Molecular Chaperone Functions in Protein Folding and Proteostasis

The major classes of molecular chaperones have highly variable sequences, sizes, and shapes, yet they all bind to unfolded proteins, limit their aggregation, and assist in their folding. Despite the central importance of this process to protein homeostasis, it has not been clear exactly how chaperones guide this process or whether the diverse families of chaperones use similar mechanisms. For the first time, recent advances in NMR spectroscopy have enabled detailed studies of how unfolded, “client” proteins interact with both ATP-dependent and ATP-independent classes of chaperones. Here, we review examples from four distinct chaperones, Spy, Trigger Factor, DnaK, and HscA-HscB, highlighting the similarities and differences between their mechanisms. One striking similarity is that the chaperones all bind weakly to their clients, such that the chaperone–client interactions are readily outcompeted by stronger, intra- and intermolecular contacts in the folded state. Thus, the relatively weak affinity of these interactions seems to provide directionality to the folding process. However, there are also key differences, especially in the details of how the chaperones release clients and how ATP cycling impacts that process. For example, Spy releases clients in a largely folded state, while clients seem to be unfolded upon release from Trigger Factor or DnaK. Together, these studies are beginning to uncover the similarities and differences in how chaperones use weak interactions to guide protein folding.

Misfolded and aggregated proteins build up in neurodegenerative illnesses, which causes neuronal dysfunction and ultimately neuronal death. In the last few years, there has been a significant upsurge in the level of interest towards the function of molecular chaperones in the control of misfolding and aggregation. The crucial molecular chaperones implicated in neurodegenerative illnesses are covered in this review article, along with a variety of their different methods of action. By aiding in protein folding, avoiding misfolding, and enabling protein breakdown, molecular chaperones serve critical roles in preserving protein homeostasis. By aiding in protein folding, avoiding misfolding, and enabling protein breakdown, molecular chaperones have integral roles in preserving regulation of protein balance. It has been demonstrated that aging, a significant risk factor for neurological disorders, affects how molecular chaperones function. The aggregation of misfolded proteins and the development of neurodegeneration may be facilitated by the aging-related reduction in chaperone activity. Molecular chaperones have also been linked to the pathophysiology of several instances of neuron withering illnesses, enumerating as Parkinson's disease, Huntington's disease, and Alzheimer's disease. Molecular chaperones have become potential therapy targets concerning with the prevention and therapeutic approach for brain disorders due to their crucial function in protein homeostasis and their connection to neurodegenerative illnesses. Protein homeostasis can be restored, and illness progression can be slowed down by methods that increase chaperone function or modify their expression. This review emphasizes the importance of molecular chaperones in the context of neuron withering disorders and their potential as therapeutic targets for brain disorders.

The price paid by the cardiovascular system for maintaining circulation is being a target of chronic forces that contribute to wear and tear. Cardiovascular tissues are in constant peril of damage from these stresses, and are dependent on highly conserved cellular machines for protection. The cellular environment is adapted to detect, repair and, if necessary, dispose of damaged proteins, in part because they are toxic to the cell. The molecular chaperone and ubiquitin-proteasome systems are the machinery that repair and degrade damaged proteins, yet little attention has been given to these systems in cardiovascular pathophysiology. Recent advances have brought protein folding and degradation closer to the forefront of cardiovascular biology. This review discusses these advances and presents new models for consideration of cardiovascular pathophysiology and therapeutics as problems of protein homeostasis. In most cases, all the information necessary for a protein to fold is contained in its primary amino acid sequence. The environment within a cell is particularly unfavorable for protein folding, however, because of constraints that occur during protein synthesis and the likelihood of intermolecular interactions that impede folding in a crowded cellular environment. An evolutionarily ancient system exists to prevent damaged or newly synthesized peptides from aggregating and to provide a microenvironment that facilitates their proper folding to a thermodynamically favorable active conformation. This system comprises the molecular chaperones, which are present in every cellular compartment to buffer and repair damaged proteins (the Table provides a partial list of mammalian chaperones). Mechanisms for removing proteins are equally critical to cellular function, and the machinery of protein degradation is similarly archaic. Although several pathways exist for protein destruction in eukaryotes, the ubiquitin-proteasome system is responsible for the majority of protein degradation and is the most tightly regulated pathway. View this table: Examples of Mammalian Chaperones and Their Functions The chaperone and ubiquitin-proteasome …

