| Hepatocellular carcinoma (HCC) is a prevalent malignancy worldwide and ranks as one of the leading causes of cancer-related death, especially in East Asia. The mechanism of HCC is highly complicated and heterogeneous that is accompanied by various molecular abnormalities. Even though many potential therapeutic targets and diagnostic biomarkers of this disease have been discovered, its mechanism is still obscure and it is necessary to identify novel oncogenes and tumor suppressors for further investigation and potential clinical application.One of the most significant character of cancer genome is the "instablity", which is resulted from the dysregulation of DNA damage repair-related proteins in cancer cells. The abnormality of DNA damage repair, not only accumulates lots of mutations (active or inhibitory) in genome of cancer cells, but also results in amplification, deletion of gene copy number, causing aberrant expression. By searching and consulting a lot of articles, this study focus on two genes whose products are eligible and haven't been studied in HCC, after eliminating the well-described genes in cancer research. These two genes are Microrchidia CW-type zinc finger (MORC2) and Structure Specific Recognition Protein 1 (SSRP1). MORC2 can function as a transcriptional repressor and it is reported it has important role in chromatin remodeling, facilitating DNA damage repairing. Recently, it is identified as an oncogene in gastric cancer. SSRP1 is involved in transcriptional regulation, DNA damage repair and cell cycle regulation. It is reported that high SSRP1 expression is associated with stem or less-differentiated cells, while low SSRP1 levels are seen in more differentiated cells.By analyzing the gene expression profile from Gene Expression Omnibus (GEO), this study not only verified that MORC2 and SSRP1 were correlated with several pathological character of HCC patients, but also demonstrated that MORC2 and SSRP1 were significantly associated with the overall and disease-free survival of the HCC patients who received surgery. Furthermore, the expression profile data showed that MORC2 and SSRP1 were both up-regulated on mRNA level in HCC. Using immnohistochemistry (IHC) and quantitative real-time PCR (qRT-PCR), this study further validated that MORC2 and SSRP1 were up-regulated on both protein and mRNA level.Next we explore the function of MORC2 and SSRP1 in HCC. To investigate the role of these two proteins in HCC progression, both loss-(siRNA transfection) and gain-function (plasmid packed by lentivirus system) models were established in several HCC cell lines and we conducted CCK-8 assay, plate colony formation assay, cell cycle assay, apoptosis detection by FACS, wound healing assay, migration and invasion assays and so on. This study demonstrated that both SSPR1 and MORC2 promote the progression of HCC; and inhibition of them blocked the progression of HCC respectively. In this study, subcutaneous xenograft and lung metastasis models were also performed. Via vivo study, we validated the function of MORC2 and SSRP1 on promoting cancer cell proliferation and metastasis.Then this study explored the downstream mechanism of MORC2 promoting HCC malignant phenotype and the upstream mechanism of SSRPloverexpression. We demonstrated that MORC2 inhibition would result in the inactivation of Hippo pathway, indicating that its dysregulation would result in the abnormal activation of Hippo pathway in HCC; We also identified microRNA-497 (miR-497) as a posttranscriptional regulator of SSRP1. Ectopic expression of miR-497 inhibited 3'-untranslated-region-coupled luciferase activity and suppressed endogenous SSRP1 expression at both messenger RNA and protein levels. Considering the underexpression of miR-497 is general in HCC, it may be a good explanation for the up-regulation of SSRP1 in HCC.Overall, this study screened and validated MORC2 and SSRP1 were crucial genes which are involved in HCC progression, providing novel potential targets for HCC treatment. |
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