Interaction of nuclear receptors with hsp90 in living cells.
The ubiquitous heat shock protein 90 (hsp90) has been shown to participate directly in the function of a wide variety of cellular signal transduction components, including steroid receptors (SRs). However, there is still no direct evidence for an in vivo association of SRs with hsp90. This study utilizes the mammalian two-hybrid system to study the ability of hsp90 to interact with various (non)liganded nuclear receptors (NRs) in vivo in mammalian cells. As bait, we used ligand-binding domain (LBD) of various NRs fused with the GAL4-DBD. hsp90/Receptor interactions were monitored in COS cells. When NR-LBDs were co-transfected along with hsp90/VP16, none (RxR(2)-LBD) or only minimal (SR-LBDs) transcription inductions were observed (1.9-4.7-fold) in the absence of ligand. Addition of ligand further abolished the observed minimal induction. As a positive control for interaction we used TIF-2, which interacts with SRs in a ligand inducible manner. When co-transfected with NR-LBDs in the absence of ligand TIF-2/VP16 induced minimal activation of transcription (1.6-4.5-fold) that was comparable to the activation induced by the NR-LBDs. In contrast, in the presence of the ligand, the activation ranged between 62- and 134-fold depending on the receptor. The results suggest that the interaction of SRs with the hsp90 is minimal when compared to a bona fide type of interaction with the co-factors.
Manninen
Tommi
T
Department of Cell Biology, Tampere Graduate School in Biomedicine, Medical School, University of Tampere, 33014 Tampere, Finland. tmannine@users.csc.fi
Purmonen
Sami
S
Ylikomi
Timo
T
D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002645:Chickens; D002522:Chlorocebus aethiops; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D008024:Ligands; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D011987:Receptors, Steroid / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection
D018841:HSP90 Heat-Shock Proteins; D008024:Ligands; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins