Nucleotide sequence and tissue-specific expression of the multifunctional protein carbamoyl-phosphate synthetase-aspartate transcarbamoylase-dihydroorotase (CAD) mRNA in Squalus acanthias.
Carbamoyl-phosphate synthetase II (CPSase II), aspartate transcarbamoylase (ATCase), and dihydroorotase (DHOase) catalyze the first three steps of de novo pyrimidine nucleotide biosynthesis, respectively. In mammalian species, these three enzyme activities exist in the cytosol in liver and other tissues as a multifunctional complex on a single polypeptide called carbamoyl-phosphate synthetase-aspartate transcarbamoylase-dihydroorotase (CAD) in the order of NH2-CPSase II-DHOase-ATCase-COOH. Previous studies provided evidence that in Squalus acanthias (spiny dogfish) these enzymes are not expressed in liver and that they exist as separate entities in the cytosol of extra-hepatic tissues such as testes and spleen (Anderson, P. M. (1989) Biochem. J. 261, 523-529). Here we report that the genes for these three enzymes are expressed in testes as a single transcript analogous to CAD in mammalian species and that these genes are not expressed in liver at levels that can be detected by Northern blots or by the polymerase chain reaction. The absence of the pyrimidine pathway in the liver may be related to the exclusive localization of glutamine synthetase in the mitochondrial matrix which provides for efficient assimilation of ammonia as glutamine for urea synthesis in these ureoosmotic species; thus glutamine may not be available for CPSase II or other amidotransferase activities in the cytosol. The amino acid sequence deduced from the nucleotide sequence of the shark CAD cDNA reported here is very similar to CAD from other species; alignment with the hamster CAD sequence shows 77% identical residues.
Hong
J
J
Department of Biochemistry and Molecular Biology, University of Minnesota, Duluth 55812, USA.
Salo
W L
WL
Anderson
P M
PM
D000595:Amino Acid Sequence; D000818:Animals; D001221:Aspartate Carbamoyltransferase / Q000737:chemistry / Q000235:genetics*; D001483:Base Sequence; D002222:Carbamoyl-Phosphate Synthase (Ammonia) / Q000737:chemistry / Q000235:genetics*; D004080:Dihydroorotase / Q000737:chemistry / Q000235:genetics*; D004284:Dogfish; D008969:Molecular Sequence Data; D009928:Organ Specificity; D012333:RNA, Messenger / Q000032:analysis* / Q000737:chemistry
D012333:RNA, Messenger; D001221:Aspartate Carbamoyltransferase; D004080:Dihydroorotase; D002222:Carbamoyl-Phosphate Synthase (Ammonia)