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Anne Marie Cirinesi - One of the best experts on this subject based on the ideXlab platform.
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Structural insights into Chaperone addiction of toxin-antitoxin systems
Nature Communications, 2019Co-Authors: Valerie Guillet, Patricia Bordes, Cécile Bon, Julien Marcoux, Virginie Gervais, Ambre Julie Sala, Suzana Dos Reis, Nawel Slama, Israel Mares-mejía, Anne Marie CirinesiAbstract:SecB Chaperones assist protein export by binding both unfolded proteins and the SecA motor. Certain SecB homologs can also control toxin-antitoxin (TA) systems known to modulate bacterial growth in response to stress. In such TA-Chaperone (TAC) systems, SecB assists the folding and prevents degradation of the antitoxin, thus facilitating toxin inhibition. Chaperone dependency is conferred by a C-terminal extension in the antitoxin known as Chaperone addiction (ChAD) sequence, which makes the antitoxin aggregation-prone and prevents toxin inhibition. Using TAC of Mycobacterium tuberculosis, we present the structure of a SecB-like Chaperone bound to its ChAD peptide. We find differences in the binding interfaces when compared to SecB–SecA or SecB-preprotein complexes, and show that the antitoxin can reach a functional form while bound to the Chaperone. This work reveals how Chaperones can use discrete surface binding regions to accommodate different clients or partners and thereby expand their substrate repertoire and functions.
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directed evolution of secb Chaperones toward toxin antitoxin systems
Proceedings of the National Academy of Sciences of the United States of America, 2017Co-Authors: Ambre Sala, Marie Pierre Castaniecornet, Anne Marie Cirinesi, Patricia Bordes, Nawel Slama, Sara Ayala, Samuel Tranier, Michele Coddeville, Lionel Mourey, Pierre GenevauxAbstract:SecB Chaperones assist protein export in bacteria. However, certain SecB family members have diverged to become specialized toward the control of toxin-antitoxin (TA) systems known to promote bacterial adaptation to stress and persistence. In such tripartite TA-Chaperone (TAC) systems, the Chaperone was shown to assist folding and to prevent degradation of its cognate antitoxin, thus facilitating inhibition of the toxin. Here, we used both the export Chaperone SecB of Escherichia coli and the tripartite TAC system of Mycobacterium tuberculosis as a model to investigate how generic Chaperones can specialize toward the control of TA systems. Through directed evolution of SecB, we have identified and characterized mutations that specifically improve the ability of SecB to control our model TA system without affecting its function in protein export. Such a remarkable plasticity of SecB Chaperone function suggests that its substrate binding surface can be readily remodeled to accommodate specific clients.
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hsp33 controls elongation factor tu stability and allows escherichia coli growth in the absence of the major dnak and trigger factor Chaperones
Journal of Biological Chemistry, 2012Co-Authors: Nicolas Bruel, Marie Pierre Castaniecornet, Anne Marie Cirinesi, Gregory Koningstein, Costa Georgopoulos, Joen Luirink, Pierre GenevauxAbstract:Abstract Intracellular de novo protein folding is assisted by cellular networks of molecular Chaperones. In Escherichia coli, cooperation between the Chaperones trigger factor (TF) and DnaK is central to this process. Accordingly, the simultaneous deletion of both Chaperone-encoding genes leads to severe growth and protein folding defects. Herein, we took advantage of such defective phenotypes to further elucidate the interactions of Chaperone networks in vivo. We show that disruption of the TF/DnaK Chaperone pathway is efficiently rescued by overexpression of the redox-regulated Chaperone Hsp33. Consistent with this observation, the deletion of hslO, the Hsp33 structural gene, is no longer tolerated in the absence of the TF/DnaK pathway. However, in contrast with other Chaperones like GroEL or SecB, suppression by Hsp33 was not attributed to its potential overlapping general Chaperone function(s). Instead, we show that overexpressed Hsp33 specifically binds to elongation factor-Tu (EF-Tu) and targets it for degradation by the protease Lon. This synergistic action of Hsp33 and Lon was responsible for the rescue of bacterial growth in the absence of TF and DnaK, by presumably restoring the coupling between translation and the downstream folding capacity of the cell. In support of this hypothesis, we show that overexpression of the stress-responsive toxin HipA, which inhibits EF-Tu, also rescues bacterial growth and protein folding in the absence of TF and DnaK. The relevance for such a convergence of networks of Chaperones and proteases acting directly on EF-Tu to modulate the intracellular rate of protein synthesis in response to protein aggregation is discussed.
