Cloning and Characterization of a Second Human CTP:Phosphocholine Cytidylyltransferase

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