De novo recruitment of Polycomb-group proteins in Drosophila embryos

Abstract

Polycomb-group (PcG)-mediated transcriptional repression of target genes can be delineated into two phases. First, following initial repression of target genes by gene-specific transcription factors, PcG proteins recognize the repressed state and assume control of the genes' repression. Second, once the silenced state is established, PcG proteins may maintain repression through an indefinite number of cell cycles. Little is understood about how PcG proteins initially recognize the repressed state of target genes and the steps leading to de novo establishment of PcG-mediated repression. We describe a genetic system in which a Drosophila PcG target gene, giant (gt), is ubiquitously repressed during early embryogenesis by a maternally expressed transcription factor, and show the temporal recruitment of components of three PcG protein complexes: PhoRC, PRC1 and PRC2. We show that de novo PcG recruitment follows a temporal hierarchy in which PhoRC stably localizes at the target gene at least 1 h before stable recruitment of PRC2 and concurrent trimethylation of histone H3 at lysine 27 (H3K27me3). The presence of PRC2 and increased levels of H3K27me3 are found to precede stable binding by PRC1.

Differentiating Drosophila female germ cells initiate Polycomb silencing by regulating PRC2-interacting proteins

“…PcG genes encode two major protein complexes, Polycomb Repressive Complex 1 (PRC1), an E3 ligase that ubiquitylates histone H2A on lysine 119 (H2AK119ub) ( Franke et al, 1992 ; Shao et al, 1999 ; Wang et al, 2004 ), and PRC2, which methylates histone H3 on lysine 27 (H3K27me1/2/3) ( Cao et al, 2002 ; Czermin et al, 2002 ; Kuzmichev et al, 2002 ; Müller et al, 2002 ). In fly embryos, silencing and PRC2-dependent H3K27me3 enrichment on PcG domains first appears as anterior-posterior patterning is being completed during a massive upregulation of zygotic gene expression during cell cycle 14 ( Alhaj Abed et al, 2018 ; Li et al, 2014 ; Pelegri and Lehmann, 1994 ). In preimplantation mouse embryos, H3K27me3 is distributed in a ‘noncanonical’ pattern throughout gene deserts and inactive loci ( Liu et al, 2016 ; Zheng et al, 2016 ) and is not further enriched on its canonical sites: CpG-rich ‘islands’ (CGIs) around enhancers and promoters of developmentally-induced genes ( Ku et al, 2008 ; Tanay et al, 2007 ).…”

Polycomb-group recruitment to a Drosophila target gene is the default state that is inhibited by a transcriptional activator

“…The maternally expressed gt activator, Cad, also is ubiquitously expressed (due to the lack of Bcd), yet gt is initially repressed in all syncytial blastoderm nuclei by Hb (fig. S1B) ( 21 ). Bcd and the terminal system, which includes Tsl, are needed to activate zygotic hb transcription ( 26 , 27 ).…”

“…Therefore, as maternally expressed Hb and Cad are degraded [after nc13; ( 27 , 30 )], they are not replaced by zygotically expressed Hb and Cad due to the bcd osk tsl maternal genotype (fig. S1B) ( 21 ). We have previously shown that both Hb and Cad bind to the gt_(-3) enhancer in these embryos, resulting in ubiquitous gt repression at the syncytial blastoderm stage and that the PcG is required to maintain gt repression when maternal Hb is degraded ( 21 , 31 ).…”

“…S1B) ( 21 ). We have previously shown that both Hb and Cad bind to the gt_(-3) enhancer in these embryos, resulting in ubiquitous gt repression at the syncytial blastoderm stage and that the PcG is required to maintain gt repression when maternal Hb is degraded ( 21 , 31 ). Of the four gt enhancers, we will focus only on gt_(-3) and therefore will simply refer to it as “enhancer.”…”

“…We have experimentally tested these models using a previously described genetic system in which a PcG target gene, giant ( gt ), is ubiquitously repressed in Drosophila embryos ( 21 ). Here, we describe a modified version of this system in which gt is ubiquitously active and compare recruitment of PcG proteins and distribution of histone modifications under these disparate conditions.…”