bluuebunny/crossref_metadata_2025 huggingface dataset:
Protein aggregation, metals and oxidative stress in neurodegenerative diseases
Authors: Tabner B.J. , El-Agnaf O.M.A. , German M.J. , Fullwood N.J. , Allsop D.
Published: August 2015

There is clear evidence implicating oxidative stress in the pathology of many different neurodegenerative diseases. ROS (reactive oxygen species) are the primary mediators of oxidative stress and many of the aggregating proteins and peptides associated with neurodegenerative disease can generate hydrogen peroxide, a key ROS, apparently through interactions with redox-active metal ions. Our recent results suggest that ROS are generated during the very early stages of protein aggregation, when protofibrils or soluble oligomers are present, but in the absence of mature amyloid fibrils. The generation of ROS during early-stage protein aggregation may be a common, fundamental molecular mechanism underlying the pathogenesis of oxidative damage, neurodegeneration and cell death in several different neurodegenerative diseases. Drugs that specifically target this process could be useful in the future therapy of these diseases.
Compartmentation of G-protein-coupled receptors and their signalling components in lipid rafts and caveolae
Authors: Insel P.A. , Head B.P. , Patel H.H. , Roth D.M. , Bundey R.A. , Swaney J.S.
Published: August 2015

G-protein-coupled receptors (GPCRs) and post-GPCR signalling components are expressed at low overall abundance in plasma membranes, yet they evoke rapid, high-fidelity responses. Considerable evidence suggests that GPCR signalling components are organized together in membrane microdomains, in particular lipid rafts, enriched in cholesterol and sphingolipids, and caveolae, a subset of lipid rafts that also possess the protein caveolin, whose scaffolding domain may serve as an anchor for signalling components. Caveolae were originally identified based on their morphological appearance but their role in compartmentation of GPCR signalling has been primarily studied by biochemical techniques, such as subcellular fractionation and immunoprecipitation. Our recent studies obtained using both microscopic and biochemical methods with adult cardiac myocytes show expression of caveolin not only in surface sarcolemmal domains but also at, or close to, internal regions located at transverse tubules/sarcoplasmic reticulum. Other results show co-localization in lipid rafts/caveolae of AC (adenylyl cyclase), in particular AC6, certain GPCRs, G-proteins and eNOS (endothelial nitric oxide synthase; NOS3), which generates NO, a modulator of AC6. Existence of multiple caveolin-rich microdomains and their expression of multiple modulators of signalling strengthen the evidence that caveolins and lipid rafts/caveolae organize and regulate GPCR signal transduction in eukaryotic cells.
Expression cloning of novel regulators of 92 kDa type IV collagenase expression
Authors: Nair R.R. , Boyd D.D.
Published: August 2015

Overexpression of the 92 kDa type IV collagenase (MMP-9) contributes to cancer progression. However, to date, there are few known regulators of expression of this metalloproteinase. We employed an expression library comprising 500000 cDNA clones to screen for novel regulators of MMP-9 expression. HT1080 cells were transiently co-transfected with an MMP-9 promoter-luciferase reporter and pools of the cDNA expression library. Positive-scoring pools were subdivided in secondary and tertiary screens, after which the regulatory cDNAs were identified by DNA sequencing. This brief review illustrates the utility of expression cloning in identifying specific regulators of MMP-9 expression.
Formation and mobilization of neutral lipids in the yeast Saccharomyces cerevisiae
Authors: Wagner A. , Daum G.
Published: August 2015

