title: Calcyclin Binding Protein/Siah-1 Interacting Protein Is a Hsp90 Binding Chaperone.
abstract: The Hsp90 chaperone activity is tightly regulated by interaction with many co-chaperones. Since CacyBP/SIP shares some sequence homology with a known Hsp90 co-chaperone, Sgt1, in this work we performed a set of experiments in order to verify whether CacyBP/SIP can interact with Hsp90. By applying the immunoprecipitation assay we have found that CacyBP/SIP binds to Hsp90 and that the middle (M) domain of Hsp90 is responsible for this binding. Furthermore, the proximity ligation assay (PLA) performed on HEp-2 cells has shown that the CacyBP/SIP-Hsp90 complexes are mainly localized in the cytoplasm of these cells. Using purified proteins and applying an ELISA we have shown that Hsp90 interacts directly with CacyBP/SIP and that the latter protein does not compete with Sgt1 for the binding to Hsp90. Moreover, inhibitors of Hsp90 do not perturb CacyBP/SIP-Hsp90 binding. Luciferase renaturation assay and citrate synthase aggregation assay with the use of recombinant proteins have revealed that CacyBP/SIP exhibits chaperone properties. Also, CacyBP/SIP-3xFLAG expression in HEp-2 cells results in the appearance of more basic Hsp90 forms in 2D electrophoresis, which may indicate that CacyBP/SIP dephosphorylates Hsp90. Altogether, the obtained results suggest that CacyBP/SIP is involved in regulation of the Hsp90 chaperone machinery.
authors.affiliation: Nencki Institute of Experimental Biology PAS, Warsaw, Poland.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Góral
authors.affiliation: Mossakowski Medical Research Centre PAS, Warsaw, Poland.
authors.forename: Paweł
authors.initials: P
authors.lastname: Bieganowski
authors.affiliation: Nencki Institute of Experimental Biology PAS, Warsaw, Poland.
authors.forename: Wiktor
authors.initials: W
authors.lastname: Prus
authors.affiliation: Nencki Institute of Experimental Biology PAS, Warsaw, Poland.
authors.forename: Łucja
authors.initials: Ł
authors.lastname: Krzemień-Ojak
authors.affiliation: Nencki Institute of Experimental Biology PAS, Warsaw, Poland.
authors.forename: Beata
authors.initials: B
authors.lastname: Kądziołka
authors.affiliation: Nencki Institute of Experimental Biology PAS, Warsaw, Poland.
authors.forename: Hanna
authors.initials: H
authors.lastname: Fabczak
authors.affiliation: Nencki Institute of Experimental Biology PAS, Warsaw, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Filipek
mesh_terms: D018797:Cell Cycle Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D000075383:S100 Calcium Binding Protein A6; D009418:S100 Proteins / Q000378:metabolism*; D015398:Signal Transduction; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D000098563:Seven in Absentia Proteins
chemical_list: D018797:Cell Cycle Proteins; D018841:HSP90 Heat-Shock Proteins; D009687:Nuclear Proteins; D000075383:S100 Calcium Binding Protein A6; D009418:S100 Proteins; C055351:S100A6 protein, human; D044767:Ubiquitin-Protein Ligases; D000098563:Seven in Absentia Proteins