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http://hdl.handle.net/10362/182771| Título: | The HSP90/R2TP Quaternary Chaperone Scaffolds Assembly of the TSC Complex |
| Autor: | Abéza, Claire Busse, Philipp Paiva, Ana C.F. Chagot, Marie Eve Schneider, Justine Robert, Marie Cécile Vandermoere, Franck Schaeffer, Christine Charpentier, Bruno Sousa, Pedro M.F. Bandeiras, Tiago M. Manival, Xavier Cianferani, Sarah Bertrand, Edouard Verheggen, Céline |
| Palavras-chave: | ATPases associated with diverse cellular activities (RUVBL1/2) heat shock protein 90 (HSP90) R2TP tuberous sclerosis complex (TSC) Biophysics Structural Biology Molecular Biology |
| Data: | 1-Dez-2024 |
| Resumo: | The R2TP chaperone is composed of the RUVBL1/RUVBL2 AAA+ ATPases and two adapter proteins, RPAP3 and PIH1D1. Together with HSP90, it functions in the assembly of macromolecular complexes that are often involved in cell proliferation. Here, proteomic experiments using the isolated PIH domain reveals additional R2TP partners, including the Tuberous Sclerosis Complex (TSC) and many transcriptional complexes. The TSC is a key regulator of mTORC1 and is composed of TSC1, TSC2 and TBC1D7. We show a direct interaction of TSC1 with the PIH phospho-binding domain of PIH1D1, which is, surprisingly, phosphorylation independent. Via the use of mutants and KO cell lines, we observe that TSC2 makes independent interactions with HSP90 and the TPR domains of RPAP3. Moreover, inactivation of PIH1D1 or the RUVBL1/2 ATPase activity inhibits the association of TSC1 with TSC2. Taken together, these data suggest a model in which the R2TP recruits TSC1 via PIH1D1 and TSC2 via RPAP3 and HSP90, and use the chaperone-like activities of RUVBL1/2 to stimulate their assembly. |
| Descrição: | Funding Information: This work was supported by grants from Centre National de la Recherche Scientifique (CNRS); Universit\u00E9 de Montpellier (UM); Universit\u00E9 de Lorraine (UL); the Agence Nationale de la Recherche [ANR-16-CE11-0032-04]; grants from the Ligue Nationale Contre le Cancer ('\u00E9quipe labellis\u00E9e'); Institut national du Cancer [INCa PLBio 2016-161]; Funda\u00E7\u00E3o para a Ci\u00EAncia e Tecnologia/Minist\u00E9rio da Ci\u00EAncia, Tecnologia e Ensino Superior (FCT/MCTES, Portugal) through national funds to iNOVA4Health (UIDB/ 04462/ 2020, UIDP/04462/2020) and the Associate Laboratory LS4FUT URE (LA/P/0087/2020). CA was supported by La ligue Contre le Cancer and Fondation pour la Recherche M\u00E9dicale. Mass spectrometry for SILAC-IP experiments was carried out using the facilities of the Montpellier Proteomics Platform (PPM, BioCampus Montpellier). We also thank MRI and MGC facilities of UMS Montpellier BioCampus. We thank C. Goujon for the gift of HEK 293T cells expressing Cas9 and Y. Abel for cloning of PIH1D1 KO cells. We thank B. Pradet-Balade, D. Helmlinger and P. Marin for their discussion on the project and M. Hourques and D. Soumayla Seyni for their help in Y2H and IP LUMIER assays. We also thank S. Boulon for reading of the manuscript. Funding Information: This work was supported by grants from Centre National de la Recherche Scientifique (CNRS); Universit\u00E9 de Montpellier (UM); Universit\u00E9 de Lorraine (UL); the Agence Nationale de la Recherche [ANR-16-CE11-0032-04] and (ANR-23-CE12-0022); grants from the Ligue Nationale Contre le Cancer ('\u00E9quipe labellis\u00E9e'); Institut national du Cancer [INCa PLBio 2016-161]; Funda\u00E7\u00E3o para a Ci\u00EAncia e Tecnologia/Minist\u00E9rio da Ci\u00EAncia, Tecnologia e Ensino Superior (FCT/MCTES, Portugal) through national funds to iNOVA4Health (UIDB/04462/2020, UIDP/04462/2020) and the Associate Laboratory LS4FUTURE (LA/P/0087/2020). CA was supported by La ligue Contre le Cancer and Fondation pour la Recherche M\u00E9dicale. Mass spectrometry for SILAC-IP experiments was carried out using the facilities of the Montpellier Proteomics Platform (PPM, BioCampus Montpellier). We also thank MRI and MGC facilities of UMS Montpellier BioCampus. We thank C. Goujon for the gift of HEK 293T cells expressing Cas9 and Y. Abel for cloning of PIH1D1 KO cells. We thank B. Pradet-Balade, D. Helmlinger and P. Marin for their discussion on the project and M. Hourques and D. Soumayla Seyni for their help in Y2H and IP LUMIER assays. We also thank S. Boulon for reading of the manuscript. C. Ab\u00E9za performed all experiments shown in Figures 1, 2, 3C, 5, 6 and 7. M-E. Chagot performed experiment shown in Figure 4A and P. Busse and A. C.F. Paiva in Figure 4B. M-C. Robert prepared the KO cell lines. F.Vandermoere analyzed proteomic data of Figure 6B and J. Schneider and C. Schaeffer proteomic data of Figures 1 and 3A. C. Ab\u00E9za and C. Verheggen prepared all the figures. C. Verheggen, E. Bertrand, B. Charpentier, P. Sousa, T. Bandeiras, X. Manival, S. Cianferani supervised the project. C. Verheggen and E. Bertrand designed the experiments and wrote the manuscript. Publisher Copyright: © 2024 The Author(s) |
| Peer review: | yes |
| URI: | http://hdl.handle.net/10362/182771 |
| DOI: | https://doi.org/10.1016/j.jmb.2024.168840 |
| ISSN: | 0022-2836 |
| Aparece nas colecções: | Home collection (ITQB) |
Ficheiros deste registo:
| Ficheiro | Descrição | Tamanho | Formato | |
|---|---|---|---|---|
| The_HSP90_R2TP_Quaternary_Chaperone.pdf | 2,93 MB | Adobe PDF | Ver/Abrir |
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