Pierre Genevaux - One of the best experts on this subject based on the ideXlab platform.
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directed evolution of secb Chaperones toward toxin antitoxin systems
Proceedings of the National Academy of Sciences of the United States of America, 2017Co-Authors: Ambre Sala, Marie Pierre Castaniecornet, Anne Marie Cirinesi, Patricia Bordes, Nawel Slama, Sara Ayala, Samuel Tranier, Michele Coddeville, Lionel Mourey, Pierre GenevauxAbstract:SecB Chaperones assist protein export in bacteria. However, certain SecB family members have diverged to become specialized toward the control of toxin-antitoxin (TA) systems known to promote bacterial adaptation to stress and persistence. In such tripartite TA-Chaperone (TAC) systems, the Chaperone was shown to assist folding and to prevent degradation of its cognate antitoxin, thus facilitating inhibition of the toxin. Here, we used both the export Chaperone SecB of Escherichia coli and the tripartite TAC system of Mycobacterium tuberculosis as a model to investigate how generic Chaperones can specialize toward the control of TA systems. Through directed evolution of SecB, we have identified and characterized mutations that specifically improve the ability of SecB to control our model TA system without affecting its function in protein export. Such a remarkable plasticity of SecB Chaperone function suggests that its substrate binding surface can be readily remodeled to accommodate specific clients.
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hsp33 controls elongation factor tu stability and allows escherichia coli growth in the absence of the major dnak and trigger factor Chaperones
Journal of Biological Chemistry, 2012Co-Authors: Nicolas Bruel, Marie Pierre Castaniecornet, Anne Marie Cirinesi, Gregory Koningstein, Costa Georgopoulos, Joen Luirink, Pierre GenevauxAbstract:Abstract Intracellular de novo protein folding is assisted by cellular networks of molecular Chaperones. In Escherichia coli, cooperation between the Chaperones trigger factor (TF) and DnaK is central to this process. Accordingly, the simultaneous deletion of both Chaperone-encoding genes leads to severe growth and protein folding defects. Herein, we took advantage of such defective phenotypes to further elucidate the interactions of Chaperone networks in vivo. We show that disruption of the TF/DnaK Chaperone pathway is efficiently rescued by overexpression of the redox-regulated Chaperone Hsp33. Consistent with this observation, the deletion of hslO, the Hsp33 structural gene, is no longer tolerated in the absence of the TF/DnaK pathway. However, in contrast with other Chaperones like GroEL or SecB, suppression by Hsp33 was not attributed to its potential overlapping general Chaperone function(s). Instead, we show that overexpressed Hsp33 specifically binds to elongation factor-Tu (EF-Tu) and targets it for degradation by the protease Lon. This synergistic action of Hsp33 and Lon was responsible for the rescue of bacterial growth in the absence of TF and DnaK, by presumably restoring the coupling between translation and the downstream folding capacity of the cell. In support of this hypothesis, we show that overexpression of the stress-responsive toxin HipA, which inhibits EF-Tu, also rescues bacterial growth and protein folding in the absence of TF and DnaK. The relevance for such a convergence of networks of Chaperones and proteases acting directly on EF-Tu to modulate the intracellular rate of protein synthesis in response to protein aggregation is discussed.
