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The TCP-1 ring complex (TRiC) is a multi-subunit group II chaperonin that assists nascent or misfolded proteins to attain their native conformation in an ATP-dependent manner. Functional studies in yeast have suggested that TRiC is an essential and generalized component of the protein-folding machinery of eukaryotic cells. However, TRiC's involvement in specific cellular processes within multicellular organisms is largely unknown because little validation of TRiC function exists in animals. Our in vivo analysis reveals a surprisingly specific role of TRiC in the biogenesis of skeletal muscle α-actin during sarcomere assembly in myofibers. TRiC acts at the sarcomere's Z-disk, where it is required for efficient assembly of actin thin filaments. Binding of ATP specifically by the TRiC subunit Cct5 is required for efficient actin folding in vivo. Furthermore, mutant α-actin isoforms that result in nemaline myopathy in patients obtain their pathogenic conformation via this function of TRiC. [ABSTRACT FROM AUTHOR]

Highlights: [•] Visualization of rotary ATPases on nanometre scale by cryo-electron microscopy. [•] In situ visualization of mitochondrial ATP synthase by electron cryo-tomography. [•] Functional implications of peripheral stalk structures and wobble theory. [Copyright &y& Elsevier]

Background Airway smooth muscle (ASM) contraction underpins airway constriction; however, underlying mechanisms for airway hyperresponsiveness (AHR) remain incompletely defined. CD151, a 4-transmembrane glycoprotein that associates with laminin-binding integrins, is highly expressed in the human lung. The role of CD151 in ASM function and its relationship to asthma have yet to be elucidated. Objective We sought to ascertain whether CD151 expression is clinically relevant to asthma and whether CD151 expression affects AHR. Methods Using immunohistochemical analysis, we determined the expression of CD151 in human bronchial biopsy specimens from patients with varying asthma severities and studied the mechanism of action of CD151 in the regulation of ASM contraction and bronchial caliber in vitro , ex vivo , and in vivo . Results The number of CD151 + ASM cells is significantly greater in patients with moderate asthma compared with those in healthy nonasthmatic subjects. From loss- and gain-of-function studies, we reveal that CD151 is required for and enhances G protein–coupled receptor (GPCR)–induced peak intracellular calcium release, the primary determinant of excitation-contraction coupling. We show that the localization of CD151 can also be perinuclear/cytoplasmic and offer an explanation for a novel functional role for CD151 in supporting protein kinase C (PKC) translocation to the cell membrane in GPCR-mediated ASM contraction at this site. Importantly, CD151 −/− mice are refractory to airway hyperreactivity in response to allergen challenge. Conclusions We identify a role for CD151 in human ASM contraction. We implicate CD151 as a determinant of AHR in vivo , likely through regulation of GPCR-induced calcium and PKC signaling. These observations have significant implications in understanding the mechanism for AHR and the efficacy of new and emerging therapeutics. [ABSTRACT FROM AUTHOR]

In this issue of Structure, Oot and colleagues present the crystal structure of the eukaryotic V-ATPase peripheral stalk in complex with one of its binding partners, revealing conformational flexibility that may be important for priming the complex for rapid disassembly in response to external stimuli. [Copyright &y& Elsevier]

Recent landmark studies applying analytical pharmacology approaches to the glucocorticoid receptor (GR) have demonstrated that different ligands can cause differential activation of distinct GR-regulated genes. Drawing on concepts of signalling bias from the field of G protein-coupled receptor (GPCR) biology, we speculate that ligand-dependent differences in GR signalling can be considered analogous to GPCR biased signalling, and thus can be quantitatively analysed in a similar way. This type of approach opens up the possibility of using rational structure-based drug optimisation strategies to improve the therapeutic selectivity of glucocorticoid drugs to maximise their efficacy and minimise adverse effects. [ABSTRACT FROM AUTHOR]

Abstract: Airway remodelling refers to the structural changes that occur in the airway wall in asthma. These include epithelial hyperplasia and metaplasia, subepithelial fibrosis, muscle cell hyperplasia and angiogenesis. These structural changes result in thickening of the airway wall, airway hyperresponsiveness (AHR), and a progressive irreversible loss of lung function. The precise sequence of events that take place during the remodelling process and the mechanisms regulating these changes remain poorly understood. It is thought that airway remodelling is initiated and promoted by repeated episodes of allergic inflammation that damage the surface epithelium of the airway. However, other mechanisms are also likely to contribute to this process. Moreover, the interrelationship between airway remodelling, inflammation and AHR has not been clearly defined. Currently, there are no effective treatments that halt or reverse the changes of airway remodelling and its effects on lung function. Glucocorticoids have been unable to eliminate the progression of remodelling changes and there is limited evidence of a beneficial effect from other available therapies. The search for novel therapies that can directly target individual components of the remodelling process should be made a priority. In this review, we describe the current understanding of the airway remodelling process and the mechanisms regulating its development. The impact of currently available asthma therapies on airway remodelling is also discussed. [Copyright &y& Elsevier]

