The pro-α1(V) collagen gene (Col5a1) is coordinately regulated by miR-29b with core promoter in cultured cells

ABSTRACTAims: Col5a1 encodes the α1 chain of type V collagen, a quantitatively minor fibrillar collagen that is critical for the formation and function of the organs in the body. MicroRNAs (miRNAs) are small noncoding RNAs that posttranscriptionally regulate biological functions by binding to the 3ʹ-untranslated region (3ʹUTR) of specific target mRNA. In this study, we investigated the posttranscriptional regulation of miRNAs on the Col5a1 gene expression. Materials and Methods: We cultured osteoblasts and fibroblasts of cell lines. To examine the 3ʹUTR activity of the Col5a1 gene, chimeric plasmids constructs containing the core promoter and 3ʹUTR of Col5a1 were generated and luciferase assays were performed. We also evaluated the role of miRNA using constructs that were mutated at the putative binding sites of miRNA. In addition, we evaluated the endogenous mRNA and protein, and luciferase activity of the Col5a1 gene after miRNA overexpression/knockdown or CRISPR/Cas9-induced knockout. Results: The luci...