Overexpression of Human Diacylglycerol Acyltransferase 1, Acyl-CoA:Cholesterol Acyltransferase 1, or Acyl-CoA:Cholesterol Acyltransferase 2 Stimulates Secretion of Apolipoprotein B-containing Lipoproteins in McA-RH7777 Cells

John J Liang,Peter Oelkers,Cuiying Guo,Pi-Chun Chu,Joseph L Dixon,Henry N Ginsberg,Stephen L Sturley

doi:10.1074/jbc.m408507200

Abstract

The relative importance of each core lipid in the assembly and secretion of very low density lipoproteins (VLDL) has been of interest over the past decade. The isolation of genes encoding diacylglycerol acyltransferase (DGAT) and acyl-CoA:cholesterol acyltransferases (ACAT1 and ACAT2) provided the opportunity to investigate the effects of isolated increases in triglycerides (TG) or cholesteryl esters (CE) on apolipoprotein B (apoB) lipoprotein biogenesis. Overexpression of human DGAT1 in rat hepatoma McA-RH7777 cells resulted in increased synthesis, cellular accumulation, and secretion of TG. These effects were associated with decreased intracellular degradation and increased secretion of newly synthesized apoB as VLDL. Similarly, overexpression of human ACAT1 or ACAT2 in McA-RH7777 cells resulted in increased synthesis, cellular accumulation, and secretion of CE. This led to decreased intracellular degradation and increased secretion of VLDL apoB. Overexpression of ACAT2 had a significantly greater impact upon assembly and secretion of VLDL from liver cells than did overexpression of ACAT1. The addition of oleic acid (OA) to media resulted in a further increase in VLDL secretion from cells expressing DGAT1, ACAT1, or ACAT2. VLDL secreted from DGAT1-expressing cells incubated in OA had a higher TG:CE ratio than VLDL secreted from ACAT1- and ACAT2-expressing cells treated with OA. These studies indicate that increasing DGAT1, ACAT1, or ACAT2 expression in McA-RH7777 cells stimulates the assembly and secretion of VLDL from liver cells and that the core composition of the secreted VLDL reflects the enzymatic activity that is elevated.

Highlights

** Present address: Dept. of Nutritional Sciences, Rutgers, The State University of New Jersey, 96 Lipman Dr, New Brunswick, NJ 08901-8525
There is a wide range of apolipoprotein B (apoB) secretion from the liver in humans studied on average American diets, and overproduction of very low density lipoproteins (VLDL) and low density lipoproteins (LDL) is a common feature of human dyslipidemias [1,2,3] The secretion of apoB in cultured primary hepatocytes and hepatoma cells is regulated primarily at the post-translational level by degradation of newly synthesized apoB [4]
Sucrose gradient ultracentrifugation indicated that the increased numbers of apoB-containing lipoproteins secreted from cells overexpressing either ACAT1 or ACAT2 were in the form of VLDL particles (Fig. 4)

Summary

Introduction

** Present address: Dept. of Nutritional Sciences, Rutgers, The State University of New Jersey, 96 Lipman Dr, New Brunswick, NJ 08901-8525. Overexpression of human DGAT1 in rat hepatoma McA-RH7777 cells resulted in increased synthesis, cellular accumulation, and secretion of TG. Overexpression of human ACAT1 or ACAT2 in McA-RH7777 cells resulted in increased synthesis, cellular accumulation, and secretion of CE.

Results

Conclusion