Heat shock protein 70 (Hsp70) molecular chaperones play critical roles in protein homeostasis. In the budding yeast Saccharomyces cerevisiae, cytosolic Hsp70 interacts with up to three types of nucleotide exchange factors (NEFs) homologous to human counterparts: Sse1/Sse2 (Heat shock protein 110 (Hsp110)), Fes1 (HspBP1), and Snl1 (Bag-1). All three NEFs stimulate ADP release; however, it is unclear why multiple distinct families have been maintained throughout eukaryotic evolution. In this study we investigate NEF roles in Hsp70 cell biology using an isogenic combinatorial collection of NEF deletion mutants. Utilizing well characterized model substrates, we find that Sse1 participates in most Hsp70-mediated processes and is of particular importance in protein biogenesis and degradation, whereas Fes1 contributes to a minimal extent. Surprisingly, disaggregation and resolubilization of thermally denatured firefly luciferase occurred independently of NEF activity. Simultaneous deletion of SSE1 and FES1 resulted in constitutive activation of heat shock protein expression mediated by the transcription factor Hsf1, suggesting that these two factors are important for modulating stress response. Fes1 was found to interact in vivo preferentially with the Ssa family of cytosolic Hsp70 and not the co-translational Ssb homolog, consistent with the lack of cold sensitivity and protein biogenesis phenotypes for fes1Δ cells. No significant consequence could be attributed to deletion of the minor Hsp110 SSE2 or the Bag homolog SNL1. Together, these lines of investigation provide a comparative analysis of NEF function in yeast that implies Hsp110 is the principal NEF for cytosolic Hsp70, making it an ideal candidate for therapeutic intervention in human protein folding disorders.

Molecular chaperones play critical roles in biological functions. They are closely involved in the maintenance of cell homeostasis, proper folding of proteins and nucleic acids, and inhibition of irreversible aggregation in denatured proteins. In addition to protein production, molecular chaperone function is widely recognized as important for peptide and protein drug delivery systems. Therefore, much effort has been made in recent decades to develop chaperone-mimetic molecules that have similar structures and biological functions to natural chaperones. These artificial molecular chaperone systems have been demonstrated to facilitate proper protein and nucleic acid folding, in addition to the formation of higher-order structures of synthetic molecules. Furthermore, the functions of these artificial systems show promising clinical applications in drug delivery and biomolecule detection. This topical review focuses on recent advances in the design, construction, characterization, and potential applications of different artificial molecular systems with distinct functional roles, such as the folding of water-soluble and membrane proteins, nucleic acids, and the self-assembly of synthetic molecules. Strategies used in the construction of some artificial molecule chaperone systems for proteins (such as pairs of amphiphilic molecules or self-assembled nanogels) and their applications as biomaterials are described. Specific examples from each design strategy are also highlighted to demonstrate the mechanisms, challenges, and limitations of the different artificial molecular systems. By highlighting the many new developments that have expanded the applications of the artificial chaperones beyond protein folding, this review aims to stimulate further studies on their design and applications.

Parkinson's disease, like many other neurodegenerative disorders, is characterized by the progressive accumulation of pathogenic protein species and the formation of intracellular inclusion bodies. The cascade by which the small synaptic protein α-synuclein misfolds to form distinctive protein aggregates, termed Lewy bodies and Lewy neurites, has been the subject of intensive research for more than a decade. Genetic and pathological studies in Parkinson's disease patients as well as experimental studies in disease models have clearly established altered protein metabolism as a key element in the pathogenesis of Parkinson's disease. Alterations in protein metabolism include misfolding and aggregation, post-translational modification and dysfunctional degradation of cytotoxic protein species. Protein folding and re-folding are both mediated by a highly conserved network of molecules, called molecular chaperones and co-chaperones. In addition to the regulatory role in protein folding, molecular chaperone function is intimately associated with pathways of protein degradation, such as the ubiquitin-proteasome system and the autophagy-lysosomal pathway, to effectively remove irreversibly misfolded proteins. Because of the central role of molecular chaperones in maintaining protein homeostasis, we herein review our current knowledge on the involvement of molecular chaperones and co-chaperones in Parkinson's disease. We further discuss the capacity of molecular chaperones to prevent or modulate neurodegeneration, an important concept for future neuroprotective strategies and summarize the current progress in preclinical studies in models of Parkinson's disease and other neurodegenerative disorders. Finally we include a discussion on the future potential of using molecular chaperones as a disease modifying therapy.

Partners in surveillance and quality control

Cellular protein homeostasis is tightly regulated by balancing protein synthesis, folding, and degradation. Cancer cells constantly modulate protein folding and degradation efficiency to survive and proliferate without accumulating toxic misfolded proteins under acute stress conditions, such as low oxygen and energy, and exposure to chemotherapeutic agents. Many cancers rely on the unfolded protein response (UPR) to cope with stress, which increases the expression of molecular chaperones and mitigates the accumulation of misfolded proteins. Thus, it is unsurprising that the Hsp70 molecular chaperone, which serves as a master regulator of proteostasis, is upregulated in many cancers. Increased Hsp70 correlates with metastasis formation and poor patient prognosis in different cancers, like breast cancer. To define the role of Hsp70 in cancer survival and identify how cancer cells compensate for chaperone-mediated proteotoxicity, we investigated the response to a specific Hsp70 inhibitor, MAL3-101, in breast cancer cells. We discovered that breast cancer cells bin into distinct groups when subjected to Hsp70 inhibition based on their sensitivity to the compound. Moreover, we demonstrate that resistant cells have higher autophagy levels compared to more sensitive lines. Autophagy was further induced by MAL3-101 in resistant breast cancer cells, as evidenced by the accumulation of both autophagy related genes and proteins, as well as by autophagic-like structures detected by confocal microscopy. These data suggest that resistance to Hsp70 inhibition arises from autophagy induction. We then discovered that Hsp70 inhibition induces the UPR pathway, triggering apoptosis in sensitive cells. In particular, the activation of the UPR transducer, PERK, is required to induce apoptosis in sensitive cells when Hsp70 was inhibited. Overall, our work positions autophagy as a critical compensatory mechanism when molecular chaperone function is overwhelmed and misfolded proteins accumulate. Furthermore, our findings delineate a distinct role of PERK in apoptosis induction upon Hsp70 inhibition, highlighting the interplay between the PERK, Hsp70, and autophagy. Citation Format: Sara Sannino, Jeffrey L. Brodsky. Modulation of protein homeostasis networks upon Hsp70 inhibition in cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1934.

