Partner: Hanna Fabczak

Nencki Institute of Experimental Biology, Polish Academy of Sciences (PL)

Recent publications
1.Góral A., Bieganowski P., Prus W., Krzemień-Ojak Ł., Kądziołka B., Fabczak H., Filipek A., Calcyclin Binding Protein/Siah-1 Interacting Protein Is a Hsp90 Binding Chaperone, PLOS ONE, ISSN: 1932-6203, DOI: 10.1371/journal.pone.0156507, Vol.11, No.6, pp.e0156507-1-18, 2016
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.

Keywords:

Chaperone proteins, Luciferase, Phosphatases, Enzyme-linked immunoassays, Recombinant proteins, Ligation assay, Plasmid construction, Luciferase assay

Affiliations:
Góral A.-other affiliation
Bieganowski P.-Mossakowski Medical Research Centre, Polish Academy of Sciences (PL)
Prus W.-other affiliation
Krzemień-Ojak Ł.-other affiliation
Kądziołka B.-other affiliation
Fabczak H.-Nencki Institute of Experimental Biology, Polish Academy of Sciences (PL)
Filipek A.-Nencki Institute of Experimental Biology, Polish Academy of Sciences (PL)
2.Bregier C., Kupikowska-Stobba B., Fabczak H., Fabczak S., CCT chaperonins and their cochaperons, Postępy Biochemii, ISSN: 0032-5422, Vol.54, No.1, pp.64-70, 2008