Towards a mechanism for histone chaperones

Histone chaperones can be broadly defined as histone-binding proteins that influence chromatin dynamics in an ATP-independent manner. Their existence reflects the importance of chromatin homeostasis and the unique and unusual biochemistry of the histone proteins. Histone supply and demand at chromatin is regulated by a network of structurally and functionally diverse histone chaperones. At the core of this network is a mechanistic variability that is only beginning to be appreciated. In this review, we highlight the challenges in determining histone chaperone mechanism and discuss possible mechanisms in the context of nucleosome thermodynamics. We discuss how histone chaperones prevent promiscuous histone interactions, and consider if this activity represents the full extent of histone chaperone function in governing chromatin dynamics. This article is part of a Special Issue entitled: Histone chaperones and Chromatin assembly.     

Histone chaperones link histone nuclear import and chromatin assembly

Histone chaperones are proteins that shield histones from nonspecific interactions until they are assembled into chromatin. After their synthesis in the cytoplasm, histones are bound by different histone chaperones, subjected to a series of posttranslational modifications and imported into the nucleus. These evolutionarily conserved modifications, including acetylation and methylation, can occur in the cytoplasm, but their role in regulating import is not well understood. As part of histone import complexes, histone chaperones may serve to protect the histones during transport, or they may be using histones to promote their own nuclear localization. In addition, there is evidence that histone chaperones can play an active role in the import of histones. Histone chaperones have also been shown to regulate the localization of important chromatin modifying enzymes. This review is focused on the role histone chaperones play in the early biogenesis of histones, the distinct cytoplasmic subcomplexes in which histone chaperones have been found in both yeast and mammalian cells and the importins/karyopherins and nuclear localization signals that mediate the nuclear import of histones. We also address the role that histone chaperone localization plays in human disease. This article is part of a Special Issue entitled: Histone chaperones and chromatin assembly.     

The chaperone-histone partnership: for the greater good of histone traffic and chromatin plasticity

Histones are highly positively charged proteins that wrap our genome. Their surface properties also make them prone to nonspecific interactions and aggregation. A class of proteins known as histone chaperones is dedicated to safeguard histones by aiding their proper incorporation into nucleosomes. Histone chaperones facilitate ordered nucleosome assembly and disassembly reactions through the formation of semi-stable histone-chaperone intermediates without requiring ATP, but merely providing a complementary protein surface for histones to dynamically interact with. Recurrent 'chaperoning' mechanisms involve the masking of the histone's positive charge and the direct blocking of crucial histone surface sites, including those required for H3-H4 tetramerization or the binding of nucleosomal DNA. This shielding prevents histones from engaging in premature or unwanted interactions with nucleic acids and other cellular components. In this review, we analyze recent structural studies on chaperone-histone interactions and discuss the implications of this vital partnership for nucleosome assembly and disassembly pathways.     

Endoplasmic reticulum chaperones stabilize nicotinic receptor subunits and regulate receptor assembly

We examined interactions between the endoplasmic reticulum (ER) chaperones calnexin (CN), ERp57, and immunological heavy chain-binding protein (BiP) and nicotinic acetylcholine receptor (nAChR) subunits. The three chaperones rapidly associate with newly synthesized nAChR subunits. Interactions between nAChR subunits and ERp57 occur via transient intermolecular disulfide bonds and do not require subunit N-linked glycosylation. The associations of ERp57 or CN with AChR subunits are long lived and prolong subunit lifetime approximately 10-fold. Coexpression of CN or ERp57 alone does not affect nAChR assembly or trafficking, but together they cause a significant decrease in nAChR expression and assembly. In contrast, associations with BiP are shorter lived and do not alter nAChR expression and assembly. However, a mutated BiP that slows its dissociation significantly increases its associations and decreases nAChR expression and assembly. Our results suggest that interactions with the chaperones regulate the levels of nAChRs assembled in the ER by stabilizing and sequestering subunits during assembly.     

