Abstract

Prior work has described novel form of highly dynamic nuclear and cytosolic protein glycosylation in which N-acetylglucosamine is O- glycosidically attached to Ser(Thr)-residues (0-GlcNAc) on a myriad of key regulatory and cytoskeletal proteins. O-GlcNAc is analogous to protein phosphorylation in several ways: 1) It is rapidly attached and removed in response to cellular stimuli and cell-cycle; 2) It occurs at sites also used by kinases; 3) It is often reciprocal with phosphorylation; 4) It is a regulatory modification.
The Specific Aims are designed to elucidate the functions of this ubiquitous modification:
Aim 1 is to further elucidate the roles of O-GlcNAc in transcription. The role of the dynamic O-GlcNAcylation of the C-terminal domain of RNA polymerase II and its basal transcription factors will be examined. The O-GlcNAcylation of the key basal transcriptional factor, TATA-binding protein (TBP) and the role of the saccharide in TBP's interactions with TAFs will be emphasized. The transcription-associated forms of O- GlcNAcase and O-GlcNAc Transferase will be identified and characterized.
Aim 2 is to continue to elucidate the regulation and diversity of O- GlcNAc Transferases (OGT). Questions that will be addressed are: 1) Regulation of OGT by tyrosine phosphorylation and 0-GlcNAcylation?; 2) Roles of the eleven tetratricopeptide repeats on OGT? 3) Phenotype of mice missing specific OGT or expressing an OGT mutated at tyrosine phosphorylation site(s)? 4) Protein and peptide specificity of the rat liver OGT? 5) How many different OGTs are there Aim 3 is to continue to clone and characterize O-GlcNAcases. The O-GlcNAcase purified from rat spleen will be cloned. Regulation and distribution of the enzyme in tissues and cells will be determined. Specific and highly potent inhibitors for the O-GlcNAcases will continue to be developed. These studies are fundamentally important to the regulation of transcription and signal transduction processes important to virtually all aspects of development and disease.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37HD013563-24
Application #
6387460
Study Section
Physiological Chemistry Study Section (PC)
Program Officer
Lock, Allan
Project Start
1998-04-01
Project End
2003-03-31
Budget Start
2001-04-01
Budget End
2002-03-31
Support Year
24
Fiscal Year
2001
Total Cost
$378,270
Indirect Cost

  Institution

Name
Johns Hopkins University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
045911138
City
Baltimore
State
MD
Country
United States
Zip Code
21218

  Publications

Hart, Gerald W; Slawson, Chad; Ramirez-Correa, Genaro et al. (2011) Cross talk between O-GlcNAcylation and phosphorylation: roles in signaling, transcription, and chronic disease. Annu Rev Biochem 80:825-58
Zachara, Natasha E; Molina, Henrik; Wong, Ker Yi et al. (2011) The dynamic stress-induced ""O-GlcNAc-ome"" highlights functions for O-GlcNAc in regulating DNA damage/repair and other cellular pathways. Amino Acids 40:793-808
Slawson, Chad; Hart, Gerald W (2011) O-GlcNAc signalling: implications for cancer cell biology. Nat Rev Cancer 11:678-84
Butkinaree, Chutikarn; Park, Kyoungsook; Hart, Gerald W (2010) O-linked beta-N-acetylglucosamine (O-GlcNAc): Extensive crosstalk with phosphorylation to regulate signaling and transcription in response to nutrients and stress. Biochim Biophys Acta 1800:96-106
Li, Xi; Molina, Henrik; Huang, Haiyan et al. (2009) O-linked N-acetylglucosamine modification on CCAAT enhancer-binding protein beta: role during adipocyte differentiation. J Biol Chem 284:19248-54
Housley, Michael P; Udeshi, Namrata D; Rodgers, Joseph T et al. (2009) A PGC-1alpha-O-GlcNAc transferase complex regulates FoxO transcription factor activity in response to glucose. J Biol Chem 284:5148-57
Zachara, Natasha E (2009) Detecting the ""O-GlcNAc-ome""; detection, purification, and analysis of O-GlcNAc modified proteins. Methods Mol Biol 534:251-79
Copeland, Ronald J; Bullen, John W; Hart, Gerald W (2008) Cross-talk between GlcNAcylation and phosphorylation: roles in insulin resistance and glucose toxicity. Am J Physiol Endocrinol Metab 295:E17-28
Cheung, Win D; Hart, Gerald W (2008) AMP-activated protein kinase and p38 MAPK activate O-GlcNAcylation of neuronal proteins during glucose deprivation. J Biol Chem 283:13009-20
Slawson, Chad; Lakshmanan, T; Knapp, Spencer et al. (2008) A mitotic GlcNAcylation/phosphorylation signaling complex alters the posttranslational state of the cytoskeletal protein vimentin. Mol Biol Cell 19:4130-40

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