Single-Cell Landscape of Transcriptional Heterogeneity and Cell Fate Decisions during Mouse Early Gastrulation

Abstract

SummaryThe mouse inner cell mass (ICM) segregates into the epiblast and primitive endoderm (PrE) lineages coincident with implantation of the embryo. The epiblast subsequently undergoes considerable expansion of cell numbers prior to gastrulation. To investigate underlying regulatory principles, we performed systematic single-cell RNA sequencing (seq) of conceptuses from E3.5 to E6.5. The epiblast shows reactivation and subsequent inactivation of the X chromosome, with Zfp57 expression associated with reactivation and inactivation together with other candidate regulators. At E6.5, the transition from epiblast to primitive streak is linked with decreased expression of polycomb subunits, suggesting a key regulatory role. Notably, our analyses suggest elevated transcriptional noise at E3.5 and within the non-committed epiblast at E6.5, coinciding with exit from pluripotency. By contrast, E6.5 primitive streak cells became highly synchronized and exhibit a shortened G1 cell-cycle phase, consistent with accelerated proliferation. Our study systematically charts transcriptional noise and uncovers molecular processes associated with early lineage decisions.

Cell Lineage Specification at Single Cell Resolution

“…These data suggest that there is possibility of increase transcriptional noise before lineage commitment as also observed in other systems [7,9].…”

“…Consistent with the in vivo phenomenon, female naïve ESCs also have two activated X chromosomes whereas one X chromosome is inactivated in female EpiSCs [6]. The new single cell transcriptome data confirmed the previous observation, moreover, they found strong association between X reactivation and Pou5f1 and one of its interacting partner Zfhx3 at E3.5; while increased expression of Dnmt3a and Zfp57 is associated with the following X inactivation, which will provide further insights into the mechanism of X reactivation and inactivation [7].…”

“…From E5.5 to E6.5, the cells begin to exit pluripotency and undergo lineage commitment. The data revealed that at E5.5, though some genes associated with anterior-posterior polarity and the primitive streak, no apparent substructure or expression of primitive streak markers were expressed, whereas at E6.5, four subgroups were identified as mentioned above, in EPI, the expression of Otx2, a gene associated with exit from pluripotency increased, along with several polycomb genes, which suggests the importance of polycomb complex in establishing transcriptional control in the non-committed epiblast cells [7].…”

“…The authors' data clearly showed genes enriched for EPI and PE respectively at E4.5; at E3.5, though subsets of EPI and PE genes were expressed, no lineage subgroups were observed. Besides identifying genes such as Fgf4 and Fgfr2 known to derive EPI and PE specification, they also found Pdgfra, Top2b, Sox17, Gata4 and Pdk2 was associated with PE fate, and Morc1, Nanog, Dppa5 and Pdpn was associated with EPI fate [7]. From E5.5 to E6.5, the cells begin to exit pluripotency and undergo lineage commitment.…”

“…To open the black box, now Mohammed et al performed single cell RNA sequencing of ICM of E3.5, EPI and PE at E4.5, E5.5 and E6.5. E3.5 cells expressed high level of pluripotency genes; at E4.5, EPI and PE was clearly separated; at E5.5, naïve pluripotency gene Nanog was down-regulated accompanied by increased primed pluripotency maker Pou3f1; at E6.5, the embryo cluster into four groups, visceral endoderm (from PE), a primitive streak population and two sub clusters of non-committed epiblast cells [7].…”