In animals, germ cells are often distinguished from somatic lineages at the earliest stages of embryogenesis. In some organisms, germ blastomeres appear to enter a state of transcriptional quiescence. For example, in the worm Caenorhabditis elegans, transcription is activated in somatic blastomeres as early as the 4-cell stage, whereas it is not initiated in germline blastomeres until the 100-cell stage. This transcriptional repression in germ blastomeres has been attributed to the PIE-1 protein, specifically localized in these cells from the first embryonic division. PIE-1 is thought to inhibit the activity of CDK-9, a cyclin-dependent kinase previously considered essential for the phosphorylation of serine 2 (CTD-Ser2) of the C-terminal domain (CTD) of RNA polymerase II and for transcription elongation. However, recent studies, showing that embryogenesis proceeds normally in a mutant strain expressing a CTD in which serines 2 is replaced by an alanine (CTD-S2A) and identifying CDK-12 as the main kinase phosphorylating CTD-Ser2, call this model into question.
To study the transcriptome of germline blastomeres in the worm C. elegans, an approach combining cell sorting and RNA sequencing (RNA-seq) was developed. Pilot analyses validated this method on a wild-type strain, enabling its use on a strain in which PIE-1 can be specifically degraded using the Auxin-Inducible Degron (AID) system. This made it possible to examine the effect of PIE-1 depletion on the transcriptome of germline blastomeres revealing that in its absence, germline blastomeres adopt a transcriptional profile close to that of somatic blastomeres, confirming the fundamental role of PIE-1 in preserving germline identity during embryogenesis.
In parallel, the fission yeast Schizosaccharomyces pombe was used to analyze the consequences of PIE-1 expression in a heterologous organism. The results showed that PIE-1 by localizing near transcription termination sites induces further transcription by RNA polymerase II beyond the termination site, leading to transcription of intergenic regions.
These observations led to the hypothesis that in C. elegans,within germinal blastomeres, PIE-1 might regulate alternative polyadenylation in 3' untranslated regions, producing longer RNA isoforms susceptible to degradation. In the absence of PIE-1, shorter isoforms could be generated, allowing accumulation of somatic transcripts and potentially degradation of maternal mRNAs via somatic protein translation.
Although further investigations are required in C. elegans to validate this hypothesis, it provides an innovative conceptual framework for understanding the role of PIE-1, independent of CTD-Ser2 phosphorylation.
