László Vígh

Dynamic, yet structured: The cell membrane three decades after the Singer–Nicolson model

The fluid mosaic membrane model proved to be a very useful hypothesis in explaining many, but certainly not all, phenomena taking place in biological membranes. New experimental data show that the compartmentalization of membrane components can be as important for effective signal transduction as is the fluidity of the membrane. In this work, we pay tribute to the Singer–Nicolson model, which is near its 30th anniversary, honoring its basic features, “mosaicism” and “diffusion,” which predict the interspersion of proteins and lipids and their ability to undergo dynamic rearrangement via Brownian motion. At the same time, modifications based on quantitative data are proposed, highlighting the often genetically predestined, yet flexible, multilevel structure implementing a vast complexity of cellular functions. This new “dynamically structured mosaic model” bears the following characteristics: emphasis is shifted from fluidity to mosaicism, which, in our interpretation, means nonrandom codistribution patterns of specific kinds of membrane proteins forming small-scale clusters at the molecular level and large-scale clusters (groups of clusters, islands) at the submicrometer level. The cohesive forces, which maintain these assemblies as principal elements of the membranes, originate from within a microdomain structure, where lipid–lipid, protein–protein, and protein–lipid interactions, as well as sub- and supramembrane (cytoskeletal, extracellular matrix, other cell) effectors, many of them genetically predestined, play equally important roles. The concept of fluidity in the original model now is interpreted as permissiveness of the architecture to continuous, dynamic restructuring of the molecular- and higher-level clusters according to the needs of the cell and as evoked by the environment.

HSP72 protects against obesity-induced insulin resistance

Patients with type 2 diabetes have reduced gene expression of heat shock protein (HSP) 72, which correlates with reduced insulin sensitivity. Heat therapy, which activates HSP72, improves clinical parameters in these patients. Activation of several inflammatory signaling proteins such as c-jun amino terminal kinase (JNK), inhibitor of κB kinase, and tumor necrosis factor-α, can induce insulin resistance, but HSP 72 can block the induction of these molecules in vitro. Accordingly, we examined whether activation of HSP72 can protect against the development of insulin resistance. First, we show that obese, insulin resistant humans have reduced HSP72 protein expression and increased JNK phosphorylation in skeletal muscle. We next used heat shock therapy, transgenic overexpression, and pharmacologic means to overexpress HSP72 either specifically in skeletal muscle or globally in mice. Herein, we show that regardless of the means used to achieve an elevation in HSP72 protein, protection against diet- or obesity-induced hyperglycemia, hyperinsulinemia, glucose intolerance, and insulin resistance was observed. This protection was tightly associated with the prevention of JNK phosphorylation. These findings identify an essential role for HSP72 in blocking inflammation and preventing insulin resistance in the context of genetic obesity or high-fat feeding.

Does the membrane's physical state control the expression of heat shock and other genes?

Membranes provide the structural framework that divides cells from their environment and that, in eukaryotic cells, permits compartmentation. They are not simply passive barriers that are liable to be damaged during environmental challenge or pathological states, but are involved in cellular responses and in modulating intracellular signalling. Recent data show that the expression of several genes, particularly those that respond to changes in temperature, ageing or disease, is influenced and/or controlled by the membranes physical state.

Heat shock proteins as emerging therapeutic targets

Chaperones (stress proteins) are essential proteins to help the formation and maintenance of the proper conformation of other proteins and to promote cell survival after a large variety of environmental stresses. Therefore, normal chaperone function is a key factor for endogenous stress adaptation of several tissues. However, altered chaperone function has been associated with the development of several diseases; therefore, modulators of chaperone activities became a new and emerging field of drug development. Inhibition of the 90 kDa heat shock protein (Hsp)90 recently emerged as a very promising tool to combat various forms of cancer. On the other hand, the induction of the 70 kDa Hsp70 has been proved to be an efficient help in the recovery from a large number of diseases, such as, for example, ischemic heart disease, diabetes and neurodegeneration. Development of membrane‐interacting drugs to modify specific membrane domains, thereby modulating heat shock response, may be of considerable therapeutic benefit as well. In this review, we give an overview of the therapeutic approaches and list some of the key questions of drug development in this novel and promising therapeutic approach.

