The changes of microRNA expression profiles and tyrosinase related proteins in MITF knocked down melanocytes

Abstract

Microphthalmia-associated transcription factor (MITF) is a master regulator in melanocyte proliferation, development, survival and melanoma formation. In melanocyte dysfunction disease, it is observed that the expressions of MITF, tyrosinase (TYR), tyrosinase related protein 1 (TYRP1) and tyrosinase related protein 2 (TYRP2)/dopachrome tautomerase (DCT) are changed, the consequence of which remains unclear. In this study, we focused on the change of microRNA (miRNA) profiles and Tyrosinase Related Proteins (TRPs) in MITF knocked down melanocytes. For the first time, we assayed the MITF-KD miRNA profiles using a miRNA microarray and found that hsa-miR-1225-3p, hsa-miR-634, hsa-miR-197, hsa-miR-766, hsa-miR-574-5p and hsa-miR-328 were upregulated, and hsa-miR-720 and hsa-miR-1308 were downregulated in MITF knocked down melanocytes. These miRNAs were validated by miRNA real time qPCR. These miRNA potential targets, especially the TRPs, were analyzed according to the miRNA database (Sanger Center). By TargetScan prediction, the hsa-miR-634 and hsa-miR-328 have poorly conserved sites on TYR and hsa-miR-197 have poorly conserved sites on TYR1. Through qPCR and western blotting we found that the expression of TYR and TYRP1 were dramatically decreased and the expression of TYRP2 was increased in MITF knocked down melanocytes (MITF-KD). These results suggested that the miRNAs may be involved in MITF regulation of TYR, TYRP1 and TYRP2, which provides a new clue for understanding the role of miRNAs in melanocyte dysfunctional disease.