Ontology highlight
ABSTRACT:
PROVIDER: PRJNA221226 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR1001900.fastq.gz | Fastqsanger.gz | |||
| SRR1001891.fastq.gz | Fastqsanger.gz | |||
| SRR1001892.fastq.gz | Fastqsanger.gz | |||
| SRR1001893.fastq.gz | Fastqsanger.gz | |||
| SRR1001894.fastq.gz | Fastqsanger.gz |
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Project description:The orchestration of protein folding and maintenance by molecular chaperones is crucial for cellular homeostasis. Among them, Hsp70 and its co-chaperone Hsp40 form a central machinery that remodels client proteins involved in numerous biological processes. Despite their essential role, how Hsp70 and Hsp40 assemble into a functional complex has remained elusive. Here, we integrated cryo-electron microscopy and nuclear magnetic resonance spectroscopy to determine the architecture of the full-length Hsp70-Hsp40 machinery. The structure of the complex in a physiological inactive state reveals distinct inhibitory mechanisms. In the active state, a new set of interactions is formed wherein the Hsp40 G/F-rich region acts as a pseudo-substrate and binds to the Hsp70, thereby directly modulating the refolding process. The G/F region maintains an Hsp40 autoinhibited state; upon binding to Hsp70, this state is disrupted, exposing a cryptic client-binding site and enabling interaction with protein clients to facilitate their refolding. The transition between these states is central to regulating the machinery’s refolding efficiency. Disrupting the Hsp40 autoinhibited state or the G/F–Hsp70 interaction leads to functional impairments that trigger a pronounced heat shock response in cells as a compensatory mechanism. Our findings provide crucial insights into the regulatory dynamics of this essential chaperone machinery, with broad implications for understanding protein homeostasis and the cellular response to stress.
2025-10-15 | GSE305556 | GEO
Project description:One-third of the human proteome is comprised of membrane proteins, which are particularly vulnerable to misfolding and often require folding assistance by molecular chaperones. Calnexin (CNX), which engages client proteins via its sugar-binding lectin domain, is one of the most abundant ER chaperones, and plays an important role in membrane protein biogenesis. Based on mass spectrometric analyses, we here show that calnexin interacts with a large number of nonglycosylated membrane proteins, indicative of additional nonlectin binding modes. We find that calnexin preferentially bind misfolded membrane proteins and that it uses its single transmembrane domain (TMD) for client recognition. Combining experimental and computational approaches, we systematically dissect signatures for intramembrane client recognition by calnexin, and identify sequence motifs within the calnexin TMD region that mediate client binding. Building on this, we show that intramembrane client binding potentiates the chaperone functions of calnexin. Together, these data reveal a widespread role of calnexin client recognition in the lipid bilayer, which synergizes with its established lectin-based substrate binding. Molecular chaperones thus can combine different interaction modes to support the biogenesis of the diverse eukaryotic membrane proteome.
2022-11-09 | PXD031140 | Pride
Project description:Cancer cells exhibit accelerated protein production to accommodate their high metabolic demands. This elevated protein load creates a dependency on endoplasmic reticulum-resident proteins and chaperones, which are required to maintain proteostasis. In this study, we identify the protein disulfide isomerases (PDIs) PDIA1 and PDIA5, which play a critical role in folding of client proteins in the ER, as important regulators of prostate cancer growth and therapy response. PDIA1 and PDIA5 are upregulated in prostate cancer and induced by the androgen receptor (AR) signalling axis. Genetic or pharmacological targeting of PDIA1/PDIA5 causes mitochondrial dysfunction, growth inhibition and apoptosis of prostate cancer cells in vitro and in vivo. Loss of PDIA1/PDIA5 activity leads to AR ubiquitination and degradation, revealing the existence of a feedback loop between these chaperones and the AR pathway. Mechanistically, PDIA1/PDIA5 regulate AR stability by catalysing disulfide bond formation in AR, an activity that requires cysteines 670 and 845 in AR’s ligand-binding domain. Importantly, targeting PDIAs sensitizes prostate cancer cells to the AR antagonist, enzalutamide. Collectively, this study reveals a novel mechanism governing AR proteostasis in prostate cancer and positions PDIA1/5 as viable therapeutic targets.
2025-10-14 | GSE288596 | GEO
Project description:Molecular chaperones such as heat-shock proteins (HSPs) help in protein folding and complex assembly processes. Their role in cytosol has been very well elucidated. Chaperones are also present in the nucleus, a compartment where proteins enter after being fully folded in the cytosol, raising an important question about chaperones function in this compartment. We have performed a systematic analysis of nuclear heat- shock protein 90 to identify regulatory functions of this chaperone in the nucleus. Combining physical and genetic interactomes with a cancer co-expression screen allowed us to define 'core functional interactors' of nuclear HSP90 consisting of five proteins. Using transcriptional studies we identified Host Cell Factor C1 (HCFC1) as a metazoan-specific transcriptional regulator that depends on HSP90 for its stability in the nucleus. We found that HSP90 is required for optimal activity of HCFC1 in transcription. Thus our study provides the first global insight into the function of nuclear HSP90.
