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|Title: ||Molecular Mechanisms of Intracellular Apolipoprotein B100 Degradation|
|Authors: ||Rutledge, Angela|
|Advisor: ||Adeli, Khosrow|
|Department: ||Laboratory Medicine and Pathobiology|
|Issue Date: ||22-Feb-2011|
|Abstract: ||Apolipoprotein B100 (apoB), a necessary component of very low density lipoproteins, is overproduced in metabolic disorders such as type II diabetes, increasing the risk of atherosclerosis. ApoB production is regulated by intracellular degradation of the protein by proteasomes, ER-60-dependent degradation, or post-endoplasmic reticulum (ER) pre-secretory proteolysis (PERPP)/autophagy. The first aim of the present study was to understand how apoB is delivered from the ER to cytosolic proteasomes. We investigated the roles of cellular factors BiP, Derlin-1, VIMP, Hrd1, and p97 in targeting apoB to proteasomes. Co-immunoprecipitation experiments indicated that these factors associate with apoB. Use of dominant negative p97 adenoviruses led to ER accumulation of apoB, demonstrating that p97 is required to extract apoB from the ER. Therefore, these factors appear to be important for proteasomal degradation of apoB.
ER-60, an ER protein with chaperone and cysteine protease activities, promotes apoB degradation through an unclear mechanism. To determine which property of ER-60 is involved, we used adenoviruses encoding wild-type ER-60 or a mutant form that lacked cysteine protease activity. Overexpression of wild-type, but not mutant, ER-60 induced apoB degradation, suggesting that ER-60 may function as a cysteine protease in apoB degradation.
We then investigated post-translational degradation of apoB in cells undergoing ER stress induced by glucosamine treatment. Treatment of hepatocytes with glucosamine induced a significant loss of cellular apoB. We speculated that oxidative stress and subsequent degradation by PERPP might be responsible. Glucosamine treatment appeared to cause only a slight increase in oxidative stress, but treatment with the antioxidants desferrioxamine or vitamin E blocked glucosamine-induced apoB degradation, suggesting involvement of PERPP/autophagy.
Overall, these studies have increased our understanding of the molecular mechanisms of intracellular apoB degradation and led to identification of critical factors and pathways that may be explored in the future to control hepatic apoB production in metabolic disease.|
|Appears in Collections:||Doctoral|
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