Drosophila Pgc protein inhibits P-TEFb recruitment to chromatin in primordial germ cells

Abstract

Germ cells are the only cells that transmit genetic information to the next generation, and they therefore must be prevented from differentiating inappropriately into somatic cells 1 . A common mechanism by which germline progenitors are protected from differentiation-inducing signals is a transient and global repression of RNA polymerase II (RNAPII)-dependent transcription 1 . In both Drosophila and Caenorhabditis elegans embryos, the repression of messenger RNA transcription during germ cell specification correlates with an absence of phosphorylation of Ser 2 residues in the carboxy-terminal domain of RNAPII (hereafter called CTD) 2 , a critical modification for transcriptional elongation 3 . Here we show that, in Drosophila embryos, a small protein encoded by polar granule component (pgc) is essential for repressing CTD Ser 2 phosphorylation in newly formed pole cells, the germline progenitors. Ectopic Pgc expression in somatic cells is sufficient to repress CTD Ser 2 phosphorylation. Furthermore, Pgc interacts, physically and genetically, with positive transcription elongation factor b (P-TEFb), the CTD Ser 2 kinase complex, and prevents its recruitment to transcription sites. These results indicate that Pgc is a cell-type-specific P-TEFb inhibitor that has a fundamental role in Drosophila germ cell specification. In C. elegans embryos, PIE-1 protein segregates to germline blastomeres, and is thought to repress mRNA transcription through interaction with P-TEFb 4-7 . Thus, inhibition of P-TEFb is probably a common mechanism during germ cell specification in the disparate organisms C. elegans and Drosophila. pgc RNA, a component of the Drosophila germ plasm 8 , has been implicated in the repression of CTD Ser 2 phosphorylation in early pole cells 9,10 . However, the mechanism underlying pgc-mediated transcriptional repression has been unknown. Initial characterization of its nucleotide sequence suggested that pgc RNA acts as a non-coding RNA 8 . Nevertheless, we have noticed that an AUG triplet, beginning at nucleotide 117 of the 0.7-kilobase (kb) transcript, is in a favourable context to serve as a translation initiation site 8 . Completion of the Drosophila genomic sequence revealed an error in our original manual sequencing in a region where a strong stem structure can form (an additional C exists between nucleotides 178 and 179). In the revised pgc sequence, the open reading frame (ORF) beginning at nucleotide 117 is capable of encoding a 71-amino-acid polypeptide (Fig. 1a). We found that this polypeptide

7SK snRNP/P-TEFb couples transcription elongation with alternative splicing and is essential for vertebrate development

“…In addition, extensive developmental defects may stem from the derepression of P-TEFb, which could in turn promote expression of otherwise silent developmental control genes that may contain paused RNAPII molecules. Importantly, transient and global inhibition of P-TEFb has been proposed to be essential for maintaining germ cell identity in C. elegans (30,31) and Drosophila (32), and transcription elongation controls cell fate specification in the Drosophila embryo (33). We therefore speculate that 7SK snRNP disintegration would lead to inappropriate differentiations and loss of identities of germ cells and multiple somatic cell lineages, causing developmental defects observed here.…”

Hidden Peptides Encoded by Putative Noncoding RNAs

“…An RNA molecule, designated pgc, was found to be localized to polar granules and was originally identified as an untranslatable RNA on the basis of the absence of conserved ORFs for products of >100 amino acids (Nakamura et al, 1996). However, completion of the Drosophila genomic sequence revealed an error in the original pgc sequence and thereby led to the identification of an ORF encoding a 71-amino acid polypeptide that was highly conserved among 12 Drosophila species (Hanyu-Nakamura et al, 2008). Immunostaining with antibodies to the Pgc polypeptide revealed its localization in pole cells.…”

Primordial Germ Cells in Mice

“…As a mechanism to inhibit transcriptional elongation, PIE-1 appears to inhibit the positive transcriptional elongation factor b (P-TEFb), which promotes the transcriptional elongation activity of RNAPII by phosphorylating the Ser-2 of the carboxy-terminal domain (CTD) of RNAPII (Zhang et al 2003). On the other hand, the polar granule component, which is a short, 71-amino-acid protein conserved onlyamong Drosophila species, inhibits the recruitment of P-TEFb to transcriptional sites (Hanyu-Nakamura et al 2008). In contrast, in mice, germ cell specification requires active transcription, but the transcriptional repressor BLIMP1 specifically shuts off gene expression for a somatic mesodermal program ) (see main text).…”