Apolipoprotein B100 exit from the endoplasmic reticulum (ER) is COPII-dependent, and its lipidation to very low density lipoprotein occurs post-ER.
Hepatic apolipoprotein B100 (apoB100) associates with lipids to form dense lipoprotein particles in the endoplasmic reticulum (ER) and is further lipidated to very low density lipoproteins (VLDL). Because the VLDL diameter can exceed 200 nm, classical ER-derived vesicles may be unable to accommodate VLDLs. Using hepatic membranes and cytosol to reconstitute ER budding, apoB100-containing vesicles sedimented distinct from those harboring more typical cargo but contained Sec23. Moreover, ER exit of apoB was inhibited by dominant-negative Sar1. Budding required Sar1 regardless of whether oleic acid (OA) was added to stimulate apoB lipidation; therefore, either large apoB100-lipoproteins reside in secretory vesicles, or full lipidation occurs post-ER. Using membranes from cells incubated in the presence or absence of OA, we determined that apoB100-lipoproteins in ER vesicles had not become lipidated to VLDLs. VLDL particles resided in the Golgi, but not the ER, after fractionation of OA-treated cells. We conclude that apoB100-lipoproteins exit the ER in COPII vesicles, but under conditions favorable for VLDL formation final lipid loading occurs post-ER.
Gusarova
Viktoria
V
Department of Medicine and The Cardiovascular Institute, Mount Sinai School of Medicine, New York, New York 10029, USA.
Brodsky
Jeffrey L
JL
Fisher
Edward A
EA
D000818:Animals; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B / Q000378:metabolism*; D015153:Blotting, Western; D022181:COP-Coated Vesicles / Q000737:chemistry*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002474:Cell-Free System; D002499:Centrifugation, Density Gradient; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D020690:GTPase-Activating Proteins; D005799:Genes, Dominant; D006056:Golgi Apparatus / Q000378:metabolism; D050356:Lipid Metabolism; D008079:Lipoproteins, VLDL / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008862:Microsomes, Liver / Q000378:metabolism; D009154:Mutation; D019301:Oleic Acid / Q000494:pharmacology; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D051381:Rats; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D053299:Apolipoprotein B-100; D001055:Apolipoproteins B; D020690:GTPase-Activating Proteins; D008079:Lipoproteins, VLDL; C079037:SEC23 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D019301:Oleic Acid