Experimental evidences for hsa-miR-497-5p as a negative regulator of SMAD3 gene expression

The SMAD family comprises of transcription factors that function as signal transducers of transforming growth factor (TGFβ) superfamily members. MiRNAs are a class of small noncoding RNAs that may play a major role in post transcriptional regulation of SMAD genes. Here, we intended to investigate if hsa-miR-497-5p is capable of regulating SMAD3 gene expression. Hsa-miR-497-5p was bioinformatically predicted as a candidate regulator of SMAD3 gene expression and then, hsa-miR-497-5p expression status was analyzed in different cell lines using RT-qPCR. Overexpression of hsa-miR-497-5p in HEK293t cells resulted in downregulation of SMAD3 which was detected by RT-qPCR and western analysis. Further, dual luciferase assay results supported direct interaction of hsa-miR-497-5p with 3'-UTR sequences of SMAD3 transcript. Overexpression of hsa-miR-497-5p in HEK293t cells resulted in cell cycle arrest in G0/G1 phase, detected by flow cytometry. Overall, accumulative results indicated that hsa-miR-497-5p by targeting SMAD3 is potentially one of the regulators of the TGFβ signaling pathway.