2019-11-08 | GSE126151 | GEO
Project description:The molecular chaperone heat shock protein 90 (HSP90) works in concert with its co-chaperones to stabilize its client proteins, which include several drivers of oncogenesis and malignant progression. Pharmacologic inhibitors of HSP90 are proposed to trigger widespread remodeling of cellular protein complexes, including dissociation of co-chaperones from HSP90, disruption of client protein signaling networks, and recruitment of the protein ubiquitination and degradation machinery. However, proteomic studies to date have focused on inhibitor-induced changes in total protein levels, often overlooking protein complex alterations. Here, we use size-exclusion chromatography in combination with mass spectrometry (SEC-MS) to characterize the changes in native protein complexes following treatment with the HSP90 inhibitor 17-AAG in the human HT29 colon adenocarcinoma cell line.
2022-12-20 | PXD033459 | Pride
Project description:Protein tyrosine kinases are involved in regulating growth and proliferation in cells and are often hyperactive in cancerous tissue. Tyrosine kinase inhibitors have been used to limit hyperactivity but become ineffective due to the appearance of resistance mutations. A common trait of hyperactive protein kinases is its strict client relationship with molecular chaperone Hsp90. However, the mechanism behind Hsp90 client kinase recognition is poorly understood. Here we measure the functional effect of thousands of single amino acid variants in the Src kinase domain to identify positions invovled in Hsp90 client recognition.
2023-05-22 | GSE218190 | GEO
Project description:Here, we present a systematic and quantitative test of the hypothesis that the composition and activities of the endoplasmic reticulum (ER) proteostasis network impact mutational tolerance of secretory pathway client proteins. We focus on influenza hemagluttinin (HA), a viral coat protein that folds in the host’s ER via a complex but well-characterized pathway. By integrating chemical methods to modulate the unfolded protein response with deep mutational scanning to assess mutational tolerance, we discover that upregulation of ER chaperones broadly enhances HA mutational tolerance across numerous sites and secondary/tertiary structure elements, including sites targeted by host antibodies. Remarkably, this host chaperone-enhanced mutational tolerance is observed at the same HA sites where mutational tolerance is most reduced by propagation at a fever-like temperature. Thus, host ER proteostasis mechanisms and temperature modulate HA mutational tolerance in opposite directions. This finding has important implications for influenza evolution, because influenza immune escape is contingent on HA possessing sufficient mutational tolerance to acquire antibody resistance while still maintaining the capacity to fold and function. More broadly, this work provides the first experimental evidence that the composition and activities of the ER proteostasis network critically define the mutational tolerance and, therefore, the evolution of secretory pathway client proteins.
2018-09-10 | GSE115168 | GEO
Project description:Heat shock protein-90 chaperone machinery is involved in the stability and activity of its client proteins. The chaperone function of Hsp90 is regulated by co-chaperones and post-translational modifications. Although structural evidence exists for Hsp90 interaction with clients, our understanding of the impact of Hsp90 chaperone function towards client activity in cells remains elusive. Here, we dissected the impact of newly identified co-chaperones in higher eukaryotes, FNIP1/2 (FNIPs) and Tsc1, towards Hsp90 client activity. Our data show that Tsc1 and FNIP2 form mutually exclusive complexes with FNIP1 and that unlike Tsc1, FNIP1/2 interact with the catalytic residue of Hsp90. Functionally, these co-chaperone complexes increase the affinity of the steroid hormone receptors glucocorticoid receptor and estrogen receptor to their ligands in vivo. Here, we provide a model for the responsiveness of the steroid hormone receptor activation upon ligand binding as a consequence of their association with specific Hsp90:co-chaperone subpopulations.
2023-05-30 | PXD030486 | Pride
Project description:Many 2-Cys-peroxiredoxins (2-Cys-Prxs) serve as dual-function proteins. Active as peroxidases under non-stress conditions, they convert into effective chaperones under stress conditions. While their peroxidase activity has been extensively studied and shown to involve cycles of redox-mediated oligomeric changes, the mechanisms by which 2-Cys-Prxs sense stress and convert into general chaperones remain to be defined. Here we focus on the Leishmania infantum mitochondrial 2-Cys-Prx (mTXNPx, Prx1m), which, in its reduced, decameric form, readily adopts chaperone function upon exposure to heat shock temperatures. This activity is crucial for parasite survival in mammalian hosts. Here we have determined the cryo-EM structure of mTXNPx in complex with a thermally unfolded client protein that identifies the flexible N-termini of mTXNPx to form a well-resolved central belt that contacts and encapsulates the unstructured client protein in the center of the decamer ring. In vivo cross-linking studies combined with quantitative in vitro cross-linking experiments further support these interactions, and demonstrate that mTXNPx decamers undergo substantial temperature-dependent structural rearrangements specifically at the dimer-dimer interfaces. These structural changes appear crucial for exposing chaperone client binding sites that are otherwise buried in the peroxidase-active protein. Based on this mechanism, we propose that mTXNPx is the founding member of heat-stress activated chaperones in parasitic mitochondria that facilitate the transition to warm-blooded host environments.
2019-02-19 | PXD010281 | Pride