Premium
Phosphorylation of Human CTPS1 by GSK3β
Author(s) -
Higgins Matthew James,
Graves Paul R,
Graves Lee M
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.6.a987-b
Subject(s) - phosphorylation , kinase , biochemistry , protein kinase c , protein phosphorylation , substrate level phosphorylation , adenosine triphosphate , biology , protein serine threonine kinases , protein kinase a , chemistry , microbiology and biotechnology
Cytidine triphosphate synthetase (CTPS) is an essential enzyme in the de novo synthesis of CTP, which is important in phospholipid metabolism and DNA synthesis. CTPS catalyzes the rate‐limiting step in the formation of cytidine nucleotides by transferring the amine from glutamine to uridine triphosphate (UTP) to form CTP. Although CTPS phosphorylation has been well characterized in yeast, relatively little is known about the phosphorylation of mammalian CTPS. In this study, we characterized the phosphorylation of human CTPS1 in human embryonic kidney (HEK) 293 cells and examined possible kinases responsible for CTPS1 phosphorylation. In yeast, CTPS1 has been shown to be phosphorylated by protein kinase A (PKA) and protein kinase C (PKC) on multiple residues. Based on the observations in yeast, we investigated whether PKA and/or PKC could phosphorylate human CTPS1. Neither activation (forskolin, TPA) or inhibition (H89, GF 109203 X) of PKA or PKC altered CTPS1 phosphorylation. However, human CTPS1 was found to be phosphorylated and we were able to identify the major sites of phosphorylation in human CTPS1 using two dimensional TLC and mass spectrometry. Interestingly, further investigation of stimuli that might cause human CTPS1 phosphorylation revealed that human CTPS1 can be phosphorylated under low serum conditions. Investigation of kinases activated under low serum conditions revealed that glycogen synthase kinase 3 beta (GSK3‐beta) can phosphorylate CTPS1 in intact cells and in vitro demonstrating CTPS1 is a new target for GSK3‐beta. Funding to LMG from NIH