Analyse fonctionnelle de la phosphorylation du co-chaperon moléculaire BAG3 et de son action dans la morpho-dynamique des cellules mitotiques
|Abstract:||Cell division is the fundamental principle of life and is based on spectacular cellular architectural changes. Many of them are driven by the accurate remodeling of mechanosensitive actin-based structures. Growing evidence suggests a close relationship between protein quality control and the spatiotemporal regulation of actin remodeling, through mechanisms that would promote protein sequestration and/or degradation. Small heat shock proteins (HSPBs) are molecular chaperones that are an integral part of the protein quality control network, which contribute to maintain proteome homeostasis. They emerge as modulators of actin-based structures under physiological conditions and as guardians of the integrity of cytoskeletal structures under stress conditions. According to the prevailing model, the assembly of HSPBs into large oligomers confers them with the ability to sequester cellular components and prevent unspecific aggregation of damaged proteins. Nevertheless, their mode of action remains elusive: the observation that some HSPBs do not form oligomers suggests another mechanism of action for these HSPBs. This is the case for HSPB8, which forms a complex with the molecular co-chaperone BAG3. The working model of this thesis is based on the initial discovery in our laboratory of a new role for this complex during cell division: BAG3 facilitates the drastic remodeling of the actin cytoskeleton required for spindle positioning and proper segregation of chromosomes, in a manner that requires HSPB8. The aim of this thesis was to identify the mechanisms whereby such a function of the BAG3-HSPB8 chaperone complex is regulated, and to investigate how the complex can facilitate mitotic actin cytoskeleton remodeling. The work presented here provides evidence that the modulation of BAG3 mitotic functions depends on its phosphorylation by the mitotic kinase CDK1 at specific residues, Thr285 and Ser386, which confers differential activity on cell rounding versus mitotic spindle positioning. Evidence also suggests that BAG3 would be phosphorylated earlier in the G2/M phase, at Ser195, which would modulate its perinuclear enrichment. Our results suggest that these phosphorylations could be involved in defining specific protein associations, in a cell-cycle dependent manner. In addition, we found that mitotic entry is marked by the stimulation of BAG3'sassociation with proteins that organize the actin cytoskeleton, such as cortactin, as well as with protein quality control actors, notable, the autophagic receptor p62/SQSTM1 and the deacetylase HDAC6. Critically, BAG3 phosphorylation and its associations with mitotic protein partners rely on its binding to HSPB8. The results suggests a model whereby the BAG3-HSPB8 complex would regulate the molecular assembly of p62/SQSTM1 into mitotic bodies that could provide a platform to sequester and facilitate protein complex assembly implicated in mitotic actin cytoskeleton remodeling. Via this mechanism, BAG3-HSPB8 could limit branched actin polymerization that depends on Arp2/3 activity, by down-modulating HDAC6 deacetylase activity towards its substrate cortactin, a process that would facilitate mitotic cell rounding. Thus, our results highlight a central role of BAG3 phosphorylation in the modulation of its mitotic action, in close relationship with its partners HSPB8 and p62. Altogether, our data contribute to a better understanding of the molecular mechanisms by which the BAG3-HSPB8 chaperone complex orchestrates the dynamic remodeling of mitotic cell structures and thereby, facilitates the cell shape changes required for mitotic progression. This study has also identified new molecular targets of the chaperone complex there are, among others, involved in the dynamics of the actin cytoskeleton. Thus, this work offers new avenues of investigation regarding the development of pathologies associated with a deregulation of the BAG3-HSPB8 complex, particularly in tumor progression.|
|Document Type:||Thèse de doctorat|
|Open Access Date:||14 June 2021|
|Collection:||Thèses et mémoires|
All documents in CorpusUL are protected by Copyright Act of Canada.