Stimulation of Akt poly-ubiquitination and proteasomal degradation in P388D1 cells by 7-ketocholesterol and 25-hydroxycholesterol.
Akt plays a role in protecting macrophages from apoptosis induced by some oxysterols. Previously we observed enhanced degradation of Akt in P388D1 moncocyte/macrophages following treatment with 25-hydroxycholesterol (25-OH) or 7-ketocholesterol (7-KC). In the present report we examine the role of the ubiquitin proteasomal pathway in this process. We show that treatment with 25-OH or 7-KC results in the accumulation of poly-ubiquitinated Akt, an effect that is enhanced by co-treatment with the proteasome inhibitor MG-132. Modification of Akt by the addition of a Gly-Ala repeat (GAr), a domain known to block ubiquitin-dependent targeting of proteins to the proteasome, resulted in a chimeric protein that is resistant to turn-over induced by 25-OH or 7-KC and provides protection from apoptosis induced by these oxysterols. These results uncover a new aspect of oxysterol regulation of Akt in macrophages; oxysterol-stimulated poly-ubiquitination of Akt and degradation by the proteasomal pathway.
Liu
June
J
Department of Urology, University of Pittsburg School of Medicine, Pittsburg, PA 15232, USA.
Netherland
Courtney
C
Pickle
Theresa
T
Sinensky
Michael S
MS
Thewke
Douglas P
DP
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001483:Base Sequence; D045744:Cell Line, Tumor; D006888:Hydroxycholesterols / Q000378:metabolism / Q000494:pharmacology*; D007653:Ketocholesterols / Q000378:metabolism / Q000494:pharmacology*; D007941:Leukemia P388; D007976:Leupeptins / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D010957:Plasmids / Q000235:genetics; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D054875:Ubiquitination / Q000187:drug effects
D006888:Hydroxycholesterols; D007653:Ketocholesterols; D007976:Leupeptins; D011480:Protease Inhibitors; D011993:Recombinant Fusion Proteins; C007997:25-hydroxycholesterol; D051057:Proto-Oncogene Proteins c-akt; D046988:Proteasome Endopeptidase Complex; C003001:7-ketocholesterol; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde