Structural studies of the TGF-βs and their receptors - insights into evolution of the TGF-β superfamily

TGF-βs are small secreted signaling proteins that function as vital regulators of cellular growth and differentiation. They signal through a single pair of receptors, known as TβR-I and TβR-II, and are among the most recently evolved members of the signaling superfamily to which they belong. This review provides an overview of the TGF-β, BMP, and activin receptor complexes that have been determined over the past several years. These structures underscore the shared ancestry of the TGF-βs with the BMPs and activins, but also provide insight as to how the TGF-βs diverged from the BMPs and activins to bind and assemble their receptors in a distinct manner. These distinctive modes of receptor binding engender the TGF-βs with high specificity for their receptors and allow them to fulfill their essential functions in vivo without interference from the many other proteins of the superfamily.     

A constitutively active Gα subunit provides insights into the mechanism of G protein activation

The activation of Gα subunits of heterotrimeric G proteins by G protein-coupled receptors (GPCRs) is a critical event underlying a variety of biological responses. Understanding how G proteins are activated will require structural and biochemical analyses of GPCRs complexed to their G protein partners, together with structure-function studies of Gα mutants that shed light on the different steps in the activation pathway. Previously, we reported that the substitution of a glycine for a proline at position 56 within the linker region connecting the helical and GTP-binding domains of a Gα chimera, designated αT*, yields a more readily exchangeable state for guanine nucleotides. Here we show that GDP-GTP exchange on αT*(G56P), in the presence of the light-activated GPCR, rhodopsin (R*), is less sensitive to the β1γ1 subunit complex than to wild-type αT*. We determined the X-ray crystal structure for the αT*(G56P) mutant and found that the G56P substitution leads to concerted changes that are transmitted to the conformationally sensitive switch regions, the α4-β6 loop, and the β6 strand. The α4-β6 loop has been proposed to be a GPCR contact site that signals to the TCAT motif and weakens the binding of the guanine ring of GDP, whereas the switch regions are the contact sites for the β1γ1 complex. Collectively, these biochemical and structural data lead us to suggest that αT*(G56P) may be adopting a conformation that is normally induced within Gα subunits by the combined actions of a GPCR and a Gβγ subunit complex during the G protein activation event.     

Is receptor oligomerization causally linked to activation of the EGF receptor kinase?

Transduction of a signal from an extracellular peptide hormone to produce an intracellular response is often mediated by a cell surface receptor, which is usually a glycoprotein. The secondary intracellular signal(s) generated after hormone binding to the receptor have been intensively studied. The nature of the primary signal generated by ligand binding to the receptor is understood less well in most cases. The particular case of the epidermal growth factor (EGF) receptor is analyzed, and evidence for or against two dissimilar models of primary signal transduction is reviewed. Evidence for the most widely accepted current model is found to be unconvincing. Evidence for the other model is substantial but indirect; a direct test of this model remains to be done.     

Molecular insights into the neutrophils activation in zebu cattle during seasonal variation

Earlier, we reported the impact of season on neutrophils’ functional competence and also hypothesized that it could be the impact of different seasons on neutrophils’ activation. In cerebration, the present study aimed to provide insights into neutrophils’ activation in terms of phosphorylation of tyrosine containing proteins during different seasons. Ten Hariana cows participated in the study and eight times blood samples were collected from each animal (twice in each month) during each season. Phosphorylation of tyrosine proteins was evaluated using western blotting and immunolocalization using a fluorescent microscope. Immuno blotting identified six tyrosine-phophorylated proteins p28, p42, p44, p58, p84, and p104 in winter and rainy seasons, whereas, p84 protein was absent in summer season. Immunolocalization revealed positive immune reactivity (IR) for tyrosine-phosphorylated proteins and significantly (p < 0.05) lower percent of neutrophils showed positive IR during the summer season as compared to winter and rainy seasons. The results of the study evidently indicate the activation of neutrophils is mediated through tyrosine phosphorylation and this may be a probable reason behind the decreased neutrophils’ functional competence during the summer. Further studies are warranted to decipher the possible association between tyrosine phosphorylation and expression of surface receptors required for the recruitment of neutrophils.     

