Interactions between microsomal triglyceride transfer protein and apolipoprotein B within the endoplasmic reticulum in a heterologous expression system

When apolipoprotein B (apoB) is expressed in heterologous cells, it is not secreted but retained and de graded within the endoplasmic reticulum (ER), We have previously characterized carboxyl-terminal truncated forms of apoB expressed in COS cells and have shown that these proteins were readily synthesized but retained within the ER and degraded, if the size of the truncated protein was larger than apoB 29, Below this size, the smaller the size of the apoB truncates, the greater the extent of secretion, although > 50% of these smaller proteins were also degraded within the ER, In the present study, we demonstrate that this secretory defect can be overcome by coexpression with microsomal triglyceride transfer protein (MTP); moreover, this complementation is inversely related to the size of apoB, Secretion of apoBs larger than B29 required the coexpression of MTP and, in the presence of MTP, was oleate-responsive, MTP, in the presence or absence of oleate supplementation, had little or no effect on the secretion of the shorter truncates, We discovered, however, that MTP was physically associated with all forms of apoB intracellularly (B13-B41), The association of MTP with apoB 41 was stable to high salt washing, as well as to low pH, suggesting that these interactions may be hydrophobic in nature, In addition to the interaction with MTP, apoB was also found to be associated with calnexin, confirming previous studies, and with proteins bearing the KDEL retention signal, However, studies on overexpression of human calnexin and tunicamycin inhibition of glycosylation showed that interaction with calnexin was not necessary for the formation or secretion of apoB 41-containing lipoproteins; moreover, in the presence of MTP, the association of calnexin with apoB 41 was transient or absent, These data suggest that for apoB to attain a folded state sufficient to escape the quality control of the ER, it needs to obtain neutral lipid (supplied by MTP), as well as its ability to keep it packaged as a rudimentary lipoprotein, dependent on its size being larger than B29.