Regulation of WDFY1 Expression by miRNAs, Transcription Factors, and IL-6 in Murine Mesangial Cells

WD40 repeat and FYVE containing protein 1 (WDFY1) functions in membrane trafficking and protein complex scaffolding. WDFY1 has been studied in the immune system and in different oncogenic conditions. Therefore, comprehensive understanding of WDFY1 regulation mechanisms is much desired. In this study, we analyzed the promoter and 5'- and 3'-untranslated regions (UTRs) of wdfy1 and identified critical sequence elements, transcription factors (TFs), and miRNAs that collaboratively regulate wdfy1 gene expression. A 3.5 kb segment of the mouse wdfy1 promoter and 5'-UTR was cloned into a luciferase expression vector and transfected into HeLa cells. Luciferase assays of promoter deletion mutants revealed approximately four-fold increased activity attributed by a 500 bp distal fragment upstream of the wdfy1 5'-UTR. Four TFs (Sp1, Ap-1, Hes1, and TCF7) were found to be critical for wdfy1 expression with binding sites spread throughout the promoter and 5'-UTR regions. Cloning of a 3.2 kb fragment of wdfy1 3'-UTR into the luciferase expression vector led to an ~3.5-fold decrease in luciferase activity. Complementary siRNA and luciferase assays mutually confirmed our findings. Most importantly, IL-6, a critical cytokine in organ inflammation, was found to promote WDFY1 expression through the upregulation of Sp1 in primary renal mesangial cells. We, therefore, identified a potential inflammation-driven WDFY1 upregulation in mice.