Principles of signaling pathway modulation for enhancing human naive pluripotency induction

Jonathan Bayerl, Muneef Ayyash, Tom Shani, Yair Shlomo Manor, Ohad Gafni, Rada Massarwa, Yael Kalma, Alejandro Aguilera-Castrejon, Mirie Zerbib, Hadar Amir, Daoud Sheban, Shay Geula, Nofar Mor, Leehee Weinberger, Segev Naveh Tassa, Vladislav Krupalnik, Bernardo Oldak, Nir Livnat, Shadi Tarazi, Shadi TawilEmilie Wildschutz, Shahd Ashouokhi, Lior Lasman, Varda Rotter, Suhair Hanna, Dalit Ben-Yosef, Noa Novershtern, Sergey Viukov, Jacob H. Hanna

Research output: Contribution to journalArticlepeer-review


Isolating human MEK/ERK signaling-independent pluripotent stem cells (PSCs) with naive pluripotency characteristics while maintaining differentiation competence and (epi)genetic integrity remains challenging. Here, we engineer reporter systems that allow the screening for defined conditions that induce molecular and functional features of human naive pluripotency. Synergistic inhibition of WNT/β-CATENIN, protein kinase C (PKC), and SRC signaling consolidates the induction of teratoma-competent naive human PSCs, with the capacity to differentiate into trophoblast stem cells (TSCs) and extraembryonic naive endodermal (nEND) cells in vitro. Divergent signaling and transcriptional requirements for boosting naive pluripotency were found between mouse and human. P53 depletion in naive hPSCs increased their contribution to mouse-human cross-species chimeric embryos upon priming and differentiation. Finally, MEK/ERK inhibition can be substituted with the inhibition of NOTCH/RBPj, which induces alternative naive-like hPSCs with a diminished risk for deleterious global DNA hypomethylation. Our findings set a framework for defining the signaling foundations of human naive pluripotency.

Original languageEnglish
Pages (from-to)1549-1565.e12
JournalCell Stem Cell
Issue number9
StatePublished - 2 Sep 2021


  • cross-species chimerisim
  • embryonic stem cells
  • extra-embryonic stem cells
  • iPSC
  • naive pluripotency
  • reprogramming


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