Largely owing to widespread deployment of microarray analysis, many of the transcriptional events associated with invasive cell migration are becoming clear. However, the transcriptional drives to invasive migration are likely modified by alternative splicing of pre-mRNAs to produce functionally distinct patterns of protein expression. Heterogenous nuclear ribonucleoprotein (hnRNP A2) is a known regulator of alternative splicing that is upregulated in a number of invasive cancer types. Here, we report that although siRNA of hnRNP A2 had little influence on the ability of cells to migrate on plastic surfaces, the splicing regulator was clearly required for cells to move effectively on three-dimensional matrices and to invade into plugs of extracellular matrix proteins. We used exon-tiling microarrays to determine that hnRNP A2 controlled approximately six individual splicing events in a three-dimensional matrix-dependent fashion, one of which influenced invasive migration. Here, we show that alternative splicing of an exon in the 5' untranslated region of a gene termed TP53INP2 is a key event downstream of hnRNP A2 that is necessary for cells to invade the extracellular matrix. Furthermore, we report that the consequences of altered TP53INP2 splicing on invasion are likely mediated via alterations in Golgi complex integrity during migration on three-dimensional matrices.
"Furthermore, in a PCa cell line (LNCaP) the drug Genistein, normally used to treat PCa, was shown to up-regulate TP53INP2, among other genes 62. Moreover, it has also been proposed that the occurrence of a specific splice variant of TP53INP2 might be a necessary event in order for malignant cells to invade the ECM 63. Finally, in 2007 a patent was filed that described using measurements of TP53INP2, among other targets, as part of a multivariate signature to monitor the progression of PCa 64. "
[Show abstract][Hide abstract] ABSTRACT: Globally, Prostate cancer (PCa) is the most frequently occurring non-cutaneous cancer, and is the second highest cause of cancer mortality in men. Serum prostate specific antigen (PSA) has been the standard in PCa screening since its approval by the American Food & Drug Administration (FDA) in 1994. Currently, PSA is used as an indicator for PCa - patients with a serum PSA level above 4ng/mL will often undergo prostate biopsy to confirm cancer. Unfortunately fewer than ~30% of these men will biopsy positive for cancer, meaning that the majority of men undergo invasive biopsy with little benefit. Despite PSA's notoriously poor specificity (33%), there is still a significant lack of credible alternatives. Therefore an ideal biomarker that can specifically detect PCa at an early stage is urgently required. The aim of this study was to investigate the potential of using deregulation of urinary proteins in order to detect Prostate Cancer (PCa) among Benign Prostatic Hyperplasia (BPH). To identify the protein signatures specific for PCa, protein expression profiling of 8 PCa patients, 12 BPH patients and 10 healthy males was carried out using LC-MS/MS. This was followed by validating relative expression levels of proteins present in urine among all the patients using quantitative real time-PCR. This was followed by validating relative expression levels of proteins present in urine among all the patients using quantitative real time-PCR. This approach revealed that significant the down-regulation of Fibronectin and TP53INP2 was a characteristic event among PCa patients. Fibronectin mRNA down-regulation, was identified as offering improved specificity (50%) over PSA, albeit with a slightly lower although still acceptable sensitivity (75%) for detecting PCa. As for TP53INP2 on the other hand, its down-regulation was moderately sensitive (75%), identifying many patients with PCa, but was entirely non-specific (7%), designating many of the benign samples as malignant and being unable to accurately identify more than one negative.
