[Show abstract][Hide abstract] ABSTRACT: Accumulating evidence suggests that microRNAs (miRNAs) can function as oncogenes or as tumor suppressor genes depending on the tissue type or target. Therefore, clarification of the specific roles of miRNAs is vital for the diagnosis and treatment of cancer. In the present study, miR-451 was found to be downregulated in hepatocellular carcinoma (HCC) tissues when compared to that in adjacent tissues. Functional analysis showed that, in vitro, miR-451 inhibited the migration of hepatoma cell lines HepG2 and SK-Hep-1. Further investigation of the molecular mechanisms identified activating transcription factor 2 (ATF2) as a target of miR-451. miR-451 inhibited ATF2 expression by binding to the 3'UTR. An in vivo assay revealed a significant negative correlation between miR-451 and ATF2 in liver cancer tissues. According to previous findings reported in the literature, the opposing functions of ATF2 are related to its subcellular localization. In the nucleus, ATF2 displays oncogenic activities in melanoma. In the present study, ATF2 exhibited a higher expression level in the nucleus in tumoral tissues of HCC as detected by immunohistochemistry. In conclusion, in this study, we identified a potential target of miR-451, ATF2, and revealed a novel role of miR-451 in the inhibition of the migratory ability of hepatoma cell lines.
[Show abstract][Hide abstract] ABSTRACT: Loss or attenuated expression of the tumor suppressor gene FHIT is associated paradoxically with poor progression of human tumors. Fhit promotes apoptosis and regulates reactive oxygen species, however, the mechanism by which Fhit inhibits tumor growth in animals remains unclear. In this study, we used a multi-discliplinary approach based on bioinformatics, small RNA library screening, human tissue analysis and a xenograft mouse model to identify a novel member of the miR-548 family in the fourth intron of the human FHIT gene. Characterization of this human-specific microRNA illustrates the importance of this class of microRNAs in tumor suppression and may influence interpretation of Fhit action in human cancer.
[Show abstract][Hide abstract] ABSTRACT: Eye absent (Eya) proteins are involved in cell fate determination in a broad spectrum of cells and tissues. Aberrant expression of Eya2 has been documented in a variety of cancers and correlates with clinical outcome. However, whether microRNAs (miRNAs) can regulate Eya2 expression remains unknown. Here, we show that miR-30a represses Eya2 expression by binding to the 3’-untranslated region of Eya2. Overexpression of Eya2 in miR-30a-transfected breast cancer cells effectively rescued the inhibition of cell proliferation and migration caused by miR-30a. Knockdown of Eya2 by small-interfering RNA (siRNA) in breast cancer cells mimicked the effect induced by miR-30a and abolished the ability of miR-30a to regulate breast cancer cell proliferation and migration. The miR-30a/Eya2 axis could regulate G1/S cell cycle progression, accompanied by the modulation of expression of cell cycle-related proteins, including cyclin A, cyclin D1, cyclin E, and c-Myc. Moreover, miR-30a expression was downregulated in breast cancer patients, and negatively correlated with Eya2, which was upregulated in breast cancer patients. These data suggest that the miR-30a/Eya2 axis may play an important role in breast cancer development and progression and that miR-30a activation or Eya2 inhibition may be a useful strategy for cancer treatment.
Biochemical and Biophysical Research Communications 01/2014; · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Thousands of long noncoding RNAs (lncRNAs) have been reported in mammalian genomes. These RNAs represent an important subset of pervasive genes involved in a broad range of biological functions. Aberrant expression of lncRNAs is associated with many types of cancers. Here, in order to explore the potential lncRNAs involved in hepatocellular carcinoma (HCC) oncogenesis, we performed lncRNA gene expression profile analysis in 3 pairs of human HCC and adjacent non-tumor (NT) tissues by microarray.
PLoS ONE 01/2014; 9(7):e101707. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: An increasing data indicates that altered microRNAs (miRNAs) participate in the radiation-induced DNA damage response. However, a correlation of mRNA and miRNA profiles across the entire genome and in response to irradiation has not been thoroughly assessed. We analyzed miRNA microarray data collected from HeLa cells after ionizing radiation (IR), quantified the expression profiles of mRNAs and performed comparative analysis of the data sets using target prediction algorithms, Gene Ontology (GO) analysis, pathway analysis, and gene network construction. The results showed that the altered miRNAs were involved in regulation of various cellular functions. miRNA-gene network analyses revealed that miR-186, miR-106b, miR-15a/b, CCND1 and CDK6 played vital role in the cellular radiation response. Using qRT-PCR, we confirmed that twenty-two miRNAs showed differential expression in HeLa cells treated with IR and some of these miRNAs affected cell cycle progression. This study demonstrated that miRNAs influence gene expression in the entire genome during the cellular radiation response and suggested vital pathways for further research.
