Novel Hsp90 partners discovered using complementary proteomic approaches

Abstract

Hsp90 is an essential eukaryotic molecular chaperone that stabilizes a large set of client proteins, many of which are involved in various cellular signaling pathways. The current list of Hsp90 interactors comprises about 200 proteins and this number is growing steadily. In this paper, we report on the application of three complementary proteomic approaches directed towards identification of novel proteins that interact with Hsp90. These methods are coimmunoprecipitation, pull down with biotinylated geldanamycin, and immobilization of Hsp90β on sepharose. In all, this study led to the identification of 42 proteins, including 18 proteins that had not been previously characterized as Hsp90 interactors. These novel Hsp90 partners not only represent abundant protein species, but several proteins were identified at low levels, among which signaling kinase Cdk3 and putative transcription factor tripartite motif-containing protein 29. Identification of tetratricopeptide-repeat-containing mitochondrial import receptor protein Tom34 suggests the involvement of Hsp90 in the early steps of translocation of mitochondrial preproteins. Taken together, our data expand the knowledge of the Hsp90 interactome and provide a further step in our understanding of the Hsp90 chaperone system.

The Assembly and Intermolecular Properties of the Hsp70-Tomm34-Hsp90 Molecular Chaperone Complex

“…Later studies showed that Tomm34 is mainly localized in cytosol and functions as a component of the large chaperone complex, which shuttles proteins from ribosomes to mitochondria (14,16). Recently, Tomm34 has been identified as an Hsp70 and Hsp90 co-chaperone (16,22,23), and, in addition to HOP (12), Tomm34 can serve as a scaffold protein by binding both Hsp70 and Hsp90. Therefore, the presented work is focused on the analysis of complex formation between full-length Tomm34 and its individual domains with Hsp70 and Hsp90 to disclose the structural relations between these proteins.…”

“…Chaperones-Tomm34 protein was described previously as an Hsp90-interacting protein (22,23). Recently, Faou et al (16) reported Tomm34 protein as a co-chaperone able to bind both Hsp70 and Hsp90 chaperones.…”

A Proteomic Investigation of Ligand-dependent HSP90 Complexes Reveals CHORDC1 as a Novel ADP-dependent HSP90-interacting Protein

“…and Geldanamycin-Several recent studies presented profiles of the human HSP90 interactome using immunopurifications and affinity capture with immobilized HSP90 (23,27). These studies effectively identified several new and novel HSP90 interactors; however, these experiments were not designed to identify interactions that are sensitive to HSP90 ligands.…”

“…Several groups have recently reported proteomics studies of the HSP90 interactome; however, these studies were focused on documenting HSP90 interactions under single experimental conditions with isolated proteins or were not designed to study how the dynamic interactions of this chaperone complex are influenced by HSP90 ligands (21)(22)(23)(24)(25)(26)(27). To gain proteome-wide resolution on the effects of ligands on the HSP90 interactome, we affinity-purified human HSP90 complexes from HEK293T cells in the presence of excess ATP, ADP, or the HSP90 inhibitor geldanamycin and used LC-MS/MS and spectral counting to quantify the effects of these ligands on the constituents of the HSP90 interactome.…”

Four-colour FRET reveals directionality in the Hsp90 multicomponent machinery

“…It supports the folding and maturation of a multitude of client proteins 8 . Many of these clients are involved in cell cycle and regulation, which makes Hsp90 an interesting cancer drug target [9][10][11][12][13] .…”