Document Detail

Ubiquitination regulates the assembly of VLDL in HepG2 cells and is the committing step of the apoB-100 ERAD pathway.
MedLine Citation:
PMID:  21421992     Owner:  NLM     Status:  MEDLINE    
Apolipoprotein B-100 (apoB-100) is degraded by endoplasmic reticulum-associated degradation (ERAD) when lipid availability limits assembly of VLDLs. The ubiquitin ligase gp78 and the AAA-ATPase p97 have been implicated in the proteasomal degradation of apoB-100. To study the relationship between ERAD and VLDL assembly, we used small interfering RNA (siRNA) to reduce gp78 expression in HepG2 cells. Reduction of gp78 decreased apoB-100 ubiquitination and cytosolic apoB-ubiquitin conjugates. Radiolabeling studies revealed that gp78 knockdown increased secretion of newly synthesized apoB-100 and, unexpectedly, enhanced VLDL assembly, as the shift in apoB-100 density in gp78-reduced cells was accompanied by increased triacylglycerol (TG) secretion. To explore the mechanisms by which gp78 reduction might enhance VLDL assembly, we compared the effects of gp78 knockdown with those of U0126, a mitogen-activated protein kinase/ERK kinase1/2 inhibitor that enhances apoB-100 secretion in HepG2 cells. U0126 treatment increased secretion of both apoB100 and TG and decreased the ubiquitination and cellular accumu-lation of apoB-100. Furthermore, p97 knockdown caused apoB-100 to accumulate in the cell, but if gp78 was concomitantly reduced or assembly was enhanced by U0126 treatment, cellular apoB-100 returned toward baseline. This indicates that ubiquitination commits apoB-100 to p97-mediated retrotranslocation during ERAD. Thus, decreasing ubiquitination of apoB-100 enhances VLDL assembly, whereas improving apoB-100 lipidation decreases its ubiquitination, suggesting that ubiquitination has a regulatory role in VLDL assembly.
Eric A Fisher; Neeraj A Khanna; Roger S McLeod
Publication Detail:
Type:  Journal Article     Date:  2011-03-18
Journal Detail:
Title:  Journal of lipid research     Volume:  52     ISSN:  0022-2275     ISO Abbreviation:  J. Lipid Res.     Publication Date:  2011 Jun 
Date Detail:
Created Date:  2011-05-16     Completed Date:  2011-09-19     Revised Date:  2013-06-30    
Medline Journal Info:
Nlm Unique ID:  0376606     Medline TA:  J Lipid Res     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1170-80     Citation Subset:  IM    
Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 1X5.
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MeSH Terms
Adenosine Triphosphatases / antagonists & inhibitors,  genetics,  metabolism*
Apolipoprotein B-100* / biosynthesis,  secretion
Butadienes / pharmacology
Cholesterol, VLDL* / metabolism
Endoplasmic Reticulum / genetics,  metabolism
Enzyme Inhibitors / pharmacology
Gene Silencing / drug effects
Hep G2 Cells
Isotope Labeling
Mitogen-Activated Protein Kinase Kinases / antagonists & inhibitors,  genetics,  metabolism*
Nitriles / pharmacology
Nuclear Proteins / antagonists & inhibitors,  genetics,  metabolism*
Proteasome Endopeptidase Complex / genetics,  metabolism
Protein Biosynthesis / drug effects*
RNA, Small Interfering / metabolism,  pharmacology
Receptors, Autocrine Motility Factor
Receptors, Cytokine / antagonists & inhibitors,  genetics,  metabolism*
Triglycerides / metabolism
Ubiquitin / genetics,  metabolism*
Ubiquitin-Protein Ligases / antagonists & inhibitors,  genetics,  metabolism*
Ubiquitination / drug effects
Reg. No./Substance:
0/Apolipoprotein B-100; 0/Butadienes; 0/Cholesterol, VLDL; 0/Enzyme Inhibitors; 0/Nitriles; 0/Nuclear Proteins; 0/RNA, Small Interfering; 0/Receptors, Cytokine; 0/Triglycerides; 0/U 0126; 0/Ubiquitin; EC Protein Kinase Kinases; EC Endopeptidase Complex; EC 3.6.1.-/Adenosine Triphosphatases; EC 3.6.1.-/p97 ATPase; EC protein, human; EC, Autocrine Motility Factor; EC Ligases

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