Germline competency of human embryonic stem cells depends on eomesodermin

Abstract

In humans, germline competency and the specification of primordial germ cells (PGCs) are thought to occur in a restricted developmental window during early embryogenesis. Despite the importance of specifying the appropriate number of PGCs for human reproduction, the molecular mechanisms governing PGC formation remain largely unexplored. Here, we compared PGC-like cell (PGCLC) differentiation from 18 independently derived human embryonic stem cell (hESC) lines, and discovered that the expression of primitive streak genes were positively associated with hESC germline competency. Furthermore, we show that chemical inhibition of TGFβ and WNT signaling, which are required for primitive streak formation and CRISPR/Cas9 deletion of Eomesodermin (EOMES), significantly impacts PGCLC differentiation from hESCs. Taken together, our results suggest that human PGC formation involves signaling and transcriptional programs associated with somatic germ layer induction and expression of EOMES.

Committing the primordial germ cell: An updated molecular perspective

“…Remarkably, this variation was positively correlated with the expression levels of T and EOMES , with higher expression of said factors reflected in enhanced differentiation potential of the clone (Yokobayashi et al, ). This correlation was reproduced in hESCs induced with a 4i‐primed intermediate stage, and additional differentiations with EOMES mutant cells showed drastic reduction in hPGCLC specification, hinting that early mesoderm‐associated transcription factors are necessary at least for the initial step of hPGCLC commitment (Figure ) (Chen et al, ). Exactly how the presence of factors such as T and EOMES engenders permissiveness of BMP‐responding cells remains to be clarified.…”

“…These data robustly demonstrated that SOX17 is necessary and sufficient for hPGC specification, placing it at the top of a genetic hierarchy driving hPGC fate in a competent milieu. Just as importantly, most other reports of in vitro hPGCLC differentiation thus far have independently verified early SOX17 expression, which has now positioned SOX17 as a major molecular footprint of nascent hPGC identity (Chen et al, ; Kobayashi et al, ; Mitsunaga et al, ; Sasaki et al, ; von Meyenn et al, ).…”

“…In humans, PRDM1 is similarly a signature gene for hPGC commitment (Figure ). PRDM1 is expressed in vivo in migratory and gonadal hPGCs (Gkountela et al, ; Gkountela et al, ; Guo et al, ; Tang et al, ), and is also among the earliest germ lineage factors upregulated upon in vitro hPGCLC induction, validating its common usage as a definitive hPGC fate marker (Chen et al, ; Irie et al, ; Irie & Surani, ; Kobayashi et al, ; Mitsunaga et al, ; Sasaki et al, ; Sugawa et al, ; von Meyenn et al, ). Attempts to differentiate PRDM1 knockout/knockdown hESCs into hPGCLCs revealed their inability to adopt germ line fate, and in particular, other germ line genes like NANOS3 were completely uninduced (Irie et al, ; Sugawa et al, ).…”

“…Additionally, their upregulation is observed after the induction of early hPGC factors like SOX17 and PRDM1 , denoting their deployment downstream of the SOX17 / PRDM1 / TFAP2C hPGC specification axis. However, another quintessential pluripotency factor, SOX2 , is virtually absent in hPGCs and hPGCLCs, contrary to its presence and necessity in the murine system (Figure ) (Chen et al, ; de Jong et al, ; Gkountela et al, ; Gkountela et al, ; Guo et al, ; Irie et al, ; Kobayashi et al, ; Mitsunaga et al, ; Sasaki et al, ; Sugawa et al, ; Tang et al, ; von Meyenn et al, ).…”

Evolutionarily diverse mechanisms of germline specification among mammals: what about us?

“…They further provided immunofluorescence staining data supporting expression of EOMES in E11-E15 cynomolgus monkey embryos at the close proximity to locations of PGC specification. Using 18 human ESC clones, a group led by Amander Clark later confirmed that germline competency of human ESCs is also dependent on expression of EOMES (23). Overall, the study of Kojima et al provided critical pieces of information to understand similarities and differences in transcriptional mechanisms involved in PGC specification.…”