Molecular chaperones: assisting assembly in addition to folding

The common perception that molecular chaperones are involved primarily with assisting the folding of newly synthesized and stress-denatured polypeptide chains ignores the fact that this term was invented to describe the function of a protein that assists the assembly of folded subunits into oligomeric structures and only later was extended to embrace protein folding. Recent work has clarified the role of nuclear chaperones in the assembly of nucleosomes and has identified a cytosolic chaperone required for mammalian proteasome assembly, suggesting that the formation of other oligomeric complexes might be assisted by chaperones.     

Procollagen folding and assembly: the role of endoplasmic reticulum enzymes and molecular chaperones

Procollagen assembly occurs within the endoplasmic reticulum, where the C-propeptide domains of three polypeptide alpha-chains fold individually, and then interact and trimerise to initiate folding of the triple helical region. This highly complex folding and assembly pathway requires the co-ordinated action of a large number of endoplasmic reticulum-resident enzymes and molecular chaperones. Disease-causing mutations in the procollagens disturb folding and assembly and lead to prolonged interactions with molecular chaperones, retention in the endoplasmic reticulum, and intracellular degradation. This review focuses predominantly on prolyl 1-hydroxylase, an essential collagen modifying enzyme, and HSP47, a collagen-specific binding protein, and their proposed roles as molecular chaperones involved in fibrillar procollagen folding and assembly, quality control, and secretion.     

Periplasmic chaperone recognition motif of subunits mediates quaternary interactions in the pilus.

The class of proteins collectively known as periplasmic immunoglobulin-like chaperones play an essential role in the assembly of a diverse set of adhesive organelles used by pathogenic strains of Gram-negative bacteria. Herein, we present a combination of genetic and structural data that sheds new light on chaperone-subunit and subunit-subunit interactions in the prototypical P pilus system, and provides new insights into how PapD controls pilus biogenesis. New crystallographic data of PapD with the C-terminal fragment of a subunit suggest a mechanism for how periplasmic chaperones mediate the extraction of pilus subunits from the inner membrane, a prerequisite step for subunit folding. In addition, the conserved N- and C-terminal regions of pilus subunits are shown to participate in the quaternary interactions of the mature pilus following their uncapping by the chaperone. By coupling the folding of subunit proteins to the capping of their nascent assembly surfaces, periplasmic chaperones are thereby able to protect pilus subunits from premature oligomerization until their delivery to the outer membrane assembly site.     

The Effect of Chemical Chaperones on the Assembly and Stability of HIV-1 Capsid Protein

Chemical chaperones are small organic molecules which accumulate in a broad range of organisms in various tissues under different stress conditions and assist in the maintenance of a correct proteostasis under denaturating environments. The effect of chemical chaperones on protein folding and aggregation has been extensively studied and is generally considered to be mediated through non-specific interactions. However, the precise mechanism of action remains elusive. Protein self-assembly is a key event in both native and pathological states, ranging from microtubules and actin filaments formation to toxic amyloids appearance in degenerative disorders, such as Alzheimer''s and Parkinson''s diseases. Another pathological event, in which protein assembly cascade is a fundamental process, is the formation of virus particles. In the late stage of the virus life cycle, capsid proteins self-assemble into highly-ordered cores, which encapsulate the viral genome, consequently protect genome integrity and mediate infectivity. In this study, we examined the effect of different groups of chemical chaperones on viral capsid assembly in vitro, focusing on HIV-1 capsid protein as a system model. We found that while polyols and sugars markedly inhibited capsid assembly, methylamines dramatically enhanced the assembly rate. Moreover, chemical chaperones that inhibited capsid core formation, also stabilized capsid structure under thermal denaturation. Correspondingly, trimethylamine N-oxide, which facilitated formation of high-order assemblies, clearly destabilized capsid structure under similar conditions. In contrast to the prevailing hypothesis suggesting that chemical chaperones affect proteins through preferential exclusion, the observed dual effects imply that different chaperones modify capsid assembly and stability through different mechanisms. Furthermore, our results indicate a correlation between the folding state of capsid to its tendency to assemble into highly-ordered structures.     

