Direct Induction Of The Three Pre-implantation Blastocyst Cell Types From Fibroblast.

Mohammad Jaber ¹ Hana Benchetrit ¹ Valery Zayat ¹ Shulamit Baror-Sebban ¹ Avital Pushtt ¹ Kiril Makedonski ¹ Ahmed Radwan ¹ Zvi Zakhaim ¹ Oren Ram ² Tommy Kaplan ³ Yosef Buganim ¹

¹Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, Hebrew University-Hadassah Medical School, Israel
²The Silberman Institute of Life Sciences and the Edmond and Lily Safra Center for Brain Science, Hebrew University of Jerusalem, Edmond J. Safra Campus, Israel
³School of Computer Science and Engineering, Hebrew University of Jerusalem, Israel

Following fertilization, totipotent cells undergo asymmetric cell divisions, resulting in three distinct cell types in the late pre-implantation blastocyst: epiblast (Epi), primitive endoderm (PrE), and trophectoderm (TE). Here, we aim to understand whether these three cell types can be induced from fibroblasts by one combination of transcription factors. By utilizing a sophisticated fluorescent knockin reporter system, we identified a combination of five transcription factors, Gata3, Eomes, Tfap2c, Myc, and Esrrb, that can reprogram fibroblasts into induced pluripotent stem cells (iPSCs), induced trophoblast stem cells (iTSCs), and induced extraembryonic endoderm stem cells (iXENs) , concomitantly. In-depth transcriptomic, chromatin, and epigenetic analyses provide insights into the molecular mechanisms that underlie the reprogramming process toward the three cell types. Mechanistically, we show that the interplay between Esrrb and Eomes during the reprogramming process determines cell fate, where high levels of Esrrb induce a XEN-like state that drives pluripotency and high levels of Eomes drive trophectodermal fate.