miR-514a-3p: a novel SHP-2 regulatory miRNA that modulates human cytotrophoblast proliferation

Src homology-2 (SH2) domain containing protein tyrosine phosphatase 2 (SHP-2), encoded by the PTPN11 gene, forms a central component of multiple signalling pathways and is required for insulin-like growth factor (IGF) induced placental growth. Altered expression of SHP-2 is associated with aberrant placental and fetal growth indicating that drugs modulating SHP-2 expression may improve adverse pregnancy outcome associated with altered placental growth. Placental PTPN11/SHP-2 expression is controlled by microRNAs (miRNAs) so SHP-2 regulatory miRNAs may have therapeutic potential, however the individual miRNA(s) regulating placental SHP-2 expression remain to be established. We performed in-silico analysis of 3’UTR target prediction databases to identify libraries of HeLa cells transfected with individual miRNA-mimetics, enriched in potential SHP-2 regulatory miRNAs. Analysis of PTPN11 levels by qPCR revealed that miR-758-3p increased, whilst miR-514a-3p reduced PTPN11 expression. miR-514a-3p and miR-758-3p expression within the human placenta was confirmed by qPCR; miR-514a-3p (but not miR-758-3p) levels inversely correlated with PTPN11 expression. To assess the interaction between these miRNAs and PTPN11/SHP-2, specific mimetics were transfected into first trimester human placental explants and explants cultured for up to 4 days. Overexpression of miR-514a-3p, but not miR-758-3p, significantly reduced PTPN11 and SHP-2 expression. microRNA-ribonucleoprotein complex (miRNP)-associated mRNA assays confirmed that this interaction was direct. miR-514a-3p overexpression attenuated IGF-I induced trophoblast proliferation (BrdU incorporation). miR-758-3p did not alter trophoblast proliferation. These data demonstrate that by modulating SHP-2 expression, miR-514a-3p is a novel regulator of IGF- signalling and proliferation in the human placenta and may have therapeutic potential in pregnancies complicated by altered placental growth.