A novel murine CTP:phosphoethanolamine cytidylyltransferase splice variant is a post-translational repressor and an indicator that both cytidylyltransferase domains are required for activity

CTP:phosphoethanolamine cytidylyltransferase (Pcyt2) has an important regulatory function in biosynthesis of the membrane phospholipid phosphatidylethanolamine. We previously determined that the full-length Pcyt2 alpha and its splice variant Pcyt2 beta are the main active isoforms of this enzyme. Here we report that mouse Pcyt2 could be spliced at Introns 7 and 8 to produce a unique third isoform, Pcyt2 gamma, in which the second cytidylyltransferase domain at the C-terminus becomes deleted. Pcyt2 gamma is ubiquitously expressed in embryonic and adult mouse tissues, and is the most abundant in the kidney, skeletal muscle and testis. Pcyt2 gamma splicing mechanism dominates over Pcyt2 beta exon-skipping mechanism in most examined tissues. Although Pcyt2 gamma maintains the N-terminal cytidylyltransferase domain as most cytidylyltransferases, the lack of the C-terminal cytidylyltransferase domain causes a complete loss of catalytic activity. However, Pcyt2 gamma interacts with the active isoform, Pcyt2 alpha, and significantly reduces Pcyt2 alpha homodimerization and activity. The inactive N-domain (H35Y, H35A) and C-domain (H244Y, H244A) mutants of Pcyt2 alpha also reduce Pcyt2 alpha homodimerization and activity. This study revealed the importance of both cytidylyltransferase (35)HYGH and (244)HIGH motifs for the activity of murine Pcyt2 alpha and established that the naturally occurring splice variant Pcyt2 gamma has a function to restrain the enzyme activity through the formation of unproductive enzyme complexes. (C) 2014 Elsevier B.V. All rights reserved.