Small heat-shock proteins regulate membrane lipid polymorphism

Thermal stress in living cells produces multiple changes that ultimately affect membrane structure and function. We report that two members of the family of small heat-shock proteins (sHsp) (α-crystallin and Synechocystis HSP17) have stabilizing effects on model membranes formed of synthetic and cyanobacterial lipids. In anionic membranes of dimyristoylphosphatidylglycerol and dimyristoylphosphatidylserine, both HSP17 and α-crystallin strongly stabilize the liquid-crystalline state. Evidence from infrared spectroscopy indicates that lipid/sHsp interactions are mediated by the polar headgroup region and that the proteins strongly affect the hydrophobic core. In membranes composed of the nonbilayer lipid dielaidoylphosphatidylethanolamine, both HSP17 and α-crystallin inhibit the formation of inverted hexagonal structure and stabilize the bilayer liquid-crystalline state, suggesting that sHsps can modulate membrane lipid polymorphism. In membranes composed of monogalactosyldiacylglycerol and phosphatidylglycerol (both enriched with unsaturated fatty acids) isolated from Synechocystis thylakoids, HSP17 and α-crystallin increase the molecular order in the fluid-like state. The data show that the nature of sHsp/membrane interactions depends on the lipid composition and extent of lipid unsaturation, and that sHsps can regulate membrane fluidity. We infer from these results that the association between sHsps and membranes may constitute a general mechanism that preserves membrane integrity during thermal fluctuations.

The small heat shock proteins and their clients

Abstract.Small heat shock proteins are ubiquitous proteins found throughout all kingdoms. One of the most notable features is their large oligomeric structures with conserved structural organization. It is well documented that small heat shock proteins can capture unfolding proteins to form stable complexes and prevent their irreversible aggregation. In addition, small heat shock proteins coaggregate with aggregation-prone proteins for subsequent, efficient disaggregation of the protein aggregates. The release of substrate proteins from the transient reservoirs, i.e. complexes and aggregates with small heat shock proteins, and their refolding require cooperation with ATP-dependent chaperone systems. The amphitropic small heat shock proteins were shown to associate with membranes, although they do not contain transmembrane domains or signal sequences. Recent studies indicate that small heat shock proteins play an important role in membrane quality control and thereby potentially contribute to the maintenance of membrane integrity especially under stress conditions.

Synechocystis HSP17 is an amphitropic protein that stabilizes heat-stressed membranes and binds denatured proteins for subsequent chaperone-mediated refolding

The small heat shock proteins (sHSPs) are ubiquitous stress proteins proposed to act as molecular chaperones to prevent irreversible protein denaturation. We characterized the chaperone activity of Synechocystis HSP17 and found that it has not only protein-protective activity, but also a previously unrecognized ability to stabilize lipid membranes. Like other sHSPs, recombinant Synechocystis HSP17 formed stable complexes with denatured malate dehydrogenase and served as a reservoir for the unfolded substrate, transferring it to the DnaK/DnaJ/GrpE and GroEL/ES chaperone network for subsequent refolding. Large unilamellar vesicles made of synthetic and cyanobacterial lipids were found to modulate this refolding process. Investigation of HSP17-lipid interactions revealed a preference for the liquid crystalline phase and resulted in an elevated physical order in model lipid membranes. Direct evidence for the participation of HSP17 in the control of thylakoid membrane physical state in vivo was gained by examining an hsp17− deletion mutant compared with the isogenic wild-type hsp17+ revertant Synechocystis cells. We suggest that, together with GroEL, HSP17 behaves as an amphitropic protein and plays a dual role. Depending on its membrane or cytosolic location, it may function as a “membrane stabilizing factor” as well as a member of a multichaperone protein-folding network. Membrane association of sHSPs could antagonize the heat-induced hyperfluidization of specific membrane domains and thereby serve to preserve structural and functional integrity of biomembranes.

