Functional analysis of the missense APOC3 mutation Ala23Thr associated with human hypotriglyceridemia

We have shown that expression of apolipoprotein (apo) C-III promotes VLDL secretion from transfected McA-RH7777 cells under lipid-rich conditions. To determine structural elements within apoC-III that confer to this function, we contrasted wild-type apoC-III with a mutant Al-a23Thr originally identified in hypotriglyceridemia subjects. Although synthesis of [H-3]glycerol-labeled TAG was comparable between cells expressing wild-type apoC-III (C3wt cells) or Ala23Thr mutant (C3AT cells), secretion of [H-3] TAG from C3AT cells was markedly decreased. The lowered [ 3 H] TAG secretion was associated with an inability of C3AT cells to assemble VLDL1. Moreover, [H-3] TAG within the microsomal lumen in C3AT cells was 60% higher than that in C3wt cells, yet the activity of microsomal triglyceride-transfer protein in C3AT cells was not elevated. The accumulated [H-3] TAG in C3AT microsomal lumen was mainly associated with lumenal IDL/LDL-like lipoproteins. Phenotypically, this [H-3] TAG fractionation profiling resembled what was observed in cells treated with brefeldin A, which at low dose specifically blocked the second-step VLDL1 maturation. Furthermore, lumenal [S-35] Ala23Thr protein accumulated in IDL/LDL fractions and was absent in VLDL fractions in C3AT cells. These results suggest that the presence of Ala23Thr protein in lumenal IDL/LDL particles might prevent effective fusion between lipid droplets and VLDL precursors. Thus, the current study reveals an important structural element residing within the N-terminal region of apoC-III that governs the second step VLDL1 maturation.-Sundaram, M., S. Zhong, M. B. Khalil, H. Zhou, Z. G. Jiang, Y. Zhao, J. Iqbal, M. M. Hussain, D. Figeys, Y. Wang, and Z. Yao. Functional analysis of the missense APOC3 mutation Ala23Thr associated with human hypotriglyceridemia. J. Lipid Res. 2010. 51: 1524-1534.