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ROLE OF APOLIPOPROTEIN A-V IN TRIGLYCERIDE METABOLISM

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abstract
ApoA-V, a 366 amino acid protein synthesized by the liver, was identified in 2001 by comparative sequence analysis. The APOA5 gene is located downstream of the AI/CIII/AIV gene cluster on human chromosome 11q23. Apolipoprotein A-V (apoA-V) plasma concentrations are low (~100-200 µg/L) in comparison to other apolipoproteins, yet exert significant effects on triglyceride homeostasis. To examine the basis for its low concentration in plasma, the secretion efficiency of apoA-V was measured in stably transfected McA-RH7777 rat hepatoma cells. Pulse-chase analysis revealed that only ~20% of newly synthesized apoA-V was secreted into culture medium within two hours post-synthesis. Similar results were obtained in transfected nonhepatic (CHO) cells. In neither cell system was apoA-V found associated with cell surface heparin sulfate proteoglycans. Similar results were also obtained when endogenous mouse apoA-V was analyzed in primary hepatocytes. In addition to its low secretion efficiency, the electrophoretic banding pattern of apoA-V suggested that the protein is susceptible to intracellular proteolysis with ~60% undergoing presecretory turnover within 2 hours post-synthesis. To study the regulation of apoA-V by lipid synthesis, stably transfected McA-RH7777 cells were treated with 0.8 mM sodium oleate. The resulting increase in triglyceride synthesis resulted in a dramatic decline in apoA-V secretion, a corresponding increase in cell-associated apoA-V, but no change in total apoA-V recovery. Reduced secretion was accompanied by movement of apoA-V onto cytosolic lipid droplets, as evidenced by apoA-V's colocalization with ADRP, a lipid droplet marker, and its flotation during sucrose gradient ultracentrifugation. The possible intracellular role of apoA-V in the regulation of apoB and TG secretion was examined by generating stably transfected doxycycline-inducible McA-RH7777 cells. Upon induction of apoA-V expression, a 32% decrease in apoB-100 secretion and a 21% decrease in triglyceride secretion were observed. In addition, apoA-V expression caused a 57% decrease in the apoA-V in the d<1.006 g/ml VLDL density fraction during equilibrium density gradient ultracentrifugation. These results indicate that 1) apoA-V inefficiently traffics within the secretory pathway but that its intracellular itinerary may be regulated by changes in TG synthesis and/or accumulation and 2) part of the effect of apoA-V on TG metabolism could be exerted at the level of hepatic TG mobilization and secretion.
subject
Apolipoprotein
Triglyceride
ApoA-V
ApoB
contributor
Blade, Anna (author)
Parks, Griffith (committee chair)
Shelness, Greg (committee member)
Parks, John (committee member)
Dawson, Paul (committee member)
Liqing, Yu (committee member)
Weinberg, Richard (committee member)
date
2009-05-28T15:57:27Z (accessioned)
2010-06-18T18:59:32Z (accessioned)
2009-05-28T15:57:27Z (available)
2010-06-18T18:59:32Z (available)
2009-05-28T15:57:27Z (issued)
degree
Molecular & Cellular Pathobiology (discipline)
identifier
http://hdl.handle.net/10339/14862 (uri)
language
en_US (iso)
publisher
Wake Forest University
rights
Release the entire work for access only to the Wake Forest University system for one year from the date below. After one year, release the entire work for access worldwide. (accessRights)
title
ROLE OF APOLIPOPROTEIN A-V IN TRIGLYCERIDE METABOLISM
type
Dissertation

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