(73) CLDN10 Drives Lineage Decision in Amnion/Primordial Germ Progenitor Cell Population at the Amnion-Epiblast Boundary of the Primate Peri-gastrula

Nikola Sekulovski¹, Amber E. Carleton¹, Anusha Rengarajan¹, Chien-Wei Lin², Maliha Kabir¹, Lauren N. Juga¹, Allison E. Whorton¹, Jenna K. Schmidt³, Thaddeus G. Golos^3,4,5, and Kenichiro Taniguchi<sup>1

1</sup>Department of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, WI 53226, USA

²Division of Biostatistics, Institute for Health and Equity, Medical College of Wisconsin, Milwaukee, WI 53226, USA

³Wisconsin National Primate Research Center (WNPRC), Madison, WI, USA

⁴Department of Obstetrics and Gynecology, University of Wisconsin - Madison School of Medicine, Madison, WI USA

⁵Department of Comparative Biosciences, University of Wisconsin - Madison School of Veterinary Medicine, Madison, WI, USA

Abstract Text:

Formation of amnion (a component of the fetal membrane) and primordial germ cells (PGC, precursor to oocyte and sperm) are seen within two weeks after implantation is initiated in humans, and are critical for continuation of pregnancy and for successful reproduction, respectively. Strikingly, a growing body of evidence from primate embryos, as well as from human pluripotent stem cell (hPSC)-derived in vitro systems, supports the notion that amnion and PGC share a common precursor. To gain comprehensive insights into this critical but poorly understood process, we examined the evolving transcriptome of a developing hPSC-derived model of amnion and PGC formation at the single cell level. Our data revealed contiguous amniogenic cell states: pluripotency-exiting, early progenitor, late progenitor, specified and maturing, each of which shows transcriptomic similarities to distinct cell types in a Carnegie stage 7 human embryo. We identified a cohort of markers specific to each of these amnion-lineage progressing states and validate selected markers for their expression in the amnion of implanted cynomolgus macaque (Macaca fascicularis) embryos. Additionally, the data unveiled a previously unrecognized CLDN10⁺ progenitor state that displays bi-potential characteristics for amnion and PGC-like cell lineages. Strikingly, we found that expression of CLDN10 is restricted to the amnion-epiblast boundary region in the human post-implantation amniotic sac embryoid model, as well as in peri-gastrula cynomolgus macaque embryos. Moreover, this boundary region displays amnion/PGC progenitor-like transcriptional characteristics – CLDN10⁺ cells are positive for amnion (TFAP2A) and PGC markers (TFAP2C). Furthermore, loss of function analysis using our stem cell model shows that CLDN10 promotes amniotic but suppresses PGC-like fate. Overall, based on the single cell transcriptomic resource in this study, we identified a CLDN10⁺ amnion/PGC progenitor-like population at the amnion-epiblast boundary of the primate peri-gastrula, and present additional molecular clues as to how amnion and PGC may be formed.