Up-regulation of the clusterin gene after proteotoxic stress: implication of HSF1-HSF2 heterocomplexes.
Clusterin is a secreted protein chaperone up-regulated in several pathologies, including cancer and neurodegenerative diseases. The present study shows that accumulation of aberrant proteins, caused by the proteasome inhibitor MG132 or the incorporation of the amino acid analogue AZC (L-azetidine-2-carboxylic acid), increased both clusterin protein and mRNA levels in the human glial cell line U-251 MG. Consistently, MG132 treatment was capable of stimulating a 1.3 kb clusterin gene promoter. Promoter deletion and mutation studies revealed a critical MG132-responsive region between -218 and -106 bp, which contains a particular heat-shock element, named CLE for 'clusterin element'. Gel mobility-shift assays demonstrated that MG132 and AZC treatments induced the formation of a protein complex that bound to CLE. As shown by supershift and chromatin-immunoprecipitation experiments, CLE is bound by HSF1 (heat-shock factor 1) and HSF2 upon proteasome inhibition. Furthermore, co-immunoprecipitation assays indicated that these two transcription factors interact. Gel-filtration analyses revealed that the HSF1-HSF2 heterocomplexes bound to CLE after proteasome inhibition have the same apparent mass as HSF1 homotrimers after heat shock, suggesting that HSF1 and HSF2 could heterotrimerize. Therefore these studies indicate that the clusterin is a good candidate to be part of a cellular defence mechanism against neurodegenerative diseases associated with misfolded protein accumulation or decrease in proteasome activity.
Loison
Fabien
F
Information et Programmation Cellulaire, UMR CNRS 6026, Interactions Cellulaires et Moléculaires, IFR 140 - Génétique Fonctionnelle Agronomie et Santé, Université de Rennes 1, France.
Debure
Laure
L
Nizard
Philippe
P
le Goff
Pascale
P
Michel
Denis
D
le Dréan
Yves
Y
D000818:Animals; D001383:Azetidinecarboxylic Acid / Q000494:pharmacology; D051152:Clusterin / Q000235:genetics* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D000076249:Heat Shock Transcription Factors; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D046912:Multiprotein Complexes / Q000378:metabolism; D061988:Proteasome Inhibitors; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D050436:Regulatory Elements, Transcriptional / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D015854:Up-Regulation / Q000235:genetics*
D051152:Clusterin; D004268:DNA-Binding Proteins; C000619706:HSF1 protein, human; D000076249:Heat Shock Transcription Factors; C000619709:Hsf1 protein, rat; D007976:Leupeptins; D046912:Multiprotein Complexes; D061988:Proteasome Inhibitors; D012333:RNA, Messenger; D014157:Transcription Factors; D001383:Azetidinecarboxylic Acid; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde