Learn more: Chaperones and Proteostasis

Endoplasmic reticulum Ca2+ increases enhance mutant glucocerebrosidase proteostasis
Nat.Chem.Biol. 2010, view full abstract in PubMed

Altering intracellular calcium levels is known to partially restore mutant enzyme homeostasis in several lysosomal storage diseases, but why? We hypothesized that endoplasmic reticulum (ER) calcium increases enhance the folding, trafficking and function of these mutant misfolding- and degradation-prone lysosomal enzymes by increasing chaperone function. Here we report that increasing ER calcium levels by reducing ER calcium efflux through the ryanodine receptor, using antagonists or RNAi, or by promoting ER calcium influx by SERCA2b overexpression enhances mutant glucocerebrosidase (GC) homeostasis in cells derived from individuals with Gaucher's disease. Post-translational regulation of the calnexin folding pathway by an elevated ER calcium concentration seems to enhance the capacity of this chaperone system to fold mutant misfolding-prone enzymes, increasing the folded mutant GC population that can engage the trafficking receptor at the expense of ER-associated degradation, increasing the lysosomal GC concentration and activity.

Chaperone-assisted degradation: multiple paths to destruction
Biol.Chem. 2010, view full abstract in PubMed

Molecular chaperones are well known as facilitators of protein folding and assembly. However, in recent years multiple chaperone-assisted degradation pathways have also emerged, including CAP (chaperone-assisted proteasomal degradation), CASA (chaperone-assisted selective autophagy), and CMA (chaperone-mediated autophagy). Within these pathways chaperones facilitate the sorting of non-native proteins to the proteasome and the lysosomal compartment for disposal. Impairment of these pathways contributes to the development of cancer, myopathies, and neurodegenerative diseases. Chaperone-assisted degradation thus represents an essential aspect of cellular proteostasis, and its pharmacological modulation holds the promise to ameliorate some of the most devastating diseases of our time. Here, we discuss recent insights into molecular mechanisms underlying chaperone-assisted degradation in mammalian cells and highlight its biomedical relevance.

HSPA1A is upregulated in periodontal ligament at early stage of tooth movement in rats
Histochem.Cell Biol. 2010, view full abstract in PubMed

Heat shock proteins (HSPs) are molecular chaperones that maintain intracellular protein homeostasis and ensure survival of cells. Continuous orthodontic force on the tooth is considered to be a type of physical stress loaded to the periodontal ligament (PDL). However, little is known about the role of HSPs during tooth movement. This study was performed to examine the expression of HSPs in the PDL during tooth movement using laser microdissection, microarray analysis, real-time RT-PCR and immunohistochemistry. Gene expression of HSPA1A in the pressure zone of the PDL was higher during 6 h of tooth movement than in the control group. Expression of HSPA1A decreased with time. HSPA1A was also detected in the pressure zone of the PDL at the protein level 24 h after the initial tissue change. These results strongly suggest that expression of HSPA1A in the PDL during early stages of tooth movement is a critical factor for tissue reaction.

Binding of a small molecule at a protein-protein interface regulates the chaperone activity of hsp70-hsp40
ACS Chem.Biol. 2010, view full abstract in PubMed

Heat shock protein 70 (Hsp70) is a highly conserved molecular chaperone that plays multiple roles in protein homeostasis. In these various tasks, the activity of Hsp70 is shaped by interactions with co-chaperones, such as Hsp40. The Hsp40 family of co-chaperones binds to Hsp70 through a conserved J-domain, and these factors stimulate ATPase and protein-folding activity. Using chemical screens, we identified a compound, 115-7c, which acts as an artificial co-chaperone for Hsp70. Specifically, the activities of 115-7c mirrored those of a Hsp40; the compound stimulated the ATPase and protein-folding activities of a prokaryotic Hsp70 (DnaK) and partially compensated for a Hsp40 loss-of-function mutation in yeast. Consistent with these observations, NMR and mutagenesis studies indicate that the binding site for 115-7c is adjacent to a region on DnaK that is required for J-domain-mediated stimulation. Interestingly, we found that 115-7c and the Hsp40 do not compete for binding but act in concert. Using this information, we introduced additional steric bulk to 115-7c and converted it into an inhibitor. Thus, these chemical probes either promote or inhibit chaperone functions by regulating Hsp70-Hsp40 complex assembly at a native protein-protein interface. This unexpected mechanism may provide new avenues for exploring how chaperones and co-chaperones cooperate to shape protein homeostasis.

ATP-independent reversal of a membrane protein aggregate by a chloroplast SRP subunit
Nat.Struct.Mol.Biol. 2010, view full abstract in PubMed

Membrane proteins impose enormous challenges to cellular protein homeostasis during their post-translational targeting, and they require chaperones to keep them soluble and translocation competent. Here we show that a novel targeting factor in the chloroplast signal recognition particle (cpSRP), cpSRP43, is a highly specific molecular chaperone that efficiently reverses the aggregation of its substrate proteins. In contrast to 'ATPases associated with various cellular activities' (AAA(+)) chaperones, cpSRP43 uses specific binding interactions with its substrate to mediate its 'disaggregase' activity. This disaggregase capability can allow targeting machineries to more effectively capture their protein substrates and emphasizes a close connection between protein folding and trafficking processes. Moreover, cpSRP43 provides the first example to our knowledge of an ATP-independent disaggregase and shows that efficient reversal of protein aggregation can be attained by specific binding interactions between a chaperone and its substrate.