Chinese Science Bulletin 12/2013; 58(36). · 1.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Long non-coding RNAs (lncRNAs), which represent a new frontier in molecular biology, play important roles in regulating gene expression at epigenetic, transcriptional and post-transcriptional levels. More and more lncRNAs have been found to play important roles in normal cell physiological activities, and participate in the development of varieties of tumors and other diseases. Previously, we have only been able to determine the function of lncRNAs through multiple mechanisms, including genetic imprinting, chromatin remodeling, splicing regulation, mRNA decay, and translational regulation. Application of technological advances to research into the function of lncRNAs is extremely important. The major tools for exploring lncRNAs include microarrays, RNA sequencing (RNA-seq), Northern blotting, real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR), fluorescence in situ hybridization (FISH), RNA interference (RNAi), RNA-binding protein immunoprecipitation (RIP), chromatin isolation by RNA purification (ChIRP), crosslinking-immunopurification (CLIP), and bioinformatic prediction. In this review, we highlight the functions of lncRNAs, and advanced methods to research lncRNA-protein interactions.
Science China. Life sciences 10/2013; 56(10):876-885. · 2.02 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: MicroRNAs (miRNAs) have been shown to be dysregulated in virus-related cancers; however, miRNA regulation of virus-related cancer development and progression remains poorly understood. Here, we report that miR-148a is repressed by hepatitis B virus (HBV) X protein (HBx) to promote cancer growth and metastasis in a mouse model of hepatocellular carcinoma (HCC). Hematopoietic pre-B cell leukemia transcription factor-interacting protein (HPIP) is an important regulator of cancer cell growth. We used miRNA target prediction programs to identify miR-148a as a regulator of HPIP. Expression of miR-148a in hepatoma cells reduced HPIP expression, leading to repression of AKT and ERK and subsequent inhibition of mTOR through the AKT/ERK/FOXO4/ATF5 pathway. HBx has been shown to play a critical role in the molecular pathogenesis of HBV-related HCC. We found that HBx suppressed p53-mediated activation of miR-148a. Moreover, expression of miR-148a was downregulated in patients with HBV-related liver cancer and negatively correlated with HPIP, which was upregulated in patients with liver cancer. In cultured cells and a mouse xenograft model, miR-148a reduced the growth, epithelial-to-mesenchymal transition, invasion, and metastasis of HBx-expressing hepatocarcinoma cells through inhibition of HPIP-mediated mTOR signaling. Thus, miR-148a activation or HPIP inhibition may be a useful strategy for cancer treatment.
The Journal of clinical investigation 01/2013; · 15.39 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cisplatin is a classic chemotherapy agent used for treating human non-small cell lung cancer (NSCLC). However, cisplatin resistance is a challenge against successful clinical use. Glutathione S-transferase P1 (GSTP1) has been reported to contribute to cisplatin resistance in many studies. MicroRNAs (miRNAs) are short non-coding RNAs that are 21-25 nucleotides in length. They play a role in post-transcriptional gene regulation by inducing repression and/or mRNA degradation. Recent studies have shown that miRNAs are responsible for cisplatin resistance. This study aims to determine whether deregulated miRNAs can sensitize human lung adenocarcinoma cells to cisplatin by targeting GSTP1. Real-time RT-PCR revealed that GSTP1 mRNA expression was 2.7 ± 0.38 folds (p=0.039) upregulated in A549/CDDP cells, compared with the parental A549 cells, while miR-513a-3p expression was 0.34 ± 0.03 folds (p=0.023) downregulated. Luciferase activity assay proved that GSTP1 was a target gene of miR-513a-3p, which was confirmed by Western blot analysis. Furthermore, CCK-8 assay showed that overexpression of miR-513a-3p could enhance cisplatin-induced apoptosis in human lung adenocarcinoma cell lines, A549/CDDP and SPC-A-1. In conclusion, our data demonstrated that miR-513a-3p can sensitize human lung adenocarcinoma cells to cisplatin by targeting GSTP1.
