title: Drosophila O-GlcNAcase Deletion Globally Perturbs Chromatin O-GlcNAcylation.
abstract: Gene expression during Drosophila development is subject to regulation by the Polycomb (Pc), Trithorax (Trx), and Compass chromatin modifier complexes. O-GlcNAc transferase (OGT/SXC) is essential for Pc repression suggesting that the O-GlcNAcylation of proteins plays a key role in regulating development. OGT transfers O-GlcNAc onto serine and threonine residues in intrinsically disordered domains of key transcriptional regulators; O-GlcNAcase (OGA) removes the modification. To pinpoint genomic regions that are regulated by O-GlcNAc levels, we performed ChIP-chip and microarray analysis after OGT or OGA RNAi knockdown in S2 cells. After OGA RNAi, we observed a genome-wide increase in the intensity of most O-GlcNAc-occupied regions including genes linked to cell cycle, ubiquitin, and steroid response. In contrast, O-GlcNAc levels were strikingly insensitive to OGA RNAi at sites of polycomb repression such as the Hox and NK homeobox gene clusters. Microarray analysis suggested that altered O-GlcNAc cycling perturbed the expression of genes associated with morphogenesis and cell cycle regulation. We then produced a viable null allele of oga (oga(del.1)) in Drosophila allowing visualization of altered O-GlcNAc cycling on polytene chromosomes. We found that trithorax (TRX), absent small or homeotic discs 1 (ASH1), and Compass member SET1 histone methyltransferases were O-GlcNAc-modified in oga(del.1) mutants. The oga(del.1) mutants displayed altered expression of a distinct set of cell cycle-related genes. Our results show that the loss of OGA in Drosophila globally impacts the epigenetic machinery allowing O-GlcNAc accumulation on RNA polymerase II and numerous chromatin factors including TRX, ASH1, and SET1.
authors.affiliation: From the NIDDK and.
authors.forename: Ilhan
authors.initials: I
authors.lastname: Akan
authors.affiliation: NCI,National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: Dona C
authors.initials: DC
authors.lastname: Love
authors.affiliation: From the NIDDK and.
authors.forename: Katryn R
authors.initials: KR
authors.lastname: Harwood
authors.affiliation: From the NIDDK and.
authors.forename: Michelle R
authors.initials: MR
authors.lastname: Bond
authors.affiliation: From the NIDDK and jah@helix.nih.gov.
authors.forename: John A
authors.initials: JA
authors.lastname: Hanover
mesh_terms: D000117:Acetylglucosamine / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D002843:Chromatin / Q000235:genetics / Q000378:metabolism*; D004330:Drosophila / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D044127:Epigenesis, Genetic / Q000235:genetics*; D047468:Immunoprecipitation; D017351:N-Acetylglucosaminyltransferases / Q000235:genetics*; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017384:Sequence Deletion; D059467:Transcriptome
chemical_list: D002843:Chromatin; D012333:RNA, Messenger; D017351:N-Acetylglucosaminyltransferases; C449880:O-GlcNAc transferase; D000117:Acetylglucosamine
keywords: Ash1; Drosophila; O-GlcNAcylation; O-linked N-acetylglucosamine (O-GlcNAc); RNA polymerase II; SET1; Trithorax; cell cycle; chromatin; polycomb