Affinity purification reveals the association of WD40 protein constitutive photomorphogenic 1 with the hetero-oligomeric TCP-1 chaperonin complex in mammalian cells.
Constitutive photomorphogenic 1 (COP1), a protein composed of a RING finger, a coiled-coil domain and seven WD40 repeats, functions as an E3 ubiquitin ligase that targets key transcription factors for ubiquitination and degradation in both higher plants and mammalian cells. While COP1 is required for light-mediated development in plants, its mammalian counterpart has been implicated in tumorigenesis. We previously showed that COP1 forms high-molecular-weight complexes in mammalian cells. Here we report our attempts in characterizing the components of the mammalian COP1 complexes by affinity purification combined with mass spectral analysis. We find that both transiently and stably expressed COP1 associates with the hetero-oligomeric TCP-1 chaperonin complex (TRiC), heat shock protein 70 (Hsp70) and BAG-family molecular chaperone regulator-2 (BAG2). In addition, stably expressed COP1 binds to major vault protein (MVP) and translocated promoter region (Tpr). The TRiC/Hsp70 complex is known to interact with and assist in the folding of a number of WD40 proteins in Saccharomyces cerevisiae. The association of WD40 protein COP1 with TRiC/Hsp70 in mammalian cells suggests that facilitating the folding of WD40 proteins may be a conserved function for TRiC/Hsp70 from yeast to mammals.
Yi
Chunling
C
Department of Molecular, Cellular and Developmental Biology, Yale University, P.O. Box 208104, 165 Prospect Street, OML 352A, New Haven, CT 06520-8104, USA.
Li
Shuting
S
Wang
Jian
J
Wei
Ning
N
Deng
Xing Wang
XW
D000818:Animals; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D056404:Chaperonin Containing TCP-1; D018833:Chaperonins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods; D006801:Humans; D056511:Metallochaperones; D008969:Molecular Sequence Data; D011485:Protein Binding; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D044767:Ubiquitin-Protein Ligases
D029681:Arabidopsis Proteins; C112111:CIAO1 protein, human; D002352:Carrier Proteins; D056511:Metallochaperones; D011993:Recombinant Fusion Proteins; C471983:AT2G32950 protein, Arabidopsis; D044767:Ubiquitin-Protein Ligases; D056404:Chaperonin Containing TCP-1; D018833:Chaperonins