ERK activation by Ca2+ ionophores depends on Ca2+ entry in lymphocytes but not in platelets, and does not conduct membrane scrambling

Rapid Ca²⁺-dependent phospholipid (PL) reorganization (scrambling) at the plasma membrane is a mechanism common to hematopoietic cells exposing procoagulant phosphatidylserine (PS). The aim of this research was to determine whether activation of the extracellular signal-regulated kinase (ERK) pathway was required for PL scrambling, based on a single report analyzing both responses induced by Ca²⁺ ionophores in megakaryoblastic HEL cells. Ca²⁺ ionophore-stimulated ERK phosphorylation was induced in platelets without external Ca²⁺, whereas exogenous Ca²⁺ entry was crucial for ERK activation in Jurkat T cells. In both cells, membrane scrambling only occurred following Ca²⁺ entry and was not blocked by inhibiting ERK phosphorylation. Furthermore, ERK proteins are strongly phosphorylated in transformed B lymphoblastic cell lines, which do not expose PS in their resting state. Overall, the data demonstrated that ERK activation and membrane scrambling are independent mechanisms. A. Arachiche, I. Badirou: These authors contributed equally to this work. Received 18 June 2008; received after revision 24 September 2008; accepted 1 October 2008     

Olfactory receptor trafficking to the plasma membrane

Olfactory receptors typically exhibit poor plasma membrane localization and functionality when heterologously expressed in most cell types. It has therefore proven difficult to effectively study olfactory receptor pharmacology and signaling mechanisms using traditional cell culture systems. Over the past few years, a variety of distinct proteins have been reported to interact with olfactory receptors and facilitate olfactory receptor trafficking to the plasma membrane in heterologous cells. Advances in this area have shed significant light on the fundamental factors governing the cell-specific control of olfactory receptor trafficking.     

Announcement

Announcement

Announcement     

Biological and Potential Therapeutic Roles of Sirtuin Deacetylases

Sirtuins comprise a unique class of nicotinamide adenine dinucleotide (NAD⁺)-dependent deacetylases that target multiple protein substrates to execute diverse biological functions. These enzymes are key regulators of clinically important cellular and organismal processes, including metabolism, cell division and aging. The desire to understand the important determinants of human health and lifespan has resulted in a firestorm of work on the seven mammalian sirtuins in less than a decade. The implication of sirtuins in medically important areas such as diabetes, cancer, cardiovascular dysfunction and neurodegenerative disease has further catapulted them to a prominent status as potential targets for nutritional and therapeutic development. Here, we present a review of published results on sirtuin biology and its relevance to human disease. Received 25 June 2008; received after revision 20 August 2008; accepted 29 August 2008     

Na+ binding to meizothrombin desF1

Meizothrombin is the physiologically active intermediate generated by a single cleavage of prothrombin at R320 to separate the A and B chains. Recent evidence has suggested that meizothrombin, like thrombin, is a Na(+)-activated enzyme. In this study we present the first X-ray crystal structure of human meizothrombin desF1 solved in the presence of the active site inhibitor PPACK at 2.1 A resolution. The structure reveals a Na(+) binding site whose architecture is practically identical to that of human thrombin. Stopped-flow measurements of Na(+) binding to meizothrombin desF1 document a slow phase of fluorescence change with a k(obs) decreasing hyperbolically with increasing [Na(+)], consistent with the existence of three conformations in equilibrium, E*, E and E:Na(+), as for human thrombin. Evidence that meizothrombin exists in multiple conformations provides valuable new information for studies of the mechanism of prothrombin activation.     

Zinc binding to peptide analogs of the structural zinc site in alcohol dehydrogenase: Implications for an entatic state

Zinc binding to the peptide replica and analogs to residues 93–115 of horse liver alcohol dehydrogenase (ADH) was examined by competition of the peptides and the chromophoric chelator 4-(2- pyridylazo)resorcinol for zinc and X-ray absorption fine structure analysis of the zinc ligands. In the enzyme, zinc is coordinated by four Cys residues. In the peptide replica, zinc is bound to three Cys and one His residue. A four-Cys zinc coordination is observed only when His is removed, leading to increased zinc stability. ADH crystal structures reveal that the ε-amino group of the conserved residue Lys323 is within H-bond distance of the backbone amide oxygens of residues 103, 105 and 108, likely stabilizing the zinc coordination in the enzyme. The peptide data thus indicate structural strain and increased energy in the zinc-binding site in the protein, characteristic of an entatic state, implying a functional nature for this zinc site. Received 3 July 2008; received after revision 11 August 2008; accepted 1 September 2008     

Molecular mechanisms of phagocytic uptake in mammalian cells

Phagocytosis is a highly conserved, complex process that has evolved to counter the constant threat posed by pathogens, effete cells and debris. Classically defined as a mechanism for internalising and destroying particles greater than 0.5 mum in size, it is a receptor-mediated, actin-driven process. The best-studied phagocytic receptors are the opsono-receptors, FcgammaR and CR3. Phagocytic uptake involves actin dynamics including polymerisation, bundling, contraction, severing and depolymerisation of actin filaments. Recent evidence points to the importance of membrane remodelling during phagocytosis, both in terms of changes in lipid composition and delivery of new membrane to the sites of particle binding. Here we review the molecular mechanisms of phagocytic uptake and some of the strategies developed by microbial pathogens to manipulate this process.     

The three-fingered protein domain of the human genome

Extracellular domains of some cellular receptors expressed in the organisms at different levels of development belong to three-fingered protein (TFP) fold. The Homo sapiens genome encodes at least 45 genes containing from one to three TFP domains (TFPDs), namely diverse paralogues of the Ly6 gene, CD59 and the receptors of activins, bone morphogenetic proteins, Mullerian inhibiting substance and transforming growth factor-β. C4.4a and urokinase/plasminogen activatory receptor contain two and three TFPD repeats, respectively. These diverse proteins have a low overall sequence similarity with each other and their hydrophobicity levels vary to a considerable degree. It is suggested that sequence differentiation within the TFPD led to distinct groups of proteins whose attributes were optimized to fit both the physicochemical properties specific to their functional microenvironment and selective targeting of their highly diversified extracellular cofactors. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Received 7 August 2008; accepted 29 August 2008     

Common variants near MC4R are associated with fat mass, weight and risk of obesity

To identify common variants influencing body mass index (BMI), we analyzed genome-wide association data from 16,876 individuals of European descent. After previously reported variants in FTO, the strongest association signal (rs17782313, P = 2.9 x 10(-6)) mapped 188 kb downstream of MC4R (melanocortin-4 receptor), mutations of which are the leading cause of monogenic severe childhood-onset obesity. We confirmed the BMI association in 60,352 adults (per-allele effect = 0.05 Z-score units; P = 2.8 x 10(-15)) and 5,988 children aged 7-11 (0.13 Z-score units; P = 1.5 x 10(-8)). In case-control analyses (n = 10,583), the odds for severe childhood obesity reached 1.30 (P = 8.0 x 10(-11)). Furthermore, we observed overtransmission of the risk allele to obese offspring in 660 families (P (pedigree disequilibrium test average; PDT-avg) = 2.4 x 10(-4)). The SNP location and patterns of phenotypic associations are consistent with effects mediated through altered MC4R function. Our findings establish that common variants near MC4R influence fat mass, weight and obesity risk at the population level and reinforce the need for large-scale data integration to identify variants influencing continuous biomedical traits.