Cloning and characterization of a second human CTP:Phosphocholine cytidylyltransferase

CTP:phosphocholine cytidylyltransferase (CCT) is a key regulator of phosphatidylcholine biosynthesis, and only a single isoform of this enzyme, CCT alpha, is known. We identified and sequenced a human cDNA that encoded a distinct CCT isoform, called CCT beta, that is derived from a gene different from that encoding CCT alpha, CCT beta transcripts were detected in human adult and fetal tissues, and very high transcript levels were found in placenta and testis, CCT beta and CCT alpha proteins share highly related, but not identical, catalytic domains followed by three amphipathic helical repeats. Like CCT alpha, CCT beta required the presence of lipid regulators for maximum catalytic activity, The amino terminus of CCT beta bears no resemblance to the amino terminus of CCT alpha, and CCT beta protein was localized to the cytoplasm as detected by indirect immunofluorescent microscopy, Whereas CCT alpha activity is regulated by reversible phosphorylation, CCT beta lacks most of the corresponding carboxyl-terminal domain and contained only 3 potential phosphorylation sites of the 16 identified in CCT alpha. Transfection of COS-7 cells with a CCT beta expression construct led to the overexpression of CCT activity, the accumulation of cellular CDP-choline, and enhanced radiolabeling of phosphatidylcholine, CCT beta protein was posttranslationally modified in COS-7 cells, resulting in slower migration during polyacrylamide gel electrophoresis. Expression of CCT beta/CCT alpha chimeric proteins showed that the amino-terminal portion of CCT beta was required for posttranslational modification. These data demonstrate that a second, distinct CCT enzyme is expressed in human tissues and provides another mechanism by which cells regulate phosphatidylcholine production.