The distal, not the proximal promoter upregulates mitochondrial glycerophosphate acyltransferase in starved and refed rat liver cells

Abstract

The effect of starvation and refeeding of liver cells on the activity of mitochondrial glycerophosphate acyltransferase (mtGPAT) was investigated. Rat liver cells were cultured in media lacking glucose, pyruvate, and fetal bovine serum for different periods of time, then placed in media, which contained those compounds, for additional time. After 24, 48 and 72 hrs of starvation and 24 hrs of refeeding of the cells, mtGPAT activity increased up to 5 fold. The viability of the cells was between 85% and 90%. Western blot results confirmed the presence of a larger amount of mtGPAT protein in the group of cells that was starved for 24, 48, and 72 hrs and refed for 24 hrs. The results of RT‐PCR showed enhancement of mRNA levels in the cells after starvation and refeeding. These results suggest that refeeding of liver cells induce an increase in mtGPAT. Additional experiments were done with liver cells transfected with pGL3‐ basic vector containing distal or proximal promoter of mtGPAT and luciferase as the reporter gene. Starvation and refeeding caused 2 to 2.5 fold increase in the luciferase activity in cells transfected with the distal promoter over control cells. No change in the luciferase activity was observed in the cells transfected with the proximal promoter. The results are in keeping with the idea that the increase in mtGPAT is regulated via the distal and not the proximal promoter. Supported by NIH grant GM‐57643.