Since energy storage is a basic metabolic process, the synthesis of neutral lipids occurs in all kingdoms of life. The yeast Saccharomyces cerevisiae, widely accepted as a model eukaryotic cell, contains two classes of neutral lipids, namely STEs (steryl esters) and TAGs (triacylglycerols). TAGs are synthesized through two pathways governed by the acyl-CoA diacylglycerol acyltransferase Dga1p and the phospholipid diacylglycerol acyltransferase Lro1p. STEs are formed by two STE synthases Are1p and Are2p, two enzymes with overlapping function, which also catalyse TAG formation, although to a minor extent. Neutral lipids are stored in the so-called lipid particles and can be utilized for membrane formation under conditions of lipid depletion. For this purpose, storage lipids have to be mobilized by TAG lipases and STE hydrolases. A TAG lipase named Tgl3p was identified as a major yeast TAG hydrolytic enzyme in lipid particles. Recently, a new family of hydrolases was detected which is required for STE mobilization in S. cerevisiae. These enzymes, named Yeh1p, Yeh2p and Tgl1p, are paralogues of the mammalian acid lipase family. The role of these proteins in biosynthesis and mobilization of TAG and STE, and the regulation of these processes will be discussed in this minireview.
Homoeostatic systems for sterols and other lipids
Authors: Garbarino J. , Sturley S.L.
Published: August 2015

Fatty acids and sterols are vital components of all eukaryotic cells. Both are used as building blocks for numerous cellular processes such as membrane biosynthesis or hormone production (sterols). Furthermore, these compounds elicit a variety of effects intracellularly as they can act as signalling molecules and regulate gene expression. The metabolism of fatty acids and sterols represents a very intricate network of pathways that are regulated in a precise manner in order to maintain lipid homoeostasis within a cell. Using the budding yeast Saccharomyces cerevisiae as a model system, we touch upon some of the aspects of achieving and maintaining this lipid homoeostasis.
Access of the substrate to the active site of squalene and oxidosqualene cyclases: comparative inhibition, site-directed mutagenesis and homology-modelling studies
Authors: Oliaro-Bosso S. , Schulz-Gasch T. , Taramino S. , Scaldaferri M. , Viola F. , Balliano G.
Published: August 2015

Substrate access to the active-site cavity of squalene-hopene cyclase from Alicyclobacillus acidocaldarious and lanosterol synthase [OSC (oxidosqualene cyclase)] from Saccharomyces cerevisiae was studied by an inhibition, mutagenesis and homology-modelling approach. Crystal structure and homology modelling indicate that both enzymes possess a narrow constriction that separates an entrance lipophilic channel from the active-site cavity. The role of the constriction as a mobile gate that permits substrate passage was investigated by experiments in which critically located Cys residues, either present in native protein or inserted by site-directed mutagenesis, were labelled with specifically designed thiol-reacting molecules. Some amino acid residues of the yeast enzyme, selected on the basis of sequence alignment and a homology model, were individually replaced by residues bearing side chains of different lengths, charges or hydrophobicities. In some of these mutants, substitution severely reduced enzymatic activity and thermal stability. Homology modelling revealed that in these mutants some critical stabilizing interactions could no longer occur. The possible critical role of entrance channel and constriction in specific substrate recognition by eukaryotic OSC is discussed.
Arf, Sec7 and Brefeldin A: a model towards the therapeutic inhibition of guanine nucleotide-exchange factors
Authors: Zeghouf M. , Guibert B. , Zeeh J.-C. , Cherfils J.
Published: August 2015

GEFs (guanine nucleotide-exchange factors), which stimulate GDP dissociation from small G-proteins, are pivotal regulators of signalling pathways activated by small G-proteins. In the case of Arf proteins, which are major regulators of membrane traffic in the cell and have recently been found to be involved in an increasing number of human diseases, GDP/GTP exchange is stimulated by GEFs that carry a catalytic Sec7 domain. Recent structural results captured snapshots of the exchange reaction, revealing that Sec7 domains secure Arf-GDP to membranes before nucleotide exchange takes place, taking advantage of a built-in structural device in Arf proteins that couples their affinity for membranes to the nature of the bound nucleotide. One of the Arf–Sec7 intermediates was trapped by BFA (Brefeldin A), an uncompetitive inhibitor of Arf activation that has been instrumental in deciphering the molecular principles of membrane traffic at the Golgi. BFA targets a low-affinity Arf–Sec7 intermediate of the exchange reaction. It binds at the Arf-GDP/Sec7 interface, thus freezing the complex in an abortive conformation that cannot proceed to nucleotide dissociation. In the cell, this results in the specific inhibition of Arf1 by a subset of its GEFs, and the efficient and reversible block of membrane traffic at the Golgi. The mechanism of BFA leads to the concept of ‘interfacial inhibition’, in which a protein–protein interaction of therapeutic interest is stabilized, rather than impaired, by a drug. Up-regulated activity of small G-proteins is involved in various human diseases, making their GEFs attractive candidates to interrupt specifically the corresponding signalling pathway. Interfacial inhibitors are proposed as an alternative to competitive inhibitors that may be explored for their inhibition.
Arfs, phosphoinositides and membrane traffic
Authors: Donaldson J.G.
Published: August 2015

Arf (ADP-ribosylation factor) GTP-binding proteins function in cells to regulate membrane traffic and structure. Arfs accomplish this task through modification of membrane lipids and the recruitment of proteins, including coat proteins and actin, to membrane surfaces. Arf1 and Arf6 are the most divergent and most studied human Arf proteins that localize predominantly to the Golgi complex and plasma membrane respectively. We have been studying the targeting of Arf1 and Arf6 to these specific compartments and the common and divergent activities that they exert on these membranes. We have found that Arf6 acts through activation of type I phosphatidylinositol 4-phosphate 5-kinases to generate phosphatidylinositol 4,5-bisphosphate and that this activity is instrumental in facilitating the actin cytoskeletal rearrangements and alterations in endosomal membrane trafficking observed with increased Arf6 activation. Arf1 can also stimulate the activity of phosphatidylinositol kinases and recruit coat proteins and actin cytoskeletal elements to the Golgi complex.
Transcriptional regulation and metabolism
Authors: Brynildsen M.P. , Wong W.W. , Liao J.C.
Published: August 2015

Understanding organisms from a systems perspective is essential for predicting cellular behaviour as well as designing gene-metabolic circuits for novel functions. The structure, dynamics and interactions of cellular networks are all vital components of systems biology. To facilitate investigation of these aspects, we have developed an integrative technique called network component analysis, which utilizes mRNA expression and transcriptional network connectivity to determine network component dynamics, functions and interactions. This approach has been applied to elucidate transcription factor dynamics in Saccharomyces cerevisiae cell-cycle regulation, detect cross-talks in Escherichia coli two-component signalling pathways, and characterize E. coli carbon source transition. An ultimate test of system-wide understanding is the ability to design and construct novel gene-metabolic circuits. To this end, artificial feedback regulation, cell–cell communication and oscillatory circuits have been constructed, which demonstrate the design principles of gene-metabolic regulation in the cell.
Chromosome positioning is largely unaffected in lymphoblastoid cell lines containing emerin or A-type lamin mutations
Authors: Meaburn K.J. , Levy N. , Toniolo D. , Bridger J.M.
Published: August 2015

Gene-poor human chromosomes are reproducibly found at the nuclear periphery in proliferating cells. There are a number of inner nuclear envelope proteins that may have roles in chromosome location and anchorage, e.g. emerin and A-type lamins. In the last decade, a number of diseases associated with tissue degeneration and premature aging have been linked with mutations in lamin A or emerin. These are termed laminopathies, with mutations in emerin causing Emery–Dreifuss muscular dystrophy. Despite highly aberrant nuclear distributions of A-type lamins and emerin in lymphoblastoid cell lines derived from patients with emerin or lamin A mutations, little or no change in chromosome location was detected.
The regulation of cell migration by PTEN
Authors: Leslie N.R. , Yang X. , Downes C.P. , Weijer C.J.
Published: August 2015

In vertebrates, the tumour suppressor PTEN (phosphatase and tensin homologue deleted on chromosome 10) regulates many cellular processes through its PtdIns(3,4,5)P3 lipid phosphatase activity, antagonizing PI3K (phosphoinositide 3-kinase) signalling. Given the important role of PI3Ks in the regulation of directed cell migration and the role of PTEN as an inhibitor of migration, it is somewhat surprising that data now indicate that PTEN is able to regulate cell migration independent of its lipid phosphatase activity. Here, we discuss the role of PTEN in the regulation of cell migration.
Myostatin: a modulator of skeletal-muscle stem cells
Authors: Walsh F.S. , Celeste A.J.
Published: August 2015

Myostatin, or GDF-8 (growth and differentiation factor-8), was first identified through sequence identity with members of the BMP (bone morphogenetic protein)/TGF-β (transforming growth factor-β) superfamily. The skeletal-muscle-specific expression pattern of myostatin suggested a role in muscle development. Mice with a targeted deletion of the myostatin gene exhibit a hypermuscular phenotype. In addition, inactivating mutations in the myostatin gene have been identified in ‘double muscled’ cattle breeds, such as the Belgian Blue and Piedmontese, as well as in a hypermuscular child. These findings define myostatin as a negative regulator of skeletal-muscle development. Myostatin binds with high affinity to the receptor serine threonine kinase ActRIIB (activin type IIB receptor), which initiates signalling through a smad2/3-dependent pathway. In an effort to validate myostatin as a therapeutic target in a post-embryonic setting, a neutralizing antibody was developed by screening for inhibition of myostatin binding to ActRIIB. Administration of this antimyostatin antibody to adult mice resulted in a significant increase in both muscle mass and functional strength. Importantly, similar results were obtained in a murine model of muscular dystrophy, the mdx mouse. Unlike the myostatin-deficient animals, which exhibit both muscle hypertrophy and hyperplasia, the antibody-treated mice demonstrate increased musculature through a hypertrophic mechanism. These results validate myostatin inhibition as a therapeutic approach to muscle wasting diseases such as muscular dystrophy, sarcopenic frailty of the elderly and amylotrophic lateral sclerosis.
Embryonic stem (ES) cells and embryonal carcinoma (EC) cells: opposite sides of the same coin
Authors: Andrews P.W. , Matin M.M. , Bahrami A.R. , Damjanov I. , Gokhale P. , Draper J.S.
Published: August 2015

Embryonal carcinoma (EC) cells are the stem cells of teratocarcinomas, and the malignant counterparts of embryonic stem (ES) cells derived from the inner cell mass of blastocyst-stage embryos, whether human or mouse. On prolonged culture in vitro, human ES cells acquire karyotypic changes that are also seen in human EC cells. They also ‘adapt’, proliferating faster and becoming easier to maintain with time in culture. Furthermore, when cells from such an ‘adapted’ culture were inoculated into a SCID (severe combined immunodeficient) mouse, we obtained a teratocarcinoma containing histologically recognizable stem cells, which grew out when the tumour was explanted into culture and exhibited properties of the starting ES cells. In these features, the ‘adapted’ ES cells resembled malignant EC cells. The results suggest that ES cells may develop in culture in ways that mimic changes occurring in EC cells during tumour progression.
Cancer stem cells: AMLs show the way
Authors: Bonnet D.
Published: August 2015

The blood-related cancer leukaemia was the first disease where human CSCs (cancer stem cells), or LSCs (leukaemic stem cells), were isolated. The haematopoietic system is one of the best tissues for investigating CSCs, since the developmental hierarchy of normal blood formation is well defined. Leukaemia can now be viewed as aberrant haematopoietic processes initiated by rare LSCs that have maintained or reacquired the capacity for indefinite proliferation through accumulated mutations and/or epigenetic changes. Yet, despite their critical importance, much remains to be learned about the developmental origin of LSCs and the mechanisms responsible for their emergence in the course of the disease. This report will review our current knowledge on LSC development and finally demonstrate how these discoveries provide a paradigm for identification of CSCs from solid tumours.
Regulation of protein synthesis by insulin
Authors: Proud C.G.
Published: August 2015

