Gene Expression Profiles of Melanocytes Over-Expressing miR-5110 in Alpaca

Previous studies have shown that miR-5110 regulates pigmentation by cotargeting melanophilin (MLPH) and WNT family member 1 (WNT1). In order to find the possible molecular mechanism for pigmentation, we examined the mRNA expression profiles in melanocytes of alpaca transfected with miR-5110, inhibitor or negative control (NC) plasmids using high-throughput RNA sequencing. The results showed that a total of 91,976 unigenes were assembled from the reads, among which 13,262 had sequence sizes greater than 2000 nucleotides. According to the KEGG pathway analysis, four pathways related to melanogenesis, the MAPK signaling pathway, Wnt signaling pathway, and cAMP signaling pathway were identified. Compared to the NC, 162 gene were upregulated and 41 genes were downregulated in melanocytes over expressed by miR-5110. The differential expressions of mRNAs Dickkopf 3 (DKK3), premelanosome protein (Pmel), insulin-like growth factor 1 receptor (IGF1R), cyclin-dependent kinase 5 (CDK5), endothelin receptor type B (Ednrb), kit ligand (Kitl), Myc, and S100 were verified using qRT-PCR, which agreed with the results of RNA sequencing. We also verified the differential expressions of mRNAs of some genes in the MAPK signaling pathway using qRT-PCR, which agreed with the results of RNA sequencing. Interestingly, several genes were screened as candidates for the melanogenesis regulated by miR-5110, including Kitl and MAPK-activated protein kinase 3 (MAPKAPK3). These findings provide new insights for further molecular studies on the effects of miR-5110 on the melanogenesis and pigmentation.