O-GlcNAcase Is an RNA Polymerase II Elongation Factor Coupled to Pausing Factors SPT5 and TIF1β.
We describe here the identification and functional characterization of the enzyme O-GlcNAcase (OGA) as an RNA polymerase II elongation factor. Using in vitro transcription elongation assays, we show that OGA activity is required for elongation in a crude nuclear extract system, whereas in a purified system devoid of OGA the addition of rOGA inhibited elongation. Furthermore, OGA is physically associated with the known RNA polymerase II (pol II) pausing/elongation factors SPT5 and TRIM28-KAP1-TIF1β, and a purified OGA-SPT5-TIF1β complex has elongation properties. Lastly, ChIP-seq experiments show that OGA maps to the transcriptional start site/5' ends of genes, showing considerable overlap with RNA pol II, SPT5, TRIM28-KAP1-TIF1β, and O-GlcNAc itself. These data all point to OGA as a component of the RNA pol II elongation machinery regulating elongation genome-wide. Our results add a novel and unexpected dimension to the regulation of elongation by the insertion of O-GlcNAc cycling into the pol II elongation regulatory dynamics.
From the Transcriptional Regulation and Biochemistry Unit, Metabolism Branch, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 30893.
Melissa
M
Resto
Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, Maryland 21702.
Bong-Hyun
BH
Kim
From the Transcriptional Regulation and Biochemistry Unit, Metabolism Branch, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 30893.
Alfonso G
AG
Fernandez
Bioinformatics Program, Boston University, Boston, Massachusetts 02215, and.|Laboratory of Epigenome Biology, NHLBI, National Institutes of Health, Bethesda, Maryland 20892.
Brian J
BJ
Abraham
Laboratory of Epigenome Biology, NHLBI, National Institutes of Health, Bethesda, Maryland 20892.
Keji
K
Zhao
From the Transcriptional Regulation and Biochemistry Unit, Metabolism Branch, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 30893, phdbiochemist@gmail.com.
Brian A
BA
Lewis
D000951:Antigens, Neoplasm / Q000737:chemistry* / Q000378:metabolism; D051548:Histone Acetyltransferases / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D006821:Hyaluronoglucosaminidase / Q000737:chemistry* / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry* / Q000378:metabolism; D061805:Transcription Elongation, Genetic / Q000502:physiology; D035602:Transcriptional Elongation Factors / Q000737:chemistry* / Q000378:metabolism; D000076183:Tripartite Motif-Containing Protein 28
D000951:Antigens, Neoplasm; D009687:Nuclear Proteins; D012097:Repressor Proteins; C112224:SUPT5H protein, human; D035602:Transcriptional Elongation Factors; D051548:Histone Acetyltransferases; C101116:TRIM28 protein, human; D000076183:Tripartite Motif-Containing Protein 28; D012319:RNA Polymerase II; C115866:OGA protein, human; D006821:Hyaluronoglucosaminidase
O-GlcNAcylation; RNA polymerase II; metabolism; transcription; transcription elongation factor