Transcriptome Comparison Between the Cultured and In Vivo Chick Primordial Germ Cells by SMART ‐Seq‐Based Single‐Cell RNA Sequencing

Primordial germ cells (PGCs), the precursors of the germline, have unique cellular characteristics to undergo long-distance migration to the embryonic gonads and have the potential to differentiate into somatic cells. Among the animal models studying PGC development, the chicken PGCs are an ideal model, since it is a rare model in which long-term PGC cultivation is applicable. Although the cultural applicability of chicken PGC makes it attractive for revealing the PGC character and its developmental processes, some differences from in vivo PGCs are known, such as the remarkable upregulation of cell proliferation and a lesser ability to reach the gonads. Understanding these differences at the molecular level is crucial. To this end, we first performed SMART-seq-based single-cell RNA sequencing to compare transcriptomes between in vivo PGCs and cultured PGCs. Our results revealed that PGC cultivation causes a shift from a MYC-dependent to a MYCN-dependent gene regulatory network (GRN) in PGCs, suggesting that this reprogramming contributes to the acquisition of proliferation ability and stem cell characteristics in cultured PGCs. Additionally, our results suggest that the MYCN-dependent GRN increases the risk of somatic differentiation, particularly in neural fate, in cultured PGCs. In addition, our transcriptome analysis identified a new cell population that shows molecular characteristics of germline-biased undifferentiated cells. Thus, our study provides fundamental molecular information to understand both the effects of PGC cultivation and the developmental process of chicken PGCs.