F 1 F o ATP synthase interchanges phosphate transfer energy and proton motive force via a rotary catalytic mechanism and isolated F 1 -ATPase subcomplexes can also hydrolyze ATP to generate rotation of their central γ rotor subunit. As ATP is hydrolyzed, the F 1 -ATPase cycles through a series of conformational states that mediates unidirectional rotation of the rotor. However, even in the absence of a rotor, the α and β subunits are still able to pass through a series of conformations, akin to those that generate rotation. Here, we use cryoelectron microscopy to establish the structures of these rotorless states. These structures indicate that cooperativity in this system is likely mediated by contacts between the β subunit lever domains, irrespective of the presence of the γ rotor subunit. These findings provide insight into how long-range information may be transferred in large biological systems. [Display omitted] • The structures of "rotorless" F 1 -ATPase under three conditions • Suggest the transitions that may occur during hydrolysis by "rotorless" F 1 -ATPase • Regions important for cooperativity in this system are highlighted ATP synthase is an important enzyme that generates the majority of cellular energy. Sobti et al. used structural analyses to show how a component of this enzyme can operate in its simplest form. The study highlights an important region that is likely responsible for coordinating changes within the enzyme. [ABSTRACT FROM AUTHOR]

Organoids, bioprinted mini-tissues and body-on-a-chip technologies are poised to transform the practice of preclinical pharmacology, with a view to achieving better predictive value. We review the need for further refinement in static and dynamic biomechanical aspects of such microenvironments. Further consideration of the developments required in perfusion systems to enable delivery of an appropriate soluble microenvironment are argued. We place particular emphasis on a major deficiency in these systems, being the absence or aberrant circadian behaviour of cells used in such settings, and consider the technical challenges that are needing to be met in order to achieve rhythm-on-a-chip. [ABSTRACT FROM AUTHOR]

Summary Chaperonins are essential biological complexes assisting protein folding in all kingdoms of life. Whereas homooligomeric bacterial GroEL binds hydrophobic substrates non-specifically, the heterooligomeric eukaryotic CCT binds specifically to distinct classes of substrates. Sulfolobales , which survive in a wide range of temperatures, have evolved three different chaperonin subunits (α, β, γ) that form three distinct complexes tailored for different substrate classes at cold, normal, and elevated temperatures. The larger octadecameric β complexes cater for substrates under heat stress, whereas smaller hexadecameric αβ complexes prevail under normal conditions. The cold-shock complex contains all three subunits, consistent with greater substrate specificity. Structural analysis using crystallography and electron microscopy reveals the geometry of these complexes and shows a novel arrangement of the α and β subunits in the hexadecamer enabling incorporation of the γ subunit. [ABSTRACT FROM AUTHOR]

The study of mechanobiology is now widespread. The impact of cell and tissue mechanics on cellular responses is well appreciated. However, knowledge of the impact of cell and tissue mechanics on pharmacological responsiveness, and its application to drug screening and mechanistic investigations, have been very limited in scope. We emphasize the need for a heightened awareness of the important bidirectional influence of drugs and biomechanics in all living systems. We propose that the term ‘mechanopharmacology’ be applied to approaches that employ in vitro systems, biomechanically appropriate to the relevant (patho)physiology, to identify new drugs and drug targets. This article describes the models and techniques that are being developed to transform drug screening and evaluation, ranging from a 2D environment to the dynamic 3D environment of the target expressed in the disease of interest. [ABSTRACT FROM AUTHOR]