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The Hsp70 Chaperone machines of Escherichia coli: a paradigm for the repartition of Chaperone functions
Molecular microbiology, 2007Co-Authors: Pierre Genevaux, Costa Panos Georgopoulos, William L. KelleyAbstract:Molecular Chaperones are highly conserved in all free-living organisms. There are many types of Chaperones, and most are conveniently grouped into families. Genome sequencing has revealed that many organisms contain multiple members of both the DnaK (Hsp70) family and their partner J-domain protein (JDP) coChaperone, belonging to the DnaJ (Hsp40) family. Escherichia coli K-12 encodes three Hsp70 genes and six JDP genes. The coexistence of these Chaperones in the same cytosol suggests that certain Chaperone-coChaperone interactions are permitted, and that Chaperone tasks and their regulation have become specialized over the course of evolution. Extensive genetic and biochemical analyses have greatly expanded knowledge of Chaperone tasking in this organism. In particular, recent advances in structure determination have led to significant insights of the underlying complexities and functional elegance of the Hsp70 Chaperone machine.
Anneclaude Gingras - One of the best experts on this subject based on the ideXlab platform.
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a quantitative Chaperone interaction network reveals the architecture of cellular protein homeostasis pathways
Cell, 2014Co-Authors: Mikko Taipale, George Tucker, Jian Peng, Irina Krykbaeva, Zhen Yuan Lin, Brett Larsen, Hyungwon Choi, Bonnie Berger, Anneclaude GingrasAbstract:Chaperones are abundant cellular proteins that promote the folding and function of their substrate proteins (clients). In vivo, Chaperones also associate with a large and diverse set of cofactors (coChaperones) that regulate their specificity and function. However, how these coChaperones regulate protein folding and whether they have Chaperone-independent biological functions is largely unknown. We combined mass spectrometry and quantitative high-throughput LUMIER assays to systematically characterize the Chaperone-coChaperone-client interaction network in human cells. We uncover hundreds of Chaperone clients, delineate their participation in specific coChaperone complexes, and establish a surprisingly distinct network of protein-protein interactions for coChaperones. As a salient example of the power of such analysis, we establish that NUDC family coChaperones specifically associate with structurally related but evolutionarily distinct β-propeller folds. We provide a framework for deciphering the proteostasis network and its regulation in development and disease and expand the use of Chaperones as sensors for drug-target engagement.
Johannes Buchner - One of the best experts on this subject based on the ideXlab platform.
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Integration of the accelerator Aha1 in the Hsp90 co-Chaperone cycle
Nature Structural & Molecular Biology, 2013Co-Authors: Jing Li, Klaus Richter, Jochen Reinstein, Johannes BuchnerAbstract:The role of co-Chaperone and Hsp90 activator Aha1 is now examined in conjunction with other co-Chaperones in vivo and in vitro , to reveal how they regulate the progression of the Hsp90 cycle. Aha1 and Cpr6 interact with and activate Hsp90 in a synergistic manner and displace the inhibitory co-Chaperone Sti1. Aha1 is eventually released from Hsp90 by p23. Heat-shock protein 90 (Hsp90) is an ATP-dependent molecular Chaperone that associates dynamically with various co-Chaperones during its Chaperone cycle. Here we analyzed the role of the activating co-Chaperone Aha1 in the progression of the yeast Hsp90 Chaperone cycle and identified a critical ternary Hsp90 complex containing the co-Chaperones Aha1 and Cpr6. Aha1 accelerates the intrinsically slow conformational transitions of Hsp90 to an N-terminally associated state but does not fully close the nucleotide-binding pocket yet. Cpr6 increases the affinity between Aha1 and Hsp90 and further stimulates the Hsp90 ATPase activity. Synergistically, Aha1 and Cpr6 displace the inhibitory co-Chaperone Sti1 from Hsp90. To complete the cycle, Aha1 is released by the co-Chaperone p23. Thus, at distinct steps during the Hsp90 Chaperone cycle, co-Chaperones selectively trap statistically distributed Hsp90 conformers and thus turn Hsp90 into a deterministic machine.