Insulin rapidly activates protein synthesis by activating components of the translational machinery including eIFs (eukaryotic initiation factors) and eEFs (eukaryotic elongation factors). In the long term, insulin also increases the cellular content of ribosomes to augment the capacity for protein synthesis. The rapid activation of protein synthesis by insulin is mediated primarily through phosphoinositide 3-kinase. This involves the activation of PKB (protein kinase B). In one case, PKB acts to phosphorylate and inactivate glycogen synthase kinase 3, which in turn phosphorylates and inhibits eIF2B. Insulin elicits the dephosphorylation and activation of eIF2B. Since eIF2B is required for recycling of eIF2, a factor required for all cytoplasmic translation initiation events, this will contribute to overall activation of protein synthesis. PKB also phosphorylates the TSC1 (tuberous sclerosis complex 1)–TSC2 complex to relieve its inhibitory action on the mTOR (mammalian target of rapamycin). Inhibition of mTOR by rapamycin markedly impairs insulin-activated protein synthesis. mTOR controls translation initiation and elongation. The cap-binding factor eIF4E can be sequestered in inactive complexes by 4E-BP1 (eIF4E-binding protein 1). Insulin elicits phosphorylation of 4E-BP1 and its release from eIF4E, allowing eIF4E to form initiation factor complexes. Insulin induces dephosphorylation and activation of eEF2 to accelerate elongation. Both effects are blocked by rapamycin. Insulin inactivates eEF2 kinase by increasing its phosphorylation at several mTOR-regulated sites. Insulin also stimulates synthesis of ribosomal proteins by promoting recruitment of their mRNAs into polyribosomes. This is inhibited by rapamycin. Several key questions remain about, for example, the mechanisms by which mTOR controls 4E-BP1 and eEF2 kinase and the control of ribosomal protein translation.
Regulation of the pyruvate dehydrogenase complex
Authors: Patel M.S. , Korotchkina L.G.
Published: August 2015

The PDC (pyruvate dehydrogenase complex) plays a central role in the maintenance of glucose homoeostasis in mammals. The carbon flux through the PDC is meticulously controlled by elaborate mechanisms involving post-translational (short-term) phosphorylation/dephosphorylation and transcriptional (long-term) controls. The former regulatory mechanism involving multiple phosphorylation sites and tissue-specific distribution of the dedicated kinases and phosphatases is not only dependent on the interactions among the catalytic and regulatory components of the complex but also sensitive to the intramitochondrial redox state and metabolite levels as indicators of the energy status. Furthermore, differential transcriptional controls of the regulatory components of PDC further add to the complexity needed for long-term tuning of PDC activity for the maintenance of glucose homoeostasis during normal and disease states.
Regulation of acetyl-CoA carboxylase
Authors: Brownsey R.W. , Boone A.N. , Elliott J.E. , Kulpa J.E. , Lee W.M.
Published: August 2015

Acetyl-CoA carboxylase (ACC) catalyses the formation of malonyl-CoA, an essential substrate for fatty acid synthesis in lipogenic tissues and a key regulatory molecule in muscle, brain and other tissues. ACC contributes importantly to the overall control of energy metabolism and has provided an important model to explore mechanisms of enzyme control and hormone action. Mammalian ACCs are multifunctional dimeric proteins (530–560 kDa) with the potential to further polymerize and engage in multiprotein complexes. The enzymatic properties of ACC are complex, especially considering the two active sites, essential catalytic biotin, the three-substrate reaction and effects of allosteric ligands. The expression of the two major isoforms and splice variants of mammalian ACC is tissue-specific and responsive to hormones and nutritional status. Key regulatory elements and cognate transcription factors are still being defined. ACC specific activity is also rapidly modulated, being increased in response to insulin and decreased following exposure of cells to catabolic hormones or environmental stress. The acute control of ACC activity is the product of integrated changes in substrate supply, allosteric ligands, the phosphorylation of multiple serine residues and interactions with other proteins. This review traces the path and implications of studies initiated with Dick Denton in Bristol in the late 1970s, through to current proteomic and other approaches that have been consistently challenging and immensely rewarding.
Remodelling Ca2+ signalling systems and cardiac hypertrophy
Authors: Berridge M.J.
Published: August 2015