Partial activation of α7 nicotinic acetylcholine receptors: insights from molecular dynamics simulations

Nicotinic acetylcholine receptors (nAChRs) are drug targets for neuronal disorders and diseases. Partial agonists for nAChRs are currently being developed as drugs for the treatment of neurological diseases for their relative safety originated from reduced excessive stimulation. In the current study, molecular docking, molecular dynamics simulations and binding energy calculations were performed to theoretically investigate the interactions between the partial agonists, 4-OH-DMXBA and tropisetron with α7-nAChR. The results suggest that the partial agonists 4-OH-DMXBA and tropisetron bind with α7-nAChR in a binding mode similar to that with AChBP. The non-conserved residues in the binding sites contribute to the orientation deviation of these partial agonists from their orientation in AChBP. Energy calculation and decomposition using MM-GB/SA suggests that the van der Waals term (ΔE_VDW) is the main driving force for the binding of the partial agonists to α7-nAChR. The molecular dynamics simulations showed that the opening of the C-loop binding with the partial agonists is in-between the openings for the binding with the full agonist and in the apo state. This conformation difference for the C-loop sheds light on the partial agonism of nAChR.     

Something in the air? New insights into mammalian pheromones

Olfaction is the dominant sensory modality for most animals and chemosensory communication is particularly well developed in many mammals. Our understanding of this form of communication has grown rapidly over the last ten years since the identification of the first olfactory receptor genes. The subsequent cloning of genes for rodent vomeronasal receptors, which are important in pheromone detection, has revealed an unexpected diversity of around 250 receptors belonging to two structurally different classes. This review will focus on the chemical nature of mammalian pheromones and the complementary roles of the main olfactory system and vomeronasal system in mediating pheromonal responses. Recent studies using genetically modified mice and electrophysiological recordings have highlighted the complexities of chemosensory communication via the vomeronasal system and the role of this system in handling information about sex and genetic identity. Although the vomeronasal organ is often regarded as only a pheromone detector, evidence is emerging that suggests it might respond to a much broader variety of chemosignals.     

Which agonist properties are important for the activation of 5-HT3A receptors?

Purpose: Why do anesthetics not activate excitatory ligand-gated ion channels such as 5-HT₃ receptors in contrast to inhibitory ligand-gated ion channels? This study examines the actions of structural closely-related 5-HT derivatives and 5-HT constituent parts on 5-HT_3A receptors with the aim of finding simpler if not minimal agonists and thus determining requirements for successful agonist action. Experimental approach: Responses to 5-HT derivatives of human 5-HT_3A receptors stably expressed in HEK 293 cells have been examined with the patch-clamp technique in the outside-out configuration combined with a fast solution exchange system. Results: Phenol, pyrrole and alkyl amines, constituents of 5-HT, even at high concentrations, cannot activate 5-HT_3A receptors but they can inhibit them. To date, tyramines are the smallest known agonists. However, an aromatic ring is not required for activation as acetylcholine is also an agonist of similar strength. Conclusion: Simultaneous interactions of adequate strength at two separate subsites within the 5-HT binding domain appear to be essential for successful agonist function. Anesthetics either fail to achieve this or the activation they produce is so weak that it is masked by a comparatively very strong inhibition.     

Novel insights into the development and maintenance of the blood–brain barrier

The blood–brain barrier (BBB) is essential for maintaining homeostasis within the central nervous system (CNS) and is a prerequisite for proper neuronal function. The BBB is localized to microvascular endothelial cells that strictly control the passage of metabolites into and out of the CNS. Complex and continuous tight junctions and lack of fenestrae combined with low pinocytotic activity make the BBB endothelium a tight barrier for water soluble moleucles. In combination with its expression of specific enzymes and transport molecules, the BBB endothelium is unique and distinguishable from all other endothelial cells in the body. During embryonic development, the CNS is vascularized by angiogenic sprouting from vascular networks originating outside of the CNS in a precise spatio-temporal manner. The particular barrier characteristics of BBB endothelial cells are induced during CNS angiogenesis by cross-talk with cellular and acellular elements within the developing CNS. In this review, we summarize the currently known cellular and molecular mechanisms mediating brain angiogenesis and introduce more recently discovered CNS-specific pathways (Wnt/β?catenin, Norrin/Frizzled4 and hedgehog) and molecules (GPR124) that are crucial in BBB differentiation and maturation. Finally, based on observations that BBB dysfunction is associated with many human diseases such as multiple sclerosis, stroke and brain tumors, we discuss recent insights into the molecular mechanisms involved in maintaining barrier characteristics in the mature BBB endothelium.     

