Characterization of factors that underlie transcriptional silencing inC. elegansoocytes

Abstract

While it has been appreciated for decades that prophase-arrested oocytes are transcriptionally silenced, the molecular pathway that promotes silencing has remained elusive. Here, using C. elegans as a model system, we demonstrate that the global chromatin compaction (GCC) pathway is responsible for genome silencing in both oocytes and spermatocytes. GCC is mediated by a collaboration between topoisomerase II, condensin II, and the H3K9me pathway. For oocytes, we further demonstrate that GCC is under cell cycle control such that it is activated specifically during diakinesis and prior to oocyte maturation. Previous work had shown that GCC silences gene expression in the primordial germ cells (PGCs) of nutrient-deprived L1 larvae, and we demonstrate here that it is also used during gametogenesis, thus showing that GCC is a versatile, signal-mediated genome silencing system.

The condensin II/TOP-2 axis silences transcription during germline specification inC. elegans

“…Previous work from our laboratory had defined the GCC pathway as a signal-mediated genome silencing system (Belew et al, 2021;2022), and thus we were interested in determining if GCC was active in the P-cells of early embryos, which are known to be transcriptionally repressed (Robert et al, 2015, Wang and Seydoux, 2013). To do so, we stained early embryos with an antibody that detects a phospho-serine 2 on the RNAPII CTD (referred to as RNAPIIpSer2 for the remainder of this study).…”

“…To do so, we stained early embryos with an antibody that detects a phospho-serine 2 on the RNAPII CTD (referred to as RNAPIIpSer2 for the remainder of this study). The presence of RNAPIIpSer2 signal has been used as a marker for actively elongating RNA polymerase II (Palancade and Bensaude, 2003), and we and others have used this approach extensively to identify transcriptionally active nuclei within a variety of C. elegans tissues (Dunn and Seydoux, 1997; Walker et al, 2004; Wong et al, 2018; Belew et al, 2021;2022). The antibody that we are using, a rabbit polyclonal that recognizes RNAPIIpSer2, has previously been validated by us for use in C. elegans embryos (Belew et al, 2021).…”

“…In fact, genome repression and activation are recurring events during germline development and gametogenesis in the roundworm C. elegans . In meiotic prophase, for example, both oocytes and spermatocytes globally repress transcription as they prepare for the meiotic divisions (Walker et al, 2007; Belew et al, 2022). In both cases transcription is repressed by a global chromatin compaction (GCC) pathway (Belew et al, 2021; 2022), comprised of topoisomerase II, condensin II, and components of the H3K9me system.…”

“…In meiotic prophase, for example, both oocytes and spermatocytes globally repress transcription as they prepare for the meiotic divisions (Walker et al, 2007; Belew et al, 2022). In both cases transcription is repressed by a global chromatin compaction (GCC) pathway (Belew et al, 2021; 2022), comprised of topoisomerase II, condensin II, and components of the H3K9me system. After fertilization, transcription remains repressed, but at this time repression is mediated by the OVA-1/2 proteins that bind to and inactivate the essential transcription factor TAF-4 (Guven-Ozkan et al, 2010).…”

“…How PIE-1 blocks serine 5 phosphorylation, mechanistically, is not understood. In this work we asked if the GCC pathway also plays a role in silencing transcription in P-cells, as it does in oocytes, spermatocytes, and the PGCs of starved L1s (Belew et al, 2021,2022). We find that the GCC per se is not required, as we found no role for the H3K9me pathway in silencing P-cell transcription.…”