A dominant-negative peroxisome proliferator-activated receptor gamma (PPARgamma) mutant is a constitutive repressor and inhibits PPARgamma-mediated adipogenesis.
The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) promotes adipocyte differentiation, exerts atherogenic and anti-inflammatory effects in monocyte/macrophages, and is believed to mediate the insulin-sensitizing action of antidiabetic thiazolidinedione ligands. As no complete PPARgamma antagonists have been described hitherto, we have constructed a dominant-negative mutant receptor to inhibit wild-type PPARgamma action. Highly conserved hydrophobic and charged residues (Leu(468) and Glu(471)) in helix 12 of the ligand-binding domain were mutated to alanine. This compound PPARgamma mutant retains ligand and DNA binding, but exhibits markedly reduced transactivation due to impaired coactivator (cAMP-response element-binding protein-binding protein and steroid receptor coactivator-1) recruitment. Unexpectedly, the mutant receptor silences basal gene transcription, recruits corepressors (the silencing mediator of retinoid and thyroid receptors and the nuclear corepressor) more avidly than wild-type PPARgamma, and exhibits delayed ligand-dependent corepressor release. It is a powerful dominant-negative inhibitor of cotransfected wild-type receptor action. Furthermore, when expressed in primary human preadipocytes using a recombinant adenovirus, this PPARgamma mutant blocks thiazolidinedione-induced differentiation, providing direct evidence that PPARgamma mediates adipogenesis. Our observations suggest that, as in other mutant nuclear receptor contexts (acute promyelocytic leukemia, resistance to thyroid hormone), dominant-negative inhibition by PPARgamma is linked to aberrant corepressor interaction. Adenoviral expression of this mutant receptor is a valuable means to antagonize PPARgamma signaling.
Gurnell
M
M
Department of Medicine, University of Cambridge, Addenbrooke's Hospital, Cambridge CB2 2QQ, United Kingdom.
Wentworth
J M
JM
Agostini
M
M
Adams
M
M
Collingwood
T N
TN
Provenzano
C
C
Browne
P O
PO
Rajanayagam
O
O
Burris
T P
TP
Schwabe
J W
JW
Lazar
M A
MA
Chatterjee
V K
VK
D000256:Adenoviridae / Q000378:metabolism; D017667:Adipocytes / Q000378:metabolism*; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D005799:Genes, Dominant; D005822:Genetic Vectors; D006801:Humans; D008024:Ligands; D008954:Models, Biological; D009154:Mutation; D056985:Nuclear Receptor Co-Repressor 2; D010957:Plasmids; D011233:Precipitin Tests; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000502:physiology*; D012097:Repressor Proteins / Q000235:genetics; D000077154:Rosiglitazone; D013844:Thiazoles / Q000494:pharmacology; D045162:Thiazolidinediones; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic; D014162:Transfection
D004268:DNA-Binding Proteins; D008024:Ligands; C539130:NCOR2 protein, human; D056985:Nuclear Receptor Co-Repressor 2; D018160:Receptors, Cytoplasmic and Nuclear; D012097:Repressor Proteins; D013844:Thiazoles; D045162:Thiazolidinediones; D014157:Transcription Factors; D000077154:Rosiglitazone; C089946:2,4-thiazolidinedione