Phospholipid Transfer Activity of Microsomal Triacylglycerol Transfer Protein Is Sufficient for the Assembly and Secretion of Apolipoprotein B Lipoproteins

Abstract

Human microsomal triacylglycerol transfer protein (hMTP) is essential for apolipoprotein B (apoB)-lipoprotein assembly and secretion and is known to transfer triacylglycerols, cholesterol esters, and phospholipids. To understand the relative importance of each lipid transfer activity, we compared the ability of hMTP and its Drosophila ortholog (dMTP) to assemble apoB lipoproteins and to transfer various lipids. apoB48 secretion was induced when co-expressed with either hMTP or dMTP in COS cells, and oleic acid supplementation further augmented secretion without altering particle density. C-terminal epitope-tagged dMTP (dMTP-FLAG) facilitated the secretion of apoB polypeptides in the range of apoB48 to apoB72 but was ϳ50% as efficient as hMTP-FLAG. Comparison of lipid transfer activities revealed that although phospholipid transfer was similar in both orthologs, dMTP was unable to transfer neutral lipids. We conclude that the phospholipid transfer activity of MTP is sufficient for the assembly and secretion of primordial apoB lipoproteins and may represent its earliest function evolved for the mobilization of lipid in invertebrates. Identification of MTP inhibitors, which selectively affect transfer of a specific lipid class, may have therapeutic potential.Lipoproteins are lipid-protein complexes that transport lipids, fatsoluble vitamins, and other hydrophobic molecules in the plasma. Apolipoprotein B (apoB) 2 is a structural protein embedded in the phospholipid monolayer on the surface of triglyceride-rich lipoproteins. It has been hypothesized that apoB contains amphipathic ␣-helical and ␤-sheet domains (1). Lipidation of the ␤-sheets is necessary for the assembly of larger apoB polypeptides into lipoprotein emulsion particles. Lipoprotein assembly begins co-translationally and microsomal triglyceride transfer protein (MTP) plays a pivotal role in this process (for reviews, see Refs. 2-6). MTP is absent in abetalipoproteinemia, a disease characterized by the deficit of plasma apoB lipoproteins and low plasma cholesterol levels (7,8). Reconstitution of MTP in heterologous systems rescues apoB secretion, whereas tissue-specific liver knock-out models recreate the lipoprotein deficiency present in abetalipoproteinemia (9, 10). The signature activity of MTP is its ability to transfer lipids between membranes. It has been demonstrated that MTP lipid transfer activity is necessary for apoB lipoprotein assembly and secretion (for reviews, see Refs. 2-6). More recent evidence suggests that MTP lipid transfer activity is also responsible for lipid accretion within the secretory pathway and that MTP potentially stabilizes lipid vesicles in the endoplasmic reticulum (reviewed in Refs. 2 and 3).MTP transfers several lipids including triacylglycerols, cholesterol esters, and phospholipids between vesicles in vitro (11)(12)(13)(14). It has been suggested that MTP transiently interacts with a membrane, extracts lipids, and then delivers them to another lipid acceptor or to nascent apoB lipoproteins (15). Kinetic studies in...

Microsomal Triglyceride Transfer Protein Activity Is Not Required for the Initiation of Apolipoprotein B-containing Lipoprotein Assembly in McA-RH7777 Cells

“…In a recent study, Rava et al (60) have supported our previous stud-ies demonstrating that the primordial apoB-containing particles are PL-rich (26 -28) and have proposed that the PL transfer activity of MTP is sufficient for the assembly and secretion of primordial apoB lipoproteins. This conclusion was based mainly on the observations derived from the effect of Drosophila MTP on the secretion of truncated forms of human apoB in COS cells (60).…”

“…In a recent study, Rava et al (60) have supported our previous stud-ies demonstrating that the primordial apoB-containing particles are PL-rich (26 -28) and have proposed that the PL transfer activity of MTP is sufficient for the assembly and secretion of primordial apoB lipoproteins. This conclusion was based mainly on the observations derived from the effect of Drosophila MTP on the secretion of truncated forms of human apoB in COS cells (60). These investigators showed that Drosophila MTP, which is defective in TAG transfer activity but has PL transfer activity equal to that of human MTP, supports the secretion of human apoB48, apoB53, and apoB72 in COS cells (60).…”

“…These investigators showed that Drosophila MTP, which is defective in TAG transfer activity but has PL transfer activity equal to that of human MTP, supports the secretion of human apoB48, apoB53, and apoB72 in COS cells (60). Our results, indicating that MTP activity is not required for the initiation of apoB assembly, are at variance with the conclusion reached by Rava et al (60). Although we do not know the exact reason for this discrepancy, we speculate that the inconsistency in the results arise, most likely, from the use of cell lines with distinctly different tissue origins.…”

“…This conclusion was based mainly on the observations derived from the effect of Drosophila MTP on the secretion of truncated forms of human apoB in COS cells (60). These investigators showed that Drosophila MTP, which is defective in TAG transfer activity but has PL transfer activity equal to that of human MTP, supports the secretion of human apoB48, apoB53, and apoB72 in COS cells (60). Our results, indicating that MTP activity is not required for the initiation of apoB assembly, are at variance with the conclusion reached by Rava et al (60).…”

Defining Lipid-Interacting Domains in the N-Terminal Region of Apolipoprotein B

“…Further apoB translation into the β1 superdomain allows apoB alone to stabilize a lipoprotein and may push off MTP. Recently, it was reported that dMTP, a drosophila MTP homolog that lacks triglyceride transfer activity, promoted apoB-containing lipoprotein assembly in COS cells (46). The authors concluded that the phospholipid transfer activity of dMTP was responsible for this function.…”