Hsa‐miR‐5582‐3P regulatory effect on TGFβ signaling through targeting of TGFβ‐R1, TGFβ‐R2, SMAD3, and SMAD4 transcripts

AbstractTransforming growth factor β (TGFβ) signaling pathway which is regulated by factors such as microRNAs (miRNAs) has pivotal roles in various cellular processes. Here, we intended to verify bioinformatics predicted regulatory effect of hsa‐miR‐5582‐3P against TGFβ/SMAD signaling pathway components. Quantitative reverse‐transcription polymerase chain reaction (RT‐qPCR) analysis indicated a negative correlation of expression between hsa‐miR‐5582‐3P against TGFβ‐R1, TGFβ‐R2, SMAD3, and SMAD4 putative target genes in all of tested cell lines. Also, hsa‐miR‐5582‐3P was significantly downregulated in glioma, breast, and ovarian tumor tissues compared with their normal pairs, detected by RT‐qPCR. Then dual luciferase assay supported direct interaction between this miRNA and TGFβ‐R1, TGFβ‐R2, SMAD3, and SMAD4, 3′ untranslated region sequences. Western blot analysis confirmed negative effect of hsa‐miR‐5582‐3P overexpression on at least TGFβ‐R1 expression. Consistently, hsa‐miR‐5582‐3P overexpression brought about downregulation of TGFβ‐R1, TGFβ‐R2, SMAD3, and SMAD4 expression in HCT‐116 cell line, followed by cell cycle arrest in sub‐G1 phase, detected by flow cytometry. Altogether, our data suggest that hsa‐miR‐5582‐3P reduces the TGFβ/SMAD signaling pathway through downregulation of TGFβ‐R1, TGFβ‐R2, SMAD3, and SMAD4 transcripts. These data introduce hsa‐miR‐5582‐3P as a potential tumor suppressors‐miR and a therapy candidate to be tested in cancers in which TGFβ/SMAD is deregulated.