Temperature-induced changes in the transbilayer distribution of phosphatidylethanolamine in mitoplasts of rainbow trout (Oncorhynchus mykiss) liver

Abstract

Increased proportions of phosphatidylethanolamine (PE) are a common adaptation of poikilothermic animals to life at cold temperatures. Although amino-phospholipids such as PE are asymmetrically distributed between the two membrane hemilayers, the impact of temperature-induced changes in phospholipid composition on PE asymmetry has not been examined. Accordingly, the distribution of PE between the inner and outer hemilayers of the inner membrane of liver mitochondria was determined in rainbow trout (Oncorhynchus mykiss) acclimated to 20°C and 5°C. Mitochondria were isolated from trout liver by a combination of differential and Nycodenz™ density-gradient centrifugation, resulting in an 11- to 15-fold enrichment in the activity of the mitochondrial enzyme marker succinate cytochrome c reductase. Interestingly, the outer mitochondrial membrane was more stable in 5°C- than 20°C-acclimated trout as indicated by the higher concentration of digitonin (0.6 vs. 0.3 mg digitonin/mg mitochondrial protein) required for the preparation of mitoplasts. Phosphatidylethanolamines in the outer hemilayer of the inner mitochondrial membrane (IMM) were identified as derivatives of 2,4,6-trinitrobenzene sulfonic acid (TNBS), a nonpenetrating reagent that reacts with primary amines. Our results demonstrate 1) a decrease in the phosphatidylcholine:phosphatidylethanolamine ratio in the IMM of 5°C- (1.43 ± 0.09) compared to 20°C- (2.31 ± 0.34) acclimated trout and 2) smaller amounts of PE in the outer hemilayer (6.0 ± 0.51%) of the IMM from 5°C- than 20°C-acclimated trout (8.1 ± 0.55%). These results indicate that the composition of the inner mitochondrial membrane is enriched in PE at cold temperatures and that this enrichment is largely restricted to, and therefore most likely to be of functional significance to, the inner hemilayer. © 1996 Wiley-Liss, Inc.