Why do leafcutter bees cut leaves? New insights into the early evolution of bees

Stark contrasts in clade species diversity are reported across the tree of life and are especially conspicuous when observed in closely related lineages. The explanation for such disparity has often been attributed to the evolution of key innovations that facilitate colonization of new ecological niches. The factors underlying diversification in bees remain poorly explored. Bees are thought to have originated from apoid wasps during the Mid-Cretaceous, a period that coincides with the appearance of angiosperm eudicot pollen grains in the fossil record. The reliance of bees on angiosperm pollen and their fundamental role as angiosperm pollinators have contributed to the idea that both groups may have undergone simultaneous radiations. We demonstrate that one key innovation--the inclusion of foreign material in nest construction--underlies both a massive range expansion and a significant increase in the rate of diversification within the second largest bee family, Megachilidae. Basal clades within the family are restricted to deserts and exhibit plesiomorphic features rarely observed among modern bees, but prevalent among apoid wasps. Our results suggest that early bees inherited a suite of behavioural traits that acted as powerful evolutionary constraints. While the transition to pollen as a larval food source opened an enormous ecological niche for the early bees, the exploitation of this niche and the subsequent diversification of bees only became possible after bees had evolved adaptations to overcome these constraints.     

Recent insights into human bronchial proteomics – how are we progressing and what is next?

Introduction: The human respiratory system is highly prone to diseases and complications. Many lung diseases, including lung cancer (LC), tuberculosis (TB), and chronic obstructive pulmonary disease (COPD) have been among the most common causes of death worldwide. Cystic fibrosis (CF), the most common genetic disease in Caucasians, has adverse impacts on the lungs. Bronchial proteomics plays a significant role in understanding the underlying mechanisms and pathogenicity of lung diseases and provides insights for biomarker and therapeutic target discoveries.

Areas covered: We overview the recent achievements and discoveries in human bronchial proteomics by outlining how some of the different proteomic techniques/strategies are developed and applied in LC, TB, COPD, and CF. Also, the future roles of bronchial proteomics in predictive proteomics and precision medicine are discussed.

Expert commentary: Much progress has been made in bronchial proteomics. Owing to the advances in proteomics, we now have better ability to isolate proteins from desired cellular compartments, greater protein separation methods, more powerful protein detection technologies, and more sophisticated bioinformatic techniques. These all contributed to our further understanding of lung diseases and for biomarker and therapeutic target discoveries.     

How do G-protein-coupled receptors work? The case of metabotropic glutamate and GABA receptors

G-protein coupled receptors (GPCRs) represent the largest membrane proteins family in animal genomes. Being the receptors for most hormones and neurotransmitters, these proteins play a central role in intercellular communication. GPCRs can be classified into several groups based on the sequence similarity of their common structural feature: the heptahelical domain. The metabotropic receptors for the main neurotransmitters glutamate and gamma-aminobutyric acid (GABA) belong to the class III of GPCRs, together with others receptors for Ca2+, for sweet and amino acid taste compounds and for some pheromones, as well as for odorants in fish. Besides their transmembrane heptahelical domain responsible for G-protein activation, most of class III receptors possess a large extracellular domain responsible for ligand recognition. The recent resolution of the structure of this binding domain of one of these receptors, the mGlu1 receptor, together with the recent demonstration that these receptors are dimers, revealed an original mechanism of activation for these GPCRs. Such data open new possibilities to develop drugs aimed at modulating these receptors, and raised a number of interesting questions on the activation mechanism of other GPCRs.     

New insights into the OSBP‒VAP cycle

VAP-A is a major endoplasmic reticulum (ER) receptor that allows this organelle to engage numerous membrane contact sites with other organelles. One highly studied example is the formation of contact sites through VAP-A interaction with Oxysterol-binding protein (OSBP). This lipid transfer protein transports cholesterol from the ER to the trans-Golgi network owing to the counter-exchange of the phosphoinositide PI(4)P. In this review, we highlight recent studies that advance our understanding of the OSBP cycle and extend the model of lipid exchange to other cellular contexts and other physiological and pathological conditions.     

Do dietary antioxidants alleviate the cost of immune activation? An experiment with greenfinches

Reactive oxygen and nitrogen species produced by metabolism and immune defenses can cause extensive damage to biomolecules. To counteract this damage, organisms rely on exogenous and endogenous antioxidants, although their relative importance in maintaining redox balance is unclear. We supplemented captive greenfinches with dietary antioxidants--carotenoids and vitamin E--and injected them with an inflammatory agent, phytohemagglutinin. Compared to controls, immune-challenged birds circulated more lipid peroxidation products but also increased total plasma antioxidativity. Carotenoid (but not vitamin E) supplementation generally reduced lipid peroxidation, but this did not compensate for the effects of immune activation. Levels of an endogenous antioxidant--uric acid--strongly contributed to plasma antioxidativity. We found no evidence that dietary antioxidants are immunostimulatory. These results demonstrate the antioxidant function of carotenoids in birds and show that simultaneous assessment of oxidative stress-driven damage, antioxidant barrier, and individual antioxidants is critical for explaining the potential costs of immune system activation.