Molecular chaperones and neuronal proteostasis

Geldanamycin (GA) bind heat-shock protein-90 (HSP-90) and destabilize its client proteins including v-Src, Bcr-Abl, RAF-1, Erb-B2, some growth factor receptors and steroid receptors. As a result, several oncoproteins are subjected to ubiquitination and proteasomal destruction by HSP-90 active compounds. HSP-90 active substances can either stop apoptosis from occurring or promote growth arrest, differentiation, and apoptosis depending on the cellular environment. Numerous preclinical models and clinical trials have demonstrated anticancer activity for a number of HSP-90 inhibitors. The well-known HSP-90 inhibitor geldanamycin’s clinical development was hampered by its hepatic toxicity. Geldanamycin at low doses can sensitize Bcr/Abl-expressing leukemia cells to death in the presence of inadequate doxorubicin concentrations by activating caspase. In another example, 17AAG in combination with taxol shows enhanced cytotoxic effects on taxol-resistant Erb-B2 overexpressing breast cancer cells. The benzoquinone ansamycin geldanamycin selectively binds to GRP94 and HSP-90 both in vivo and in vitro. When cells are treated with geldanamycin, HSP-90’s molecular chaperone function is changed. This prevents some cytosolic proteins from maturing, reduces their activity, and/or modifies their stability. On the other hand, nothing is known about GRP94’s function in protein folding or how geldanamycin affects this endoplasmic reticulum (ER) homologue of HSP-90. In this work, we show that geldanamycin is a strong inducer of the cellular stress response in the ER, leading to the transcriptional up-regulation of ER chaperones and production of the gadd153/CHOP transcription factor in a range of cell lines. Here we mention the anticancerous activity of HSP-90 (Heat Shock Protein 90) Inhibitor geldanamycin and some researches in field of anticancerous activity of Geldanamycin.

] In fact, recent studies have demonstrated experimentally that increasing the burden of misfolded proteins in the heart can contribute to the development of cardiac dysfunction. In this review, we discuss the role of heat shock proteins (HSPs) in common cardiac diseases, including cardiac hypertrophy, heart failure, and ischemia/reperfusion injury. Furthermore, we delineate the many specific mechanisms by which these chaperones, cochaperones, and heat shock factor (HSF) transcription factors have been found to be cardioprotective in experimental models. Lastly, we review recent studies involving drugs that are being developed (and currently used) to increase the expression (and presumably function) of chaperone/cochaperone systems that may be applicable to the treatment of common cardiac diseases and familial cardiac diseases with a pathogenesis that includes a major component of misfolded proteins (eg, desminopathies).

Review: Mechanisms of Disaggregation and Refolding of Stable Protein Aggregates by Molecular Chaperones

The Hsp70 and Hsp90 molecular chaperone systems are critical regulators of protein homeostasis (proteostasis) in eukaryotes under normal and stressed conditions. The Hsp70 and Hsp90 systems physically and functionally interact to ensure cellular proteostasis. Co-chaperones interact with Hsp70 and Hsp90 to regulate and to promote their molecular chaperone functions. Mammalian Hop, also called Stip1, and its budding yeast ortholog Sti1 are eukaryote-specific co-chaperones, which have been thought to be essential for substrate (“client”) transfer from Hsp70 to Hsp90. Substrate transfer is facilitated by the ability of Hop to interact simultaneously with Hsp70 and Hsp90 as part of a ternary complex. Intriguingly, in prokaryotes, which lack a Hop ortholog, the Hsp70 and Hsp90 orthologs interact directly. Recent evidence shows that eukaryotic Hsp70 and Hsp90 can also form a prokaryote-like binary chaperone complex in the absence of Hop, and that this binary complex displays enhanced protein folding and anti-aggregation activities. The canonical Hsp70-Hop-Hsp90 ternary chaperone complex is essential for optimal maturation and stability of a small subset of clients, including the glucocorticoid receptor, the tyrosine kinase v-Src, and the 26S/30S proteasome. Whereas many cancers have increased levels of Hop, the levels of Hop decrease in the aging human brain. Since Hop is not essential in all eukaryotic cells and organisms, tuning Hop levels or activity might be beneficial for the treatment of cancer and neurodegeneration.

the endoplasmic reticulum (ER) serves as a way station during the biogenesis of nearly every integral membrane and secreted protein synthesized in eukaryotic cells. As such, its primary role is to facilitate the folding of nascent polypeptides. Thus plentiful among the ER resident proteins are 1 )

Protein homeostasis networks in physiology and disease