Heterochromatin Protein 1A Collaborates with Polycomb/Trithorax Group Proteins to Regulate Drosophila Transcription

Maintenance of the integrity of the genome is vital to survival and the small non-histone protein Heterochromatin protein 1 (HP1) plays an important role in the process. Here we demonstrate two roles for HP1 in regulating transcription in the early Drosophila embryo. HP1, together with its telomeric binding partner, HOAP, play a role in regulating the Drosophila sex determination pathway. Specifically, these proteins regulate the critical decision in this pathway, firing of the establishment promoter of the master switch gene, Sex-lethal (Sxl). Female-specific activation of this promoter, SxlPe, is essential to females as it provides SXL protein to initiate the autoregulatory feedback loop, which ensures productive female-specific splicing of the Sxl transcripts that are transcribed from the maintenance promoter (SxlPm), which is active in both sexes. HOAP mutants show inappropriate SxlPe expression in males and incorrect splicing of SxlPm-derived transcripts, while females show premature activation of SxlPe. These results implicate HOAP in regulating the promoter. HP1 mutants, by contrast, display SxlPm splicing defects in both sexes, suggestive of inappropriate expression of SxlPe in both sexes. Chromatin immunoprecipitation assays show both proteins are associated with SxlPe sequences. In embryos from HP1 mutant mothers and Sxl mutant fathers, female viability and RNA polymerase II (RNAPII) recruitment to SxlPe are severely compromised. Our genetic and biochemical assays indicate a repressing activity for HOAP and both activating and repressing roles of HP1 at SxlPe. These findings prompted us to further investigate the role of chromatin modifying proteins in regulating the sex determination decision. Consistent with important role for HP1 in regulating transcription, we also find that HP1 collaborates with Polycomb/trithorax Group (PcG/trxG) proteins to regulate gene expression in the early embryo. PcG/trxG proteins were first identified in Drosophila as repressors (PcG) and activators (trxG) of the homeotic genes, serving to pattern the body axis. Mutations in PcG/trxG genes can antagonize or synergize with those in the HP1 chromatin modifying system, suggesting the two systems work together to set the chromatin state and influence transcription. These proteins regulate the chromatin by placing either repressive H3K27me3 (PcG) or activating H3K4me3 (trxG) marks. In early embryogenesis, we find that promoters have both H3K27me3/H3K4me3 marks suggesting that they are bivalent. Both marks appear to be dependent on HP1 in the early embryo, which is not the case in later larval stages. We speculate that as the early embryo organizes its genome into heterochromatin and euchromatin, a transition between the HP1/H3K9me silencing system to the PcG/H3K27me3 system takes place. This transition is sensitive to changes in HP1. This work reveals a novel complex process in which two distinct chromatin systems collaborate to regulate gene expression.