Mapping The Epigenomics Of Chromodomain Proteins In C.elegans

Epigenetic regulation of histone post-translational modifications is critical for genome stability,gene expression and other fundamental biological processes.It is related to aging and lifespan,cancer,and neural pathologies.This area shows enormous potentials for application in human health and medicines customized for individual people.Histone modifications are dynamic and reversible,which can be regulated by"writers","erasers",and "readers".Modifications are deposited by writers and removed by erasers.Histone readers specifically recognize histone modifications by their conserved domains.They interpret information of the "histone code" and mediate functional outcomes by recruiting and interacting with other epigenetic effectors.Chromodomain is an evolutionally conserved motif across eukaryotic species,which functions as a histone methylation reader and recognizes H3K4me,H3K9me,H3K27me,etc.Previous works have revealed that chromodomain proteins,such as HP1 and Pc,played important roles in epigenetic regulation.However,the function and regulating mechanisms of most chromodomain proteins in vivo are still unclear.Here,by combining CRISPR/Cas9 gene-editing,ChIP-seq analysis,fluorescence imaging,genetic screen,and in vitro biochemistry,we mapped the epigenomics of chromodomain proteins in the model organism,C.elegans.Firstly,we systematically mapped the expression and chromatin binding signatures of chromodomain proteins.We used CRISPR/Cas9 and constructed fluorescently tagged transgenic strains by knocking in GFP tag in situ to chromodomain genes.By ChIP-seq data analysis,we characterized putative histone modifications recognized by chromodomain proteins,and delineated an epigenetic regulatory network in which chromodomain proteins participated.Secondly,we generated a comprehensive sub-cellular localization map of chromodomain proteins by fluorescence imaging.Employing sub-cellular localization as a tool,we studied function and regulation of two chromodomain proteins,CEC-5 and UAD-2,in detail.We utilized chromodomain protein sub-cellular localizations as reporters and performed a RNAi screen based on epigenetic factor candidates.MET-2,an H3K9me1/2 writer,and UBC-9,UBA-2,two SUMOylation factors,are required for nuclear puncta formed by CEC-5.CEC-5 is an H3K9mel/2 reader,which is proved by ChIP and in vitro biochemical assays.MET-2 is required for CEC-5 association with heterochromatin.Furthermore,a forward genetic screen via chemical mutagenesis identified a conserved Arginine124 of CEC-5’s chromodomain,which is essential for CEC-5’s association with heterochromatin.CEC-5 is required for the lifespan regulation of C.elegans.Thus,our work shows an evidence of H3K9me1/2 regulating lifespan.piRNA genes typically localize to clusters on the genome.UAD-2,an H3K27me3 reader,promotes piRNA cluster transcription together with SNAPc and USTC complexes.In this work,we revealed that SUMOylation pathway suppressed piRNA production by inhibiting binding of UAD-2 to piRNA clusters upon heat stress.Additionally,another H3K9me1/2 reader,CEC-8,responds to heat stress and accumulates to nucleolus.Interestingly,in the process of studying environmental stress,we accidently found that starvation induced trans-splicing of translation related genes,which is mediated by SNAPc complex and TOFU-5.SNAPc complex and TOFU-5 may activate the response by promoting transcription of SL RNAs.Together,our work will not only serve as a resource to explore chromodomain proteins,but also elaborate the critical function of chromodomain proteins in aging,piRNA expression,and stress response.Our findings will lay the foundations for further investigation of chromodomain in eukaryotes and provide new insights into epigenetic regulation network of histone modifications and human diseases.