Essential Role of Adhesion GPCR, GPR123, for Human Pluripotent Stem Cells and Reprogramming towards Pluripotency

Abstract

G-protein-coupled receptors (GPCRs) are the largest family of cell surface receptors. They modulate key physiological functions and are required in diverse developmental processes including embryogenesis, but their role in pluripotency maintenance and acquisition during the reprogramming towards hiPSCs draws little attention. Meanwhile, it is known that more than 106 GPCRs are overexpressed in human pluripotent stem cells (hPSCs). Previously, to identify novel effectors of reprogramming, we performed a high-throughput RNA interference (RNAi) screening assay and identified adhesion GPCR, GPR123, as a potential reprogramming effector. Its role has not been explored before. Herein, by employing GPR123 RNAi we addressed the role of GPR123 for hPSCs. The suppression of GPR123 in hPSCs leads to the loss of pluripotency and differentiation, impacted colony morphology, accumulation of cells at the G2 phase of the cell cycle, and absence of the scratch closure. Application of the GPR123 RNAi at the initiation stage of reprogramming leads to a decrease in the percentage of the “true” hiPSC colonies, a drop in E-cadherin expression, a decrease in the percentage of NANOG+ nuclei, and the absence of actin cytoskeleton remodeling. Together this leads to the absence of the alkaline-phosphatase-positive hiPSCs colonies on the 18th day of the reprogramming process. Overall, these data indicate for the first time the essential role of GPR123 in the maintenance and acquisition of pluripotency.

Retroelement-Age Clocks: Epigenetic Age Captured by Human Endogenous Retrovirus and LINE-1 DNA methylation states

“…In LINE-1-Age, we observed 8 CpGs each located at the L1FLnl_10q26.3e (cg03230038, cg09292354, cg09699225, cg13754569, cg17005927, cg18635896, cg20654619, and cg25120459) and L1FLnl_1p31.3z (cg00318766, cg08413469, cg08825216, cg16166651, cg18167921, cg19634693, cg20322613, cg26064634) LINEs. CpGs in L1FLnl_10q26.3e were downstream of the 3’UTR of the ADGRA1 gene that encodes a G-protein-coupled receptor that has been shown to play an essential role for human pluripotent stem cells and reprogramming towards pluripotency( 36 ). CpGs in L1FLnl_1p31.3z were located in an enhancer region of the DEPDC1 gene that has been shown as a novel cell cycle related gene that regulates mitotic progression and has been shown to inhibit aging-related transcription factor FOXO3 ( 37 , 38 ).…”