Translocation of GPAT4 from the endoplasmic reticulum to the mitochondria may alter efficiency of TAG synthesis in liver cells

Synthesis of triacylglycerol (TAG) is critical for the formation of lipoproteins and lipid droplets and is the main constituent of mammalian fat. The rate of TAG synthesis is controlled by the enzyme glycerol-3-phosphate acyltransferase (GPAT). GPAT4, an isoform of GPAT, is found predominantly in the endoplasmic reticulum (ER), while GPAT1 is found in the mitochondria. Although GPAT4 and GPAT1 both produce lysophosphatidic acid (LPA), it is not known whether the output of phospholipids whose synthesis was initiated in the outer-mitochondrial membrane (OMM) differ from those that are initiated in the ER. It was hypothesized that GPAT4 and GPAT1 have non-interchangeable outputs within the cell and that the introduction of GPAT4 to the mitochondria would not affect TAG output. To determine if there was a difference between the outputs of GPAT 1 and GPAT4, primary liver cells were infected with adenovirus to express GPAT4 on the mitochondria (Ad-Tom20-GPAT4, “mtGPAT4-FLAG”). We measured GPAT enzyme activity in total particulate prepared from primary mouse hepatocytes and total particulate prepared from a mouse hepatoma cell line, Hepa 1-6, infected with these adenoviruses. The introduction of mtGPAT4-FLAG had an inconclusive change in activity in primary mouse hepatocytes while it had a doubling effect on enzyme specific activity in Hepa 1-6 cells as compared to the GFP control adenovirus (Ad-T20-GFP-BirA, “GFP control adenovirus”). These results indicate conflicting information about the introduction of mtGPAT4-FLAG. Future experiments within Hepa 1-6 cells or possibly whole animal models should be considered for further analysis on the effect of incorporating mtGPAT4-FLAG. Understanding what affects the efficiency of TAG synthesis is important for providing a better understanding of diseases that are influenced by TAG production and maintenance, such as diabetes mellitus, lipodystrophy, and heart disease.Bachelor of Scienc