MICRORNA-193B FUNCTIONS AS A TUMOR SUPPRESSOR IN MALIGNANT MELANOMA
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Cutaneous melanoma is an increasingly common skin cancer characterized by aggressive metastatic growth and poor prognosis. The mechanisms behind melanoma progression are not fully understood, but emerging evidence suggests that a group of newly discovered small regulatory RNAs, named microRNAs (miRNAs), plays an important role. miRNAs are ~ 22 nucleotide single strand non-coding RNAs that post-transcriptionally regulate gene expression by binding to target messenger RNAs (mRNAs), leading to mRNA degradation and translation inhibition. Abnormal expression of miRNAs has been observed in human malignancies and is associated with tumorigenesis. The main goals of this thesis are to investigate miRNA dysregulation in melanoma and to identify potential miRNAs involved in melanoma pathogenesis. Initially, the expression of 470 miRNAs was profiled in 8 metastatic melanoma and 8 benign nevus tissue samples. We discovered unique miRNA expression profiles and identified differentially expressed miRNAs in melanomas as compared to nevi. miR-193b was one of the most significantly downregulated miRNAs in melanoma, and its function and regulatory targets were unknown. Subsequently, in vitro functional studies revealed that ectopic expression of miR-193b in melanoma cells drastically repressed cell proliferation and migration. Although it does not directly induce apoptosis in melanoma cells, miR-193b does sensitize these cells to ABT-737-mediated cell death. In concert with functional studies, gene expression analysis and in silico target prediction were performed to globally screen for mRNA targets of miR-193b. We identified eighteen genes as candidates in that they were downregulated by miR-193b and contained predicted miR-193b binding sites. Based on their known biological functions, three genes were particularly interesting: cyclin D1 (CCND1), myeloid cell leukemia sequence 1 (Mcl-1), and stathmin 1 (STMN1). CCND1 and Mcl-1 are two well-known melanoma oncogenes, and we validated their role in cell proliferation and apoptosis respectively. Furthermore, using similar approach, we were the first to identify STMN1 as a novel melanoma oncogene. We demonstrated that CCND1, Mcl-1, and STMN1 were directly regulated by miR-193b. During melanoma progression, reduced expression of miR-193b may promote cell proliferation, migration and survival. Taken together, this thesis describes the dysregulation of miRNAs in melanoma and demonstrates that miR-193b functions as a tumor suppressor.