Regulation of carbamoyl phosphate synthetase by MAP kinase

• DOI: 10.1038/35002111
• Publication Date: 2002-07-26T08:41:25Z
• Authors: Lee M. Graves; Hedeel I. Guy; Piotr Kozlowski; Min Huang; Eduardo Lazarowski; R. Marshall Pope; Matthew A. Collins; Erik N. Dahlstrand; H. Shelton Earp; David R. Evans
• Keywords: Flavonoids; 0303 health sciences; Mesocricetus; MAP Kinase Signaling System; Molecular Sequence Data; Phosphoribosyl Pyrophosphate; Cell Line; Enzyme Activation; 03 medical and health sciences; Allosteric Regulation; Multienzyme Complexes; Cricetinae; Aspartate Carbamoyltransferase; Mutagenesis, Site-Directed; Animals; Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing); Pyrimidine Nucleotides; Amino Acid Sequence; Enzyme Inhibitors; Mitogen-Activated Protein Kinases; Phosphorylation; Dihydroorotase

The de novo synthesis of pyrimidine nucleotides is required for mammalian cells to proliferate. The rate-limiting step in this pathway is catalysed by carbamoyl phosphate synthetase (CPS II), part of the multifunctional enzyme CAD. Here we describe the regulation of CAD by the mitogen-activated protein (MAP) kinase cascade. When phosphorylated by MAP kinase in vitro or activated by epidermal growth factor in vivo, CAD lost its feedback inhibition (which is dependent on uridine triphosphate) and became more sensitive to activation (which depends upon phosphoribosyl pyrophosphate). Both these allosteric regulatory changes favour biosynthesis of pyrimidines for growth. They were accompanied by increased epidermal growth factor-dependent phosphorylation of CAD in vivo and were prevented by inhibition of MAP kinase. Mutation of a consensus MAP kinase phosphorylation site abolished the changes in CAD allosteric regulation that were stimulated by growth factors. Finally, consistent with an effect of MAP kinase signalling on CPS II activity, epidermal growth factor increased cellular uridine triphosphate and this increase was reversed by inhibition of MAP kinase. Hence these studies may indicate a direct link between activation of the MAP kinase cascade and de novo biosynthesis of pyrimidine nucleotides.