Membranes: a meeting point for lipids, proteins and therapies

•  Introduction •  Membrane lipid composition •  Membrane lipid structure •  Membrane lipid organization ‐  Why so many different lipids? ‐  Lipid mixing and demixing ‐  Lateral pressure ‐  Surface electrostatics •  Role of lipids in cell functions •  Lipid influence in transmembrane protein function ‐  Prokaryotic potassium channel (KcsA) ‐  Mechanosensitive channels ‐  Voltage‐gated potassium channel (KvAP) ‐  Nicotinic acetylcholine receptor (nAcChR) ‐  G protein‐coupled receptors ‐  Other examples •  Non‐permanent proteins in membranes ‐  Proteins that interact reversibly with the bilayers ‐  Proteins that interact irreversibly with the bilayers ‐  Proteins that interact weakly with the membrane ‐  Proteins that interact strongly with the membrane ‐  G proteins and their interactions with membranes ‐  Small monomeric G proteins: the Ras and Ras‐like family ‐  Protein kinase C •  Membrane microdomains and lipid mediators in the control of heat‐shock protein response ‐  Stress sensing and signalling: the membrane sensor theory ‐  Hsp signalling in cancer and diabetes ‐  The role of membrane microdomains ‐  Lipid mediators of the stress response •  A subpopulation of Hsps can interact with and translocate through membranes ‐  Hsp90 in eukaryotic membranes ‐  Hsp70 in cell membranes ‐  Hsp27‐membrane interactions ‐  Secreted Hsps ‐  Representative cases where Hsps interact with membranes or release from the cells •  Concluding remarks

Docosahexaenoic Acid Reduces Amyloid β Production via Multiple Pleiotropic Mechanisms

Alzheimer disease is characterized by accumulation of the β-amyloid peptide (Aβ) generated by β- and γ-secretase processing of the amyloid precursor protein (APP). The intake of the polyunsaturated fatty acid docosahexaenoic acid (DHA) has been associated with decreased amyloid deposition and a reduced risk in Alzheimer disease in several epidemiological trials; however, the exact underlying molecular mechanism remains to be elucidated. Here, we systematically investigate the effect of DHA on amyloidogenic and nonamyloidogenic APP processing and the potential cross-links to cholesterol metabolism in vivo and in vitro. DHA reduces amyloidogenic processing by decreasing β- and γ-secretase activity, whereas the expression and protein levels of BACE1 and presenilin1 remain unchanged. In addition, DHA increases protein stability of α-secretase resulting in increased nonamyloidogenic processing. Besides the known effect of DHA to decrease cholesterol de novo synthesis, we found cholesterol distribution in plasma membrane to be altered. In the presence of DHA, cholesterol shifts from raft to non-raft domains, and this is accompanied by a shift in γ-secretase activity and presenilin1 protein levels. Taken together, DHA directs amyloidogenic processing of APP toward nonamyloidogenic processing, effectively reducing Aβ release. DHA has a typical pleiotropic effect; DHA-mediated Aβ reduction is not the consequence of a single major mechanism but is the result of combined multiple effects.

Bimoclomol, a heat shock protein co-inducer, acts by the prolonged activation of heat shock factor-1 ☆ ☆☆

The novel hydroxylamine derivative, bimoclomol, has been shown previously to act as a co-inducer of several heat shock proteins (Hsp-s), enhancing the amount of these proteins produced following a heat shock compared to heat shock alone. Here we show that the co-inducing effect of bimoclomol on Hsp expression is mediated via the prolonged activation of the heat shock transcription factor (HSF-1). Bimoclomol effects are abolished in cells from mice lacking HSF-1. Moreover, bimoclomol binds to HSF-1 and induces a prolonged binding of HSF-1 to the respective DNA elements. Since HSF-1 does not bind to DNA in the absence of stress, the bimoclomol-induced extension of HSF-1/DNA interaction may contribute to the chaperone co-induction of bimoclomol observed previously. These findings indicate that bimoclomol may be of value in targeting HSF-1 so as to induce up-regulation of protective Hsp-s in a non-stressful manner and for therapeutic benefit.