Journal of Cancer 01/2014; 5(2):103-14. DOI:10.7150/jca.6890 · 3.27 Impact Factor
"Despite the fact that hnRNP D suppresses expression of many cell cycle regulatory factors by promoting degradation of their mRNAs (57), overexpression of hnRNP D has also been shown to prevent cellular senescence (53) and to lead to tumorigenesis (72). Expression of hnRNP A2/B1 protein is deregulated and overexpressed in many different cancer forms and has been shown to drive tumorigenesis (73) and control invasive cell migration (74) and epithelial-mesenchymal transition (75). This is interesting as high levels of hnRNP A2/B1 enhances splicing of the HPV-16 early pre-mRNAs to generate E7 mRNAs and inhibits HPV-16 late mRNAs splicing, thereby creating the HPV-16 gene expression profile seen in HPV-16 high grade lesions or cancers in which the HPV-16 genome has not integrated into the host cell chromosomes. "
[Show abstract][Hide abstract] ABSTRACT: Human papillomavirus type 16 (HPV-16) 5'-splice site SD3632 is used exclusively to produce late L1 mRNAs. We identified a 34-nt splicing inhibitory element located immediately upstream of HPV-16 late 5'-splice site SD3632. Two AUAGUA motifs located in these 34 nt inhibited SD3632. Two nucleotide substitutions in each of the HPV-16 specific AUAGUA motifs alleviated splicing inhibition and induced late L1 mRNA production from episomal forms of the HPV-16 genome in primary human keratinocytes. The AUAGUA motifs bind specifically not only to the heterogeneous nuclear RNP (hnRNP) D family of RNA-binding proteins including hnRNP D/AUF, hnRNP DL and hnRNP AB but also to hnRNP A2/B1. Knock-down of these proteins induced HPV-16 late L1 mRNA expression, and overexpression of hnRNP A2/B1, hnRNP AB, hnRNP DL and the two hnRNP D isoforms hnRNP D37 and hnRNP D40 further suppressed L1 mRNA expression. This inhibition may allow HPV-16 to hide from the immune system and establish long-term persistent infections with enhanced risk at progressing to cancer. There is an inverse correlation between expression of hnRNP D proteins and hnRNP A2/B1 and HPV-16 L1 production in the cervical epithelium, as well as in cervical cancer, supporting the conclusion that hnRNP D proteins and A2/B1 inhibit HPV-16 L1 mRNA production.
Nucleic Acids Research 09/2013; 41(22). DOI:10.1093/nar/gkt803 · 9.11 Impact Factor
"The ability of hnRNP A1 to control hnRNP A2/B1 expression appears to be relevant for the biology of the cell because increased expression of hnRNP A1 in cancer cells has been shown to correlate with downregulation of hnRNP A2/B1 (42). Several observations indicate a prominent role of hnRNP A2/B1 in the alternative splicing of genes implicated in cellular migration and invasion (13,30,63). Indeed, upregulation of hnRNP A2/B1 correlates with poor prognosis in patients with glioma and its overexpression is sufficient to confer to mouse fibroblasts the ability to form high-grade sarcomas in nude mice (13). "
[Show abstract][Hide abstract] ABSTRACT: Epithelial-to-mesenchymal transition (EMT) is an embryonic program used by cancer cells to acquire invasive capabilities becoming metastatic. ΔRon, a constitutively active isoform of the Ron tyrosine kinase receptor, arises from skipping of Ron exon 11 and provided the first example of an alternative splicing variant causatively linked to the activation of tumor EMT. Splicing of exon 11 is controlled by two adjacent regulatory elements, a silencer and an enhancer of splicing located in exon 12. The alternative splicing factor and oncoprotein SRSF1 directly binds to the enhancer, induces the production of ΔRon and activates EMT leading to cell locomotion. Interestingly, we now find an important role for hnRNP A1 in controlling the activity of the Ron silencer. HnRNP A1 is able to antagonize the binding of SRSF1 and prevent exon skipping. Notably, hnRNP A1, by inhibiting the production of ΔRon, activates the reversal program, namely the mesenchymal-to-epithelial transition, which instead occurs at the final metastasis sites. Also, hnRNP A1 affects Ron splicing by regulating the expression level of hnRNP A2/B1, which similarly to SRSF1 can promote ΔRon production. These results shed light on how splicing regulation contributes to the tumor progression and provide potential targets to develop anticancer therapies.
Nucleic Acids Research 07/2013; 41(18). DOI:10.1093/nar/gkt579 · 9.11 Impact Factor
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