Phospholipid Modifying Enzymes And The Regulation Of Golgi Complex Structure And Function
The secretory pathway in cells is critical for the precise synthesis, modification and directed transport of proteins, lipids, and carbohydrates. Phospholipid metabolizing enzymes are thought to contribute to the transport of cargo within the cell by altering the lipid composition of membranes and allowing them to form transport carriers including vesicles and membrane tubules. The specific phospholipid-modifying enzymes have not been fully discovered, but evidence suggests roles for both lysophospholipid acyltransferases (LPATs) and phospholipase A2 (PLA2) enzymes in mediating membrane trafficking. Using inhibitor-based studies, PLA2 activity has been shown to be important for Golgi trafficking and structure. PLA2 antagonists block the formation of membrane tubules from the Golgi. Conversely, LPAT inhibitors cause the formation of Golgi membrane tubules. From those observations, we hypothesize a role for PLA2 enzymes in generating membrane tubules and a role for LPATs in the negative regulation of membrane tubules, which are important for Golgi structure and function. In this study I explore the functions of a novel protein, lysophosphatidic acid acyltransferase 3 (LPAAT3), in phospholipid metabolism, maintaining Golgi structure, membrane trafficking, and membrane tubule regulation. Using biochemical approaches, I have shown that LPAAT3 generates phosphatidic acid in Golgi membranes. LPAAT3’s activity is critical for maintaining a compact Golgi structure. Furthermore, LPAAT3 expression alters both retrograde and anterograde trafficking by regulating the formation of membranes tubules from the Golgi. The role of PLA2 enzymes in trafficking was also explored using inhibitors to study trafficking from the Trans Golgi Network (TGN) to the cell surface. PLA2 inhibitors blocked trafficking to the plasma membrane and caused the accumulation of cargo in the TGN. Furthermore, the formation of TGN tubules, whose fission is regulated by protein kinase D (PKD), is dependent on PLA2 activity as well. I show that both LPATs and PLA2s are important factors in regulating membrane trafficking within cells. They do this by altering the phospholipids within membranes and by promoting or regulating the formation of membrane vesicles and membrane tubules. These results contribute to our understanding of membrane trafficking and the phospholipids involved in those pathways.
dissertation or thesis