Molecular chaperones: new proteins--new functions]

A new class of cellular proteins named "molecular chaperones" has been described recently. Chaperones prevent from improper interactions either within or between polypeptide chains, which could produce incorrect structures. Chaperones assist in assembly or disassembly of oligomeric structures and in protein transport across membranes. There are three conservative families of chaperones and a few unrelated members. Some of them appeared to be stress proteins with yet unknown function. Mechanism of action and specificity of chaperone binding are under investigation now.     

The catalytic activity of Ubp6 enhances maturation of the proteasomal regulatory particle

The 26S proteasome is a 2.5 MDa macromolecular machine responsible for targeted protein degradation. Recently, four chaperones were identified that promote the assembly of the 19S regulatory particle (RP). Here, we probe the dynamic architecture of the proteasome by applying quantitative proteomics and mass spectrometry (MS) of intact complexes to provide a detailed characterization of how Ubp6 assists this assembly process. Our MS data demonstrate stoichiometric binding of chaperones and Ubp6 to the basal part of the RP. Genetic interactions of Ubp6 with Hsm3, but not with the other chaperones, indicate a functional overlay with Hsm3. Our biochemical data identified Ubp6 as an additional member of the Hsm3 module. Deletions of ubp6 with hsm3 perturb 26S proteasome assembly, which we attribute to an accumulation of ubiquitylated substrates on these assembly precursors. We therefore propose that Ubp6 facilitates proteasomal assembly by clearing ubiquitylated substrates from assembly precursors by its deubiquitylating activity.     

Water and molecular chaperones act as weak links of protein folding networks: energy landscape and punctuated equilibrium changes point towards a game theory of proteins

Water molecules and molecular chaperones efficiently help the protein folding process. Here we describe their action in the context of the energy and topological networks of proteins. In energy terms water and chaperones were suggested to decrease the activation energy between various local energy minima smoothing the energy landscape, rescuing misfolded proteins from conformational traps and stabilizing their native structure. In kinetic terms water and chaperones may make the punctuated equilibrium of conformational changes less punctuated and help protein relaxation. Finally, water and chaperones may help the convergence of multiple energy landscapes during protein-macromolecule interactions. We also discuss the possibility of the introduction of protein games to narrow the multitude of the energy landscapes when a protein binds to another macromolecule. Both water and chaperones provide a diffuse set of rapidly fluctuating weak links (low affinity and low probability interactions), which allow the generalization of all these statements to a multitude of networks.     

Hsp110 is required for spindle length control

Systematic affinity purification combined with mass spectrometry analysis of N- and C-tagged cytoplasmic Hsp70/Hsp110 chaperones was used to identify new roles of Hsp70/Hsp110 in the cell. This allowed the mapping of a chaperone-protein network consisting of 1,227 unique interactions between the 9 chaperones and 473 proteins and highlighted roles for Hsp70/Hsp110 in 14 broad biological processes. Using this information, we uncovered an essential role for Hsp110 in spindle assembly and, more specifically, in modulating the activity of the widely conserved kinesin-5 motor Cin8. The role of Hsp110 Sse1 as a nucleotide exchange factor for the Hsp70 chaperones Ssa1/Ssa2 was found to be required for maintaining the proper distribution of kinesin-5 motors within the spindle, which was subsequently required for bipolar spindle assembly in S phase. These data suggest a model whereby the Hsp70-Hsp110 chaperone complex antagonizes Cin8 plus-end motility and prevents premature spindle elongation in S phase.     

乙型肝炎病毒与细胞相互作用的研究进展

乙型肝炎病毒( HBV) 是一种类反转录( retroid) 病毒, 其DNA 基因组短小, 编码能力有限。但HBV 却能成功地在不同宿主个体中繁衍, 并在人群中广泛传播。本文着重对HBV 与宿主细胞关系的研究进展进行综述, 以利于阐明HBV 复制及其在宿主细胞内持续存在的特性。这些内容主要来自于本实验室的研究。文中讨论的HBV 与宿主之间的相互作用主要包括: 细胞伴侣分子在HBV 反转录起始和核衣壳组装中的作用,宿主通过调控核衣壳磷酸化对病毒衣壳成熟的调节以及与HBV 持续存在相关的核内附加体( episomal) 病毒DNA 的形成。