THE AMINO-TERMINUS OF APOLIPOPROTEIN-B IS NECESSARY BUT NOT SUFFICIENT FOR MICROSOMAL TRIGLYCERIDE TRANSFER PROTEIN RESPONSIVENESS

Human apolipoprotein (apo) B mediates the formation of neutral Lipid-c ontaining lipoproteins in the liver and intestine. The association of apoB with lipid is thought to be promoted by the microsomal triglyceri de transfer protein complex. We have reconstituted lipoprotein assembl y in an insect cell line that normally does not support this process. Expression of human microsomal triglyceride transfer protein (MTP) and apolipoprotein B48 (apoB48) together enabled Sf-21 insect cells to se crete similar to 60-fold more lipoprotein-associated triacylglycerol t han control cells. This dramatic effect demonstrates that effective pa rtitioning of triacylglycerol into the secretory pathway requires an e ndoplasmic reticulum-associated neutral lipid transporter (provided by MTP) and an apolipoprotein to shuttle the lipid through the pathway. Expression of the human apoB48 gene in insect cells resulted in secret ion of the protein product, Including both MTP subunits with apoB48 an d oleic acid specifically increased apoB48 secretion 8-fold over indiv idual subunits alone. To assess whether specific regions of apoB are n ecessary for MTP responsiveness, nine apoB segments were expressed. Th ese included NH2-terminal segments as well as internal and COOH-termin al regions of apoB fused with a heterologous signal sequence, ApoB seg ments containing the NH2-terminal 17% of the protein mere secreted and responded to NTP activity; however, a segment containing only the NH2 -terminal 17% of the protein was not significantly responsive to MTP. Segments lacking the NH2 terminus were not MTP-responsive, and five of six of these proteins were trapped intracellularly but, in certain ca ses, could be rescued by fusion to apoB17. These results suggest that the NH2 terminus of apoB is necessary but not sufficient for MTP respo nsiveness.