Use of acyltransferase inhibitors to block vesicular traffic between the ER and Golgi complex.

This article describes the use of acyltransferase inhibitors as probes for studying the potential role of lysophospholipid acyltransferases (LPAT) in intracellular membrane trafficking in the secretory and endocytic pathways. The small molecule inhibitors that are described here were originally found as acyl-CoA:cholesterol acyltransferase (ACAT) inhibitors. One of these, CI-976 (2,2-methyl-N-(2,4,6,-trimethoxyphenyl)dodecanamide), was also found to be a potent LPAT inhibitor. CI-976 is a small, hydrophobic, membrane-permeant compound and both in vivo and in vitro studies have shown that it, but not other ACAT inhibitors, has a profound effect on multiple membrane trafficking pathways in eukaryotic cells including: (1) inhibition of COPII vesicle budding from the endoplasmic reticulum (ER), (2) inhibition of transferrin and transferrin receptor export from the endocytic recycling compartment, and (3) stimulation of tubule-mediated retrograde trafficking of Golgi membranes to the ER. Here we describe the use of CI-976 and other ACAT inhibitors for studies with both cultured mammalian cells and in vitro reconstitution assays, with a particular emphasis on COPII vesicle budding from the ER. All of these studies strongly suggest that CI-976-sensitive LPATs play a role in coated vesicle fission, and therefore, CI-976 is a valuable addition to the arsenal of small molecule inhibitors that can be used to study secretory and endocytic membrane trafficking pathways.