A novel murine CTP:phosphoethanolamine cytidylyltransferase splice variant is a post-translational repressor and an indicator that both cytidylyltransferase domains are required for activity |
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Authors: | Zvezdan Pavlovic Ratnesh Kumar SinghMarica Bakovic |
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Affiliation: | Department of Human Health and Nutritional Sciences, University of Guelph, 50 Stone Road East, Guelph, Ontario N1G2W1, Canada |
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Abstract: | CTP:phosphoethanolamine cytidylyltransferase (Pcyt2) has an important regulatory function in biosynthesis of the membrane phospholipid phosphatidylethanolamine. We previously determined that the full-length Pcyt2α and its splice variant Pcyt2β 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, Pcyt2γ, in which the second cytidylyltransferase domain at the C-terminus becomes deleted. Pcyt2γ is ubiquitously expressed in embryonic and adult mouse tissues, and is the most abundant in the kidney, skeletal muscle and testis. Pcyt2γ splicing mechanism dominates over Pcyt2β exon-skipping mechanism in most examined tissues. Although Pcyt2γ 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, Pcyt2γ interacts with the active isoform, Pcyt2α, and significantly reduces Pcyt2α homodimerization and activity. The inactive N-domain (H35Y, H35A) and C-domain (H244Y, H244A) mutants of Pcyt2α also reduce Pcyt2α homodimerization and activity. This study revealed the importance of both cytidylyltransferase 35HYGH and 244HIGH motifs for the activity of murine Pcyt2α and established that the naturally occurring splice variant Pcyt2γ has a function to restrain the enzyme activity through the formation of unproductive enzyme complexes. |
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Keywords: | Pcyt2, CTP:phosphoethanolamine cytidylyltransferase PE, phosphatidylethanolamine EK, ethanolamine kinase P-Etn, phosphoethanolamine EPT, CDP-ethanolamine:1,2-diacylglycerol ethanolaminephosphotransferase PfECT, Plasmodium falciparum Pcyt2 GAPDH, glyceraldehyde 3-phosphate dehydrogenase GCT, CTP:glycerol-3-phosphate cytidylyltransferase CCT1, CTP:phosphocholine cytidylyltransferase 1 HMGCS1, HMG-CoA synthase LDLR, LDL receptor HMGCR, 3-hydroxy-3-methylglutaryl coenzyme A reductase PCSK9, proprotein convertase subtilisin/kexin type 9 ASM, acid sphingomyelinase |
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