Glucocorticoids (GCs) have impressive anti-inflammatory and immunosuppressive effects and show a diversity of actions across a variety of cell phenotypes. Implicit in efforts to optimize GCs as anti-inflammatory agents for any or all indications is the notion that the relevant mechanism(s) of action of GCs are fully elucidated. However, recent advances in understanding GC signalling mechanisms have revealed remarkable complexity and contextual dependence, calling into question whether the mechanisms of action are sufficiently well-described to embark on optimization. In the current review, we address evidence for differences in the mechanism of action in different cell types and contexts, and discuss contrasts in mechanisms of glucocorticoid insensitivity, with a focus on asthma and Chronic Obstructive Pulmonary Disease (COPD). Given this complexity, we consider the potential breadth of impact and selectivity of strategies directed to reversing the glucocorticoid insensitivity. [ABSTRACT FROM AUTHOR]

Myocardial infarction (MI) and its resultant heart failure remains a major cause of death in the world. The current treatments for patients with MI are revascularization with thrombolytic agents or interventional procedures. These treatments have focused on restoring blood flow to the ischemic tissue to prevent tissue necrosis and preserve organ function. The restoration of blood flow after a period of ischemia, however, may elicit further myocardial damage, called reperfusion injury. Pharmacological interventions, such as antioxidant and Ca 2+ channel blockers, have shown premises in experimental settings; however, clinical studies have shown limited success. Thus, there is a need for the development of novel therapies to treat reperfusion injury. The therapeutic potential of glucocorticoid-regulated anti-inflammatory mediator annexin-A1 (ANX-A1) has recently been recognized in a range of systemic inflammatory disorders. ANX-A1 binds to and activates the family of formyl peptide receptors (G protein-coupled receptor family) to inhibit neutrophil activation, migration and infiltration. Until recently, studies on the cardioprotective actions of ANX-A1 and its peptide mimetics (Ac2-26, CGEN-855A) have largely focused on its anti-inflammatory effects as a mechanism of preserving myocardial viability following I–R injury. Our laboratory provided the first evidence of the direct protective action of ANX-A1 on myocardium, independent of inflammatory cells in vitro. We now review the potential for ANX-A1 based therapeutics to be seen as a “triple shield” therapy against myocardial I–R injury, limiting neutrophil infiltration and preserving both cardiomyocyte viability and contractile function. This novel therapy may thus represent a valuable clinical approach to improve outcome after MI. [ABSTRACT FROM AUTHOR]

Highlights: [•] A strategy for treating chronic respiratory disease is to reduce airspace fibrin. [•] Small molecule inhibitors of PAI-1 show beneficial effects in lung injury models. [•] uPA is an important mediator of lung inflammation and remodeling in injury and disease. [•] Preclinical characterization of uPA inhibitors as therapy for lung disease is required. [ABSTRACT FROM AUTHOR]

Synthetic glucocorticoids are among the most commonly used prescription medicines. Nevertheless, their clinical efficacy is accompanied by dose- and indication-limiting acute and chronic adverse effects. Intrinsic or acquired resistance to glucocorticoid actions may also limit clinical efficacy. In chronic inflammatory conditions there has been a considerable focus on understanding mechanism(s) of resistance in cells with a primary immune and/or inflammatory function. However, it has become increasingly accepted that a substantial part of the efficacy of glucocorticoid treatments derives from actions on ‘structural’ cell types (smooth muscle, fibroblasts, epithelia). In this article we review the mechanism of action of glucocorticoids on structural cells and contrast knowledge of resistance mechanisms between structural and inflammatory cell types, using asthma as an exemplar chronic inflammatory condition associated with glucocorticoid resistance. [ABSTRACT FROM AUTHOR]

The estradiol metabolite, 2-methoxyestradiol (2MEO), is currently being evaluated in Phase II clinical trials for the treatment of solid tumours and is undergoing preclinical evaluation for inflammatory conditions. The anti-proliferative/cytotoxic/pro-apoptotic effects on tumour and endothelial cells have conferred potential on this metabolite for a synergistic impact on tumour growth. Exploitation of this synergy of 2MEO has previously required the combination of well-established cytotoxic agents with newer anti-angiogenic agents. This article reviews the pharmacology of 2MEO and describes the limitations inherent in its residual estrogen receptor affinity. The extent to which the metabolite 2MEO embodies an optimised therapeutic candidate is discussed. The challenges involved in using rational (3D QSAR-based) drug design to optimise the activity profile of analogues of 2MEO to provide additional members of this new class of anti-tumour/anti-inflammatory drug are also outlined. [Copyright &y& Elsevier]

Details the symposium of pharmacological scientists concerning airway smooth muscle (ASM) research in May 2002 in Durham, North Carolina. Structure of the symposium; Citations of the attendees; Highlights on the aspects of ASM.