Journal
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
Volume 465, Issue 2, Pages 347-358Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.abb.2007.06.033
Keywords
triacylglycerol; glycerolipid; testis; steatosis; gpam
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Funding
- FIC NIH HHS [R03 TW006034] Funding Source: Medline
- NIDDK NIH HHS [R56 DK056598, R01 DK056598-22, R01 DK056598, K01 DK059931] Funding Source: Medline
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Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the initial and rate-limiting step in glycerolipid synthesis. Several mammalian GPAT activities have been recognized, including N-ethylmaleimide (NEM)-sensitive isoforms in microsomes and mitochondria and an NEM-resistant form in mitochondrial outer membrane (GPAT1). We have now cloned a second mitochondrial isoform, GPAT2 from mouse testis. The open-reading frame encodes a protein of 798 amino acids with a calculated mass of 88.8 kDa and 27% amino acid identity to GPAT1. Testis mRNA expression was 50-fold higher than in liver or brown adipose tissue, but the specific activity of NEM-sensitive GPAT in testis mitochondria was similar to that in liver. When Cos-7 cells were transiently transfected with GPAT2, NEM-sensitive GPAT activity increased 30%. Confocal microscopy confirmed a mitochondrial location. incubation of GPAT2-transfected Cos-7 cells with trace (3 mu M, 0.25 mu Ci) [1-C-14]oleate for 6 h increased incorporation of [C-14]oleate into TAG 84%. In contrast, incorporation into phospholipid species was lower than in control cells. Although a polyclonal antibody raised against full-length GPAT1 detected ail similar to 89-kDa band in liver and testis from GPAT1 null mice and both 89- and 80-kDa bands in BAT from the knockout animals, the GPAT2 protein expressed in Cos-7 cells was only 80 kDa. In vitro translation showed a single product of 89 kDa. Unlike GPAT1, GPAT2 mRNA abundance in liver was not altered by fasting or refeeding. GPAT2 is likely to have a specialized function in testis. (C) 2007 Elsevier Inc. All rights reserved.
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