately increases cyclin B1 resulting in cell death via mitotic catastrophe.
The p21-activated kinase 1 (Pak1) is a serine/threonine kinase whose activity is regulated by both Rho GTPases and AGC kinase family members. It plays a role in cytoskeletal remodeling and cell motility as well as cell proliferation, angiogenesis, tumorigenesis and metastasis. An involvement of Pak1 in renal cell carcinoma (RCC), which remains highly refractory to chemotherapy and radiotherapy, remains to be investigated. Pak1 expression, phosphorylation and kinase activity were examined in RCC cell lines and human tissue from normal and renal carcinoma. We report increased Pak1 expression and constitutive activity in the membrane and nucleus but not the cytoplasm of resected human RCC. To study a role for Pak1 in RCC, we developed 786-0 clones that expressed either a kinase-active Pak1L83,L86 2 different Pak1 dominant negative mutants, Pak1R299 and Pak1L83,L86,R299 or Pak1 siRNA. The expression of Pak1L83,L86 increased 786-0 proliferation, motility and anchorage independent growth, while the dominant negative mutants and Pak1 siRNA abrogated these effects. In addition, Pak1L83,L86 conferred resistance to 5-fluorouracil with a 40%+/-10% increase in cell viability. Conversely, Pak1L83,L86,R299, Pak1R299 and Pak1 siRNA conferred sensitivity with a 65.2%+/-5.5%, 69.2%+/-3.3% and 73.0%+/-8.4% loss in viability, respectively. Finally, Pak1 plays a role in renal tumor growth in vivo. Only 33% of mice developed tumors in the Pak1L83,L86,R299 group and no tumors developed from Pak1R299 cell challenge. Together these findings point to Pak1 as an exciting target for therapy of renal cancer, which remains highly refractory to existing treatments.
Molecular epidemiology, pathogenesis and prevention of gastric cancer
- Stadtlander