Chaperone networks: tipping the balance in protein folding diseases
Neurobiol.Dis. 2010, view full abstract in PubMed

Adult-onset neurodegeneration and other protein conformational diseases are associated with the appearance, persistence, and accumulation of misfolded and aggregation-prone proteins. To protect the proteome from long-term damage, the cell expresses a highly integrated protein homeostasis (proteostasis) machinery to ensure that proteins are properly expressed, folded, and cleared, and to recognize damaged proteins. Molecular chaperones have a central role in proteostasis as they have been shown to be essential to prevent the accumulation of alternate folded proteotoxic states as occurs in protein conformation diseases exemplified by neurodegeneration. Studies using invertebrate models expressing proteins associated with Huntington's disease, Alzheimer's disease, ALS, and Parkinson's disease have provided insights into the genetic networks and stress signaling pathways that regulate the proteostasis machinery to prevent cellular dysfunction, tissue pathology, and organismal failure. These events appear to be further amplified by aging and provide evidence that age-related failures in proteostasis may be a common element in many diseases.

Chaperones and multitasking proteins in the nucleolus: networking together for survival?
Trends Biochem.Sci. 2010, view full abstract in PubMed

The nucleolus has emerged as a key player that regulates cell growth, survival and the recovery from stress. Progress in proteomics made it possible to sequence the nucleolar proteome under different physiological conditions. Together with other research, this work revealed the presence of multiple chaperones and co-chaperones in the nucleolus. Molecular chaperones are components of a larger network that promotes protein homeostasis, thereby providing continuous adaptation to a changing environment. Recent studies suggest that the cellular chaperone network is divided into individual branches which orchestrate specific functions. Input from separate branches is then combined to 'fine-tune' the cellular proteostasis network. Based on the latest developments in nucleolar and chaperone biology, we speculate that a unique network comprising chaperones, co-chaperones and multitasking proteins is located in nucleoli. This network supports and regulates fundamental biological processes, including ribosome biogenesis, cell signaling, and the stress response.

Identification and molecular analysis of a stress-inducible Hsp70 from Sciaenops ocellatus
Fish.Shellfish.Immunol. 2010, view full abstract in PubMed

Hsp70 proteins are a family of molecular chaperones that are involved in many aspects of protein homeostasis. In this study, an Hsp70 homologue (SoHsp70) was identified from red drum Sciaenops ocellatus and analyzed at molecular level. The open reading frame of SoHsp70 is 1920 bp and intronless, with a 5'-untranslated region (UTR) of 399 bp and a 3'-UTR of 241 bp. The deduced amino acid sequence of SoHsp70 shares 84-92% overall identities with the Hsp70s of a number of fish species. In silico analysis identified in SoHsp70 three conserved Hsp70 domains involved in nucleotide and substrate binding. The coding sequence of SoHsp70 was subcloned into Escherichia coli, from which recombinant SoHsp70 was purified and, upon ATPase assay, found to exhibit apparent ATPase activity. Expressional analysis showed that constitutive expression of SoHsp70 was detectable in heart, liver, spleen, kidney, brain, blood, and gill. Experimental challenges with poly(I:C) and bacterial pathogens of Gram-positive and Gram-negative nature induced SoHsp70 expression in kidney to different levels. Stress-responsive analysis of SoHsp70 expression in primary cultures of red drum hepatocytes showed that acute heat shock treatment elicited a rapid induction of SoHsp70 expression which appeared after 10 min and 30 min of treatment. Exposure of hepatocytes separately to iron, copper, mercury, and hydrogen peroxide significantly upregulated SoHsp70 expression in time-dependent manners. Vaccination of red drum with a Streptococcus iniae bacterin was also found to induce SoHsp70 expression. Furthermore, recombinant SoHsp70 enhanced the immunoprotective effect of a subunit vaccine. Taken together, these results suggest that SoHsp70 is a stress-inducible protein that is likely to play a role in immunity and in coping with environmental and biological stresses.

Physicochemical determinants of chaperone requirements
J.Mol.Biol. 2010, view full abstract in PubMed

We describe a series of stringent relationships between abundance, solubility and chaperone usage of proteins. Based on these relationships, we show that the need of Escherichia coli proteins for the chaperonin GroEL can be predicted with 86% accuracy. Furthermore, from the observation that the abundance and solubility of proteins depend on the physicochemical properties of their amino acid sequences, we demonstrate that the requirement for GroEL can also be predicted directly from the sequences with 90% accuracy. These results indicate that the physicochemical properties of the amino acid sequences represent an essential component of the cellular quality control system that ensures the maintenance of protein homeostasis in living systems.

Structure and function of archaeal prefoldin, a co-chaperone of group II chaperonin
Front Biosci. 2010, view full abstract in PubMed

Molecular chaperones are key cellular components involved in the maintenance of protein homeostasis and other unrelated functions. Prefoldin is a chaperone that acts as a co-factor of group II chaperonins in eukaryotes and archaea. It assists proper folding of protein by capturing nonnative proteins and delivering it to the group II chaperonin. Eukaryotic prefoldin is a multiple subunit complex composed of six different polypeptide chains. Archaeal prefoldin, on the other hand, is a heterohexameric complex composed of two alpha and four beta subunits, and forms a double beta barrel assembly with six long coiled coils protruding from it like a jellyfish with six tentacles. Based on the structural information of the archaeal prefoldin, substrate recognition and prefoldin-chaperonin binding mechanisms have been investigated. In this paper, we review a series of studies on the molecular mechanisms of archaeal PFD function. Particular emphasis will be placed on the molecular structures revealed by X-ray crystallography and molecular dynamics induced by binding to nonnative protein substrates.