Regulated Co-translational ubiquitination of apolipoprotein B100. A new paradigm for proteasomal degradation of a secretory protein.
Presentation of a wild-type secretory protein, apolipoprotein B100 (apoB), to the cytosol for ubiquitin-proteasome proteolysis has been observed in HepG2 cells. A currently accepted model for proteasomal degradation of secretory proteins is retrograde translocation of the substrate polypeptides from the lumen of endoplasmic reticulum (ER) back to the cytosol. In this report, we present evidence that newly synthesized apoB becomes exposed to the cytosol and targeted to the proteasomes in a co-translational manner. Thus, after protein translation was synchronized with puromycin, partially synthesized apoB polypeptides were found to be conjugated to ubiquitin. The magnitude of co-translational ubiquitination and subsequent degradation of apoB was increased when cells were pretreated with either herbimycin A to induce cytosolic Hsp70 or with an inhibitor of microsomal triglyceride transfer protein; both treatments impede translocation of nascent apoB across the ER membrane. These treatments also decreased secretion of apoB and increased its degradation via the ubiquitin-proteasome pathway. We suggest that translocation arrest with subsequent co-translational exposure to the cytosol provides an alternative model to explain how mammalian secretory proteins can overcome topological segregation by the ER membrane and undergo degradation by the ubiquitin-proteasome pathway.
Zhou
M
M
Department of Medicine, Columbia University, New York, New York 10032, USA.
Fisher
E A
EA
Ginsberg
H N
HN
D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B / Q000235:genetics* / Q000378:metabolism*; D016227:Benzoquinones; D016466:CHO Cells; D006528:Carcinoma, Hepatocellular; D006224:Cricetinae; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis; D006801:Humans; D047029:Lactams, Macrocyclic; D008113:Liver Neoplasms; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D011691:Puromycin / Q000494:pharmacology; D011809:Quinones / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis; D017828:Rifabutin / Q000031:analogs & derivatives; D014162:Transfection; D014407:Tumor Cells, Cultured; D014452:Ubiquitins / Q000378:metabolism*
D053299:Apolipoprotein B-100; D001055:Apolipoproteins B; D016227:Benzoquinones; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D018840:HSP70 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D009097:Multienzyme Complexes; D011809:Quinones; D011994:Recombinant Proteins; D014452:Ubiquitins; C067713:lactacystin; D017828:Rifabutin; D011691:Puromycin; C020754:herbimycin; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D000111:Acetylcysteine