Lung cancer (Amsterdam, Netherlands) 06/2012; 77(3):488-94. · 3.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: MicroRNAs may function to promote or suppress tumor development, depending on the cellular context. The important role of microRNAs in regulating molecular pathways underlying tumorigenesis has been emphasized in hepatocellular carcinoma (HCC). MicroRNAs regulate gene expression via post-transcriptional mechanisms by inhibiting translation or by degrading mRNA. In this study, we show that microRNA-1 (miR-1) and microRNA-499 (miR-499) are capable of repressing the expression of the ets1 proto-oncogene, which plays a fundamental role in the extracellular matrix (ECM) degradation, a process required for tumor cell invasion and migration. We used luciferase reporter assays to demonstrate that miR-1 and miR-499 target the 3' untranslated region (UTR) of ets1. Overexpression of miR-1 and miR-499 in HepG2 cells led to downregulation of ets1 mRNA and protein as assessed by quantitative reverse transcription PCR and western blot analysis. Furthermore, overexpression of miR-1 and miR-499 inhibited the invasion and migration of HepG2 cells in matrigel invasion and transwell migration assays, respectively. These results suggest that miR-1 and miR-499 may play an important role in the pathogenesis of HCC by regulating ets1.
[Show abstract][Hide abstract] ABSTRACT: Valosin containing protein (VCP)/p97 plays various important roles in cells. Moreover, elevated expression of VCP in hepatocellular carcinoma (HCC) is correlated with increased incidence of recurrence. But the role of VCP in HCC progression in vitro and in vivo is unclear. And there are few reports about the regulation mechanism on the expression of VCP in HCC. In this study, it was identified that the level of VCP was frequently increased in human HCC tissues. In addition, down-regulation of VCP with siRNAs could dramatically suppress the genesis and progression of tumor in vivo. It was found that miR-129-5p directly inhibited the expression of VCP in several HCC cell lines. Meanwhile, the level of VCP in HCC tissues was negatively associated with the level of miR-129-5p. Our further investigation showed that the enhanced expression of miR-129-5p also suppressed tumor growth in vivo. Moreover, it was revealed that miR-129-5p could inhibit the degradation of IκBα and increase the apoptosis and reduce the migration of HCC cells by suppressing the expression of VCP. Our results revealed that the expression of VCP was directly regulated by miR-129-5p and this regulation played an important role in the progression of HCC.
PLoS ONE 01/2012; 7(4):e35800. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Most intronic micro-RNAs are coexpressed with their host genes, suggesting that they may play similar roles. The function of miR-483 remains unknown, although it is embedded in an intron of Igf2 gene, which is an activator of hepatocellular carcinoma proliferation. In the present study, we provide evidence that Igf2-derived miR-483 can induce proliferation in hepatocellular carcinoma cells. The miR-483 promotion of proliferation was analysed by soft agar colony formation assay and proliferation curve assay. The effect of miR-483 on Socs3 expression was examined by Western blot and a reporter assay. Our results revealed that Igf2-derived intronic miR-483 was identified by the application of 94G6, an inhibitor of Igf2 at the transcriptional level. All results from the (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) MTT assay, the proliferation curve assay and the soft agar colony formation assay showed that miR-483 promoted the proliferation of hepatocellular carcinoma cells. Finally, Socs3, a putative target predicted by bioinformatics, was regulated by miR-483 at mRNA and protein levels. Direct binding with the 3' UTR was identified by a luciferase activity assay. Our findings demonstrate that Igf2-derived intronic miR-483, through downregulation of its target Socs3, regulates hepatoma cell proliferation and thus may serve as a potential target for hepatocellular carcinoma therapy.