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Chaperones and protein folding
2012Co-Authors: Arthur L. Horwich, Johannes Buchner, Robert G. Smock, Lila M. Gierasch, H.r. SaibilAbstract:The folding and translocation of many newly synthesized proteins in the cell is kinetically assisted by ubiquitous, abundant, specialized proteins known as molecular Chaperones. These components generally recognize hydrophobic surfaces, exposed specifically by non-native conformations, through their own solvent-exposed hydrophobic surfaces, with different classes of Chaperone recognizing such surfaces in the context of extended (Hsp70) vs. collapsed (Hsp60/chaperonin) topology of substrate protein. Such binding prevents substrate proteins from misfolding and from forming multimolecular aggregates. Chaperone-bound proteins are then released from Hsp70 and Hsp60 machines via the binding of ATP to Chaperone domains physically separated from the substrate protein binding domains, via allosterically directed conformational changes. Molecular Chaperones also act under stress conditions, where polypeptide chains are subject to misfolding, preventing aggregation and restoring the native state. The small heat shock proteins (sHsps) are oligomeric assemblies that participate with the other Chaperones in binding non-native states under such conditions. Finally, Hsp90 is an abundant clamp-shaped Chaperone that participates in binding and maturation of a variety of substrate proteins via an ATP-directed cycle. This chapter reviews the structure and mechanism of action of these Chaperones, with special attention directed to the variety of biophysical methods employed to reaching our current understanding.
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3.10 Chaperones and Protein Folding
Comprehensive Biophysics, 2012Co-Authors: Arthur L. Horwich, Johannes Buchner, Robert G. Smock, Lila M. Gierasch, H.r. SaibilAbstract:The folding and translocation of many newly synthesized proteins in the cell is kinetically assisted by ubiquitous, abundant, specialized proteins known as molecular Chaperones. These components generally recognize hydrophobic surfaces, exposed specifically by non-native conformations, through their own solvent-exposed hydrophobic surfaces, with different classes of Chaperone recognizing such surfaces in the context of extended (Hsp70) vs. collapsed (Hsp60/chaperonin) topology of substrate protein. Such binding prevents substrate proteins from misfolding and from forming multimolecular aggregates. Chaperone-bound proteins are then released from Hsp70 and Hsp60 machines via the binding of ATP to Chaperone domains physically separated from the substrate protein binding domains, via allosterically directed conformational changes. Molecular Chaperones also act under stress conditions, where polypeptide chains are subject to misfolding, preventing aggregation and restoring the native state. The small heat shock proteins (sHsps) are oligomeric assemblies that participate with the other Chaperones in binding non-native states under such conditions. Finally, Hsp90 is an abundant clamp-shaped Chaperone that participates in binding and maturation of a variety of substrate proteins via an ATP-directed cycle. This chapter reviews the structure and mechanism of action of these Chaperones, with special attention directed to the variety of biophysical methods employed to reaching our current understanding.
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The Hsp90 Chaperone machinery: Conformational dynamics and regulation by co-Chaperones
Biochimica et biophysica acta, 2011Co-Authors: Joanna Soroka, Johannes BuchnerAbstract:Hsp90 is a dimeric molecular Chaperone required for the activation and stabilization of numerous client proteins many of which are involved in essential cellular processes like signal transduction pathways. This activation process is regulated by ATP-induced large conformational changes, co-Chaperones and posttranslational modifications. For some co-Chaperones, a detailed picture on their structures and functions exists, for others their contributions to the Hsp90 system is still unclear. Recent progress on the conformational dynamics of Hsp90 and how co-Chaperones affect the Hsp90 Chaperone cycle significantly increased our understanding of the gearings of this complex molecular machinery. This article is part of a Special Issue entitled: Heat Shock Protein 90 (Hsp90).
