Abstract
Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) hold great promise for regenerative medicine, disease treatment, and organ transplantation. As the ethical issue of human ESCs and similarity of pig in human genome and physiological characteristics, the porcine iPSCs (piPSCs) have become an ideal alternative study model. N6-methyladenosine (m6A) methylation is the most prevalent modification in eukaryotic mRNAs, regulating the self-renewal and differentiation of pluripotency stem cells. However, the explicit m6A-regulating machinery remains controversial. Here, we demonstrate that m6A modification and its modulators play a crucial role in mediating piPSCs pluripotency. In brief, loss of METTL3 significantly impairs self-renewal and triggers differentiation of piPSCs by interfering JAK2 and SOCS3 expression, further inactivating JAK2–STAT3 pathway, which then blocks the transcription of KLF4 and SOX2. We identify that both of JAK2 and SOSC3 have m6A modification at 3′UTR by m6A-seq analysis. Dual-luciferase assay shows that METTL3 regulates JAK2 and SOCS3 expression in an m6A-dependent way. RIP-qPCR validates JAK2 and SOCS3 are the targets of YTHDF1 and YTHDF2, respectively. SiMETTL3 induced lower m6A levels of JAK2 and SOCS3 lead to the inhibition of YTHDF1-mediated JAK2 translation and the block of YTHDF2-dependent SOCS3 mRNA decay. Subsequently, the altered protein expressions of JAK2 and SOCS3 inhibit JAK2–STAT3 pathway and then the pluripotency of piPSCs. Collectively, our work uncovers the critical role of m6A modification and its modulators in regulating piPSCs pluripotency and provides insight into an orchestrated network linking the m6A methylation and SOCS3/JAK2/STAT3 pathway in pluripotency regulation.
Highlights
Embryonic stem cells (ESCs) offer great hope for regenerative medicine, organ transplantation, and drug development
METTL3 is required for pluripotent stem cells (piPSCs) self-renewal and pluripotency We first examined the m6A methyltransferase METTL3 expression of piPSCs in retinoic acid (RA)-induced differentiation and revealed a gradual decrease in METTL3 levels (Fig. 1a)
Liquid chromatographytandem mass spectrometry (LC-MS/MS) analysis of global m6A level in purified mRNA from cells with or without METTL3 knockdown showed that METTL3 ablation leads to a significant reduction (~ 80%) of m6A on mRNA (Fig. 1d), confirming the methylation activity of METTL3 in piPSCs
Summary
Embryonic stem cells (ESCs) offer great hope for regenerative medicine, organ transplantation, and drug development. These cells provide a powerful model system for studies of cellular identity and early mammalian development[1]. Page 2 of 15 171 porcine induced pluripotent stem cells (piPSCs) become an ideal alternative resource, which holds unprecedented promise for human regenerative medicine, disease treatment, and organ transplantation. The mechanisms of porcine embryonic development and the pluripotent regulation network remain largely unknown. This dynamic epigenetic modification has been demonstrated to regulate RNA stability, translation, alternative splicing, and nuclear export[11,12,13,14]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