Molecular and Cellular Biochemistry 11/2011; 361(1-2):337-43. · 2.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: microRNAs (miRNAs) are a versatile class of non-coding RNAs involved in regulation of various biological processes. miRNA-122 (miR-122) is specifically and abundantly expressed in human liver. In this study, we employed 3'-end biotinylated synthetic miR-122 to identify its targets based on affinity purification. Quantitative RT-PCR analysis of the affinity purified RNAs demonstrated a specific enrichment of several known miR-122 targets such as CAT-1 (also called SLC7A1), ADAM17 and BCL-w. Using microarray analysis of affinity purified RNAs, we also discovered many candidate target genes of miR-122. Among these candidates, we confirmed that protein kinase, interferon-inducible double-stranded RNA-dependent activator (PRKRA), a Dicer-interacting protein, is a direct target gene of miR-122. miRNA quantitative-RT-PCR results indicated that miR-122 and small interfering RNA against PRKRA may facilitate the accumulation of newly synthesized miRNAs but did not detectably affect endogenous miRNAs levels. Our findings will lead to further understanding of multiple functions of this hepato-specific miRNA. We conclude that miR-122 could repress PRKRA expression and facilitate accumulation of newly synthesized miRNAs.
Nucleic Acids Research 09/2011; 40(2):884-91. · 8.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: MicroRNAs (miRNAs) are endogenous noncoding RNAs that downregulate gene expression by inhibiting translation or promoting mRNA degradation. Although over one-third of miRNAs are located within the intronic regions of transcription units, the expression of such "intron-derived" (or "intronic") miRNAs and their relationship with their host gene remain a mystery. Here, we show that miR-483-5p, which is located within the second intron of the insulin-like growth factor (Igf2) gene, is downregulated in mouse Hepa1-6 cells in a direct correlation with the Igf2 transcript by chromeceptin, an inhibitor of Igf2 at the transcriptional level. Furthermore, miR-483-5p overexpression and knockdown regulates the suppressor of cytokine signalling 3 (Socs3) and Igf2 mRNAs, respectively. Finally, Socs3, a key putative leptin-resistant factor in obesity, is identified as a direct target of miR-483-5p. These data suggest that miR-483-5p can be coexpressed together with its host gene, Igf2, and revealed the link between this miRNA and the IGF2 growth factor. In addition, these observations not only provide supporting evidence for the codependent expression of intronic miRNAs and their host genes in vitro, but also give insight into the role of miR-483-5p in metabolism regulation and tumourigenesis.
Molecular and Cellular Endocrinology 02/2011; 333(1):96-101. · 4.04 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ionizing radiation (IR) causes severe cellular damage both directly and indirectly and disrupts RNA integrity. RNA strand
breaks are the most frequent type of damage caused by IR. RNA damage is involved in the development of degenerative diseases,
including Alzheimer’s disease and Parkinson’s disease. However, the mechanism of mRNA damage and any resulting pathophysiological
outcomes are poorly understood. This is partly because there is a lack of sensitive tools to monitor damage randomly occurring
in RNA, especially RNA strand break damage in a given RNA. In this work, a method using the reverse transcription polymerase
chain reaction (RT-PCR) after poly(A) addition to 3′-end of RNA to determine RNA strand break damage in a specific RNA by
poly(A) polymerase has been developed. The levels of damage in specific mRNAs, including ABL1, TP53, GADD45A and ATR from IR-treated HeLa cells were examined. Strand breaks were detected in all mRNAs examined. The study provides a novel and
sensitive method based on 3′-end poly(A)-tailing RT-PCR to monitor RNA strand break damage.
KeywordsRNA damage–RNA strand break–ionizing radiation–poly(A) polymerase–RT-PCR
Chinese Science Bulletin 01/2011; 56(30):3172-3177. · 1.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This work presents two methods, the Least-square and Bayesian method, to solve the multiple mapping problem in extracting gene expression profiles through the next-generation sequencing. We parallel the tag sequences to genome, and partition them to improving the methods’ efficiency. The essential feature of these methods is that they can solve the multiple mapping problem between genes and short-reads, while generating almost the same estimation in single-mapping situation as the traditional approaches. These two methods are compared by simulation and a real example, which was generated from radiation-induced lung cancer cells (A549), through mapping short-reads to human ncRNA database. The results show that the Bayesian method, as realized by Gibbs sampler, is more efficient and robust than the Least-square method.
Frontiers of Mathematics in China 01/2011; 6(6). · 0.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recently, some miRNAs have been reported to be connected closely with the development of human hepatocellular carcinoma. However, the functions of these miRNAs in HCC remain largely undefined.
