The overall goal of this research is to gain insight into the mechanism of the regulation of protein synthesis initiation in animal cells. The rate of protein synthesis in animal cells is regulated by the status of the phosphorylation of the smallest alpha-subunit of eukaryotic initiation factor 2 (Eif2). p67, a cellular glycoprotein, protects eIF2alpha from phosphorylation by active kinases, PKR and HCR. The O-GlcNAc moieties of p67 are required for its activity. In response to stress such as viral infection, mitosis, and possibly others, p67 becomes inactive because of the high level of the activity of p67-deglycosylase which removes the O-GlcNAc moieties from p67. In this research the detailed structure-function relationship of p67 will be studied by genetic and biochemical analysis.
Specific aims of this research are to identify and perform functional analyses for (i) p67 dimerization domain(s), (ii) p67 domains that are binding to eIF2alpha, eIF2gamma, PKR, and p67-DG, (iii) GlcNAc modification sites of p67, (iv) the phosphorylation site(s) where PKR transfers its phosphoryl group to p67, (v) the """"""""lysine boxes"""""""" and an """"""""acid box"""""""" of p67 in the regulation of protein synthesis, and (vi) the methionine aminopeptidase activity of rat p67. Further experimentations will be performed to understand the regulatory roles of p67 and p67-deglycosylase in the initiation of protein synthesis during cellular stress responses such as terminal differentiation, and viral infections. To obtain a comprehensive picture about the regulation of the p67 activity, p67-deglycosylase which is specific to p67, will be purified, cloned, and characterized. This research is expected to provide valuable information regarding the mechanism of protein synthesis initiation in normal, cancerous, and virus- infected animal cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
7R01GM059190-02
Application #
6181437
Study Section
Physiological Chemistry Study Section (PC)
Program Officer
Rhoades, Marcus M
Project Start
1999-09-01
Project End
2003-08-31
Budget Start
2000-09-01
Budget End
2001-08-31
Support Year
2
Fiscal Year
2000
Total Cost
$70,073
Indirect Cost
Name
Kent State University at Kent
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
City
Kent
State
OH
Country
United States
Zip Code
44242
Ghosh, Arnab; Datta, Rekha; Majumdar, Avijit et al. (2006) The N-terminal lysine residue-rich domain II and the 340-430 amino acid segment of eukaryotic initiation factor 2-associated glycoprotein p67 are the binding sites for the gamma-subunit of eIF2. Exp Cell Res 312:3184-203
Datta, Bansidhar; Datta, Rekha; Majumdar, Avijit et al. (2005) The stability of eukaryotic initiation factor 2-associated glycoprotein, p67, increases during skeletal muscle differentiation and that inhibits the phosphorylation of extracellular signal-regulated kinases 1 and 2. Exp Cell Res 303:174-82
Datta, Bansidhar; Majumdar, Avijit; Datta, Rekha et al. (2004) Treatment of cells with the angiogenic inhibitor fumagillin results in increased stability of eukaryotic initiation factor 2-associated glycoprotein, p67, and reduced phosphorylation of extracellular signal-regulated kinases. Biochemistry 43:14821-31
Datta, Bansidhar; Datta, Rekha; Ghosh, Arnab et al. (2004) Eukaryotic initiation factor 2-associated glycoprotein, p67, shows differential effects on the activity of certain kinases during serum-starved conditions. Arch Biochem Biophys 427:68-78
Datta, Rekha; Tammali, Ravinder; Datta, Bansidhar (2003) Negative regulation of the protection of eIF2alpha phosphorylation activity by a unique acidic domain present at the N-terminus of p67. Exp Cell Res 283:237-46
Datta, Rekha; Choudhury, Papiya; Ghosh, Arnab et al. (2003) A glycosylation site, 60SGTS63, of p67 is required for its ability to regulate the phosphorylation and activity of eukaryotic initiation factor 2alpha. Biochemistry 42:5453-60
Datta, Bansidhar; Datta, Rekha (2003) Mutation at the acidic residue-rich domain of eukaryotic initiation factor 2 (eIF2alpha)-associated glycoprotein p67 increases the protection of eIF2alpha phosphorylation during heat shock. Arch Biochem Biophys 413:116-22
Datta, R; Choudhury, P; Bhattacharya, M et al. (2001) Protection of translation initiation factor eIF2 phosphorylation correlates with eIF2-associated glycoprotein p67 levels and requires the lysine-rich domain I of p67. Biochimie 83:919-31
Datta, B (2000) MAPs and POEP of the roads from prokaryotic to eukaryotic kingdoms. Biochimie 82:95-107