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HOW ChaperoneS FOLD PROTEINS
Biological chemistry, 1998Co-Authors: M Beissinger, Johannes BuchnerAbstract:Chaperones are a functionally related group of proteins assisting protein folding in the cell under physiological and stress conditions. They share the ability to recognize and bind nonnative proteins thus preventing unspecific aggregation. The underlying functional principles of the different Chaperone classes are beginning to be understood. A landmark feature of molecular Chaperones is the involvement of energy-dependent reactions in the folding process. Nucleotide binding to ATP-dependent Chaperones (e.g. GroEL, Hsp70, Hsp90) leads to sometimes large conformational changes in the Chaperone which allow to shift between high- and low-affinity states for substrate proteins. Interestingly, the ATPase activity which is the key determinant for functional cycles is tightly regulated by a set of co-Chaperones. While for ATP-dependent Chaperones binding sites for nucleotide and protein are found in one protein, in the case of ATP-independent Chaperones (e. g. sHsps, SecB) the energy-dependent step is performed by another Chaperone (Hsp70, SecA). Therefore, the ATP-independent Chaperones can be regarded as efficient 'holding' components. Cooperation of different Chaperone machineries creates a synergistic network of folding helpers in the cell, which allows to maintain protein homeostasis under conditions nonpermissive for spontaneous folding.
Gabriela Chiosis - One of the best experts on this subject based on the ideXlab platform.
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Chaperome Networks - Redundancy and Implications for Cancer Treatment.
Advances in experimental medicine and biology, 2020Co-Authors: Pengrong Yan, Tai Wang, Monica L. Guzman, Radu I. Peter, Gabriela ChiosisAbstract:The chaperome is a large family of proteins composed of Chaperones, co-Chaperones and a multitude of other factors. Elegant studies in yeast and other organisms have paved the road to how we currently understand the complex organization of this large family into protein networks. The goal of this chapter is to provide an overview of chaperome networks in cancer cells, with a focus on two cellular states defined by chaperome network organization. One state characterized by chaperome networks working in isolation and with little overlap, contains global chaperome networks resembling those of normal, non-transformed, cells. We propose that in this state, redundancy in chaperome networks results in a tumor type unamenable for single-agent chaperome therapy. The second state comprises chaperome networks interconnected in response to cellular stress, such as MYC hyperactivation. This is a state where no redundant pathways can be deployed, and is a state of vulnerability, amenable for chaperome therapy. We conclude by proposing a change in how we discover and implement chaperome inhibitor strategies, and suggest an approach to chaperome therapy where the properties of chaperome networks, rather than genetics or client proteins, are used in chaperome inhibitor implementation.
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Adapting to stress — chaperome networks in cancer
Nature Reviews Cancer, 2018Co-Authors: Suhasini Joshi, Tai Wang, Thaís L. S. Araujo, Sahil Sharma, Jeffrey L. Brodsky, Gabriela ChiosisAbstract:In this Opinion, Joshi et al. argue that in cancer cells, a state of chaperome hyperconnectivity is obtained by increasing the interaction strength among chaperome machinery members. These chaperome scaffolding platforms act to increase the functional diversity of oncogenic processes and have implications for the development of chaperome inhibitors. In this Opinion article, we aim to address how cells adapt to stress and the repercussions chronic stress has on cellular function. We consider acute and chronic stress-induced changes at the cellular level, with a focus on a regulator of cellular stress, the chaperome, which is a protein assembly that encompasses molecular Chaperones, co-Chaperones and other co-factors. We discuss how the chaperome takes on distinct functions under conditions of stress that are executed in ways that differ from the one-on-one cyclic, dynamic functions exhibited by distinct molecular Chaperones. We argue that through the formation of multimeric stable chaperome complexes, a state of chaperome hyperconnectivity, or networking, is gained. The role of these chaperome networks is to act as multimolecular scaffolds, a particularly important function in cancer, where they increase the efficacy and functional diversity of several cellular processes. We predict that these concepts will change how we develop and implement drugs targeting the chaperome to treat cancer.