The expression profiles of miR-193b were compared between HCC tissues and adjacent normal liver tissues using qRT-PCR method. This method was also be used to screen the potential target genes of miR-193b. A luciferase reporter assay was conducted to confirm target association. Finally, the functional effect of miR-193b in hepatoma cells was examined further.
miR-193b was significantly down-regulated in most of the HCC tissues compared to the matching non-tumoural liver tissues. Furthermore, ectopic expression of miR-193b dramatically suppressed the ability of hepatoma cells to form colonies in vitro and to develop tumours in nude mice. CCND1 and ETS1 were revealed to be regulated by miR-193b directly. By regulating the expressions of these oncogenes, miR-193b induced cell cycle arrest and inhibited the invasion and migration of hepatoma cells.
miR-193b may function as a tumour suppressor in the development of HCC by acting on multiple tumourigenic pathways.
European journal of cancer (Oxford, England: 1990) 10/2010; 46(15):2828-36. · 4.12 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Previous studies have suggested that microRNAs (miRNAs) may play important roles in tumorigenesis, but little is known about the functions of most miRNAs in cancer development. In the present study, we set up a cell-based screen using a luciferase reporter plasmid carrying the whole approximately 4.7 kb 3'-UTR of estrogen receptor alpha (ERalpha) mRNA cotransfected with a synthetic miRNA expression library to identify potential ERalpha-targeting miRNAs. Among all the miRNAs, miR-22 was found to repress robustly the luciferase signal in both HEK-293T and ERalpha-positive MCF-7 cells. Mutation of the target site was found to abrogate this repression effect of miR-22, whereas antagonism of endogenous miR-22 in MDA-MB-231 cells resulted in elevated reporter signals. We assessed the miR-22 expression patterns in five breast cancer cell lines and 23 clinical biopsies and revealed that there is a significant inverse association between the miR-22 levels and ERalpha protein expression. To evaluate the potential of miR-22 as a potential therapeutic intervention, we found that reduction of endogenous ERalpha protein levels and suppression of cancer cell growth could be achieved in MCF-7 cells by miR-22 overexpression in a way that can be recapitulated by the introduction of specific small interfering RNA against ERalpha. The phenomena can be rescued by the reintroduction of ERalpha. Taken together, our data indicate that miR-22 was frequently downregulated in ERalpha-positive human breast cancer cell lines and clinical samples. Direct involvement in the regulation of ERalpha may be one of the mechanisms through which miR-22 could play a pivotal role in the pathogenesis of breast cancer.
[Show abstract][Hide abstract] ABSTRACT: The pleckstrin homology (PH) domain-containing protein casein kinase 2 interacting protein-1 (CKIP-1) plays an important role in regulation of bone formation and muscle differentiation. How CKIP-1 localization is determined remains largely unclear. We observed that isolated CKIP-1-PH domain was predominantly localized in the nucleus and the C-terminus of CKIP-1 counteracted its nuclear localization. The net charge of basic residues and a serine-rich motif within the PH domain plays a pivotal role in the localization switch of both full-length CKIP-1 and the isolated PH domain. We propose that the N-terminal PH domain and C-terminal auto-inhibitory region of CKIP-1 coordinate to determine its subcellular localization and the nucleus-plasma membrane shuttling.
[Show abstract][Hide abstract] ABSTRACT: In recent years, some miRNAs have been reported to be connected closely with the development of human hepatocellular carcinoma. In our previous studies, a set of miRNAs were revealed to be dysregulated in HCC tissues. However, the functions of these miRNAs in HCC remain largely undefined.
The expression profiles of miR-183 were compared between HCC tissues and adjacent normal liver tissues using qRT-PCR method. This method was used to screen the potential target genes of miR-183. A luciferase reporter assay was conducted to confirm target association. Finally, the functional effect of miR-183 in hepatoma cells was examined.
Among the 25 HCC samples analyzed, microRNA-183 was significantly up-regulated (twofold to 367-fold) in 17 samples compared with the matching nontumoral liver tissues. Programmed cell death 4 (PDCD4) was identified as the target gene of miR-183. Moreover, PDCD4 is a proapoptotic molecule involved in TGF-beta1-induced apoptosis in human HCC cells, we found that miR-183 transfectants were resistant to apoptosis induced by TGF-beta1.
We conclude that miR-183 can inhibit apoptosis in human HCC cells by repressing the PDCD4 expression, and miR-183 may play an important role in HCC development.