In cardiac cells, Ca2+ signals appear as brief transients responsible for controlling both contraction and transcription. Information may be encoded in these digital signals through changes in both frequency and shape. An increase in Ca2+ signalling contributes to a process of phenotypic remodelling during hypertrophy. The increase in Ca2+ that drives the larger contractions may be responsible for switching on a second process of signalosome remodelling to down-regulate the Ca2+ signalling pathway. It is a change in the properties of the Ca2+ transient that seems to carry the information responsible for the remodelling of the cardiac gene transcription programme that leads first to hypertrophy and then to congestive heart failure.
The impact of molecular biology on drug discovery
Authors: Belfield G.P. , Delaney S.J.
Published: August 2015

The discipline of molecular biology has become increasingly important in recent times for the process of drug discovery. We describe the impact of molecular biology across the whole process of drug discovery and development, including (i) the identification and validation of new drug targets, (ii) the development of molecular screens to find new candidate drugs, and (iii) the generation of safety data and competences leading to enhanced clinical efficacy. We also speculate on emerging developments in drug discovery where it seems likely that molecular biology will play an even more vital role in the generation of future therapies.
Recent Advances in Membrane Biochemistry
Authors: East J. Malcolm , Michelangeli Francesco
Published: August 2015

This Biochemical Society Annual Symposium on Recent Advances in Membrane Biochemistry was organized to bring together experts from across the spectrum of biomembrane disciplines from the biological to the biophysical/structural, with the intention of promoting interactions and collaborations across the field. We were keen that the potential for improving human health that stems from a deeper understanding of membrane structure/function should be acknowledged, especially in the light of the increasing numbers of membrane protein structures that continue to be made available to the biomembrane community. This foreword provides an idea of what was communicated in the various sessions and, we hope, gives an impression of the excitement generated by the speakers and delegates at this over-subscribed Symposium.
Understanding the yeast host cell response to recombinant membrane protein production
Authors: Bawa Zharain , Bland Charlotte E. , Bonander Nicklas , Bora Nagamani , Cartwright Stephanie P. , Clare Michelle , Conner Matthew T. , Darby Richard A.J. , Dilworth Marvin V. , Holmes William J. , Jamshad Mohammed , Routledge Sarah J. , Gross Stephane R. , Bill Roslyn M.
Published: August 2015

Membrane proteins are drug targets for a wide range of diseases. Having access to appropriate samples for further research underpins the pharmaceutical industry's strategy for developing new drugs. This is typically achieved by synthesizing a protein of interest in host cells that can be cultured on a large scale, allowing the isolation of the pure protein in quantities much higher than those found in the protein's native source. Yeast is a popular host as it is a eukaryote with similar synthetic machinery to that of the native human source cells of many proteins of interest, while also being quick, easy and cheap to grow and process. Even in these cells, the production of human membrane proteins can be plagued by low functional yields; we wish to understand why. We have identified molecular mechanisms and culture parameters underpinning high yields and have consolidated our findings to engineer improved yeast host strains. By relieving the bottlenecks to recombinant membrane protein production in yeast, we aim to contribute to the drug discovery pipeline, while providing insight into translational processes.
Switching the centromeres on and off: epigenetic chromatin alterations provide plasticity in centromere activity stabilizing aberrant dicentric chromosomes
Authors: Sato Hiroshi , Saitoh Shigeaki
Published: August 2015

The kinetochore, which forms on a specific chromosomal locus called the centromere, mediates interactions between the chromosome and the spindle during mitosis and meiosis. Abnormal chromosome rearrangements and/or neocentromere formation can cause the presence of multiple centromeres on a single chromosome, which results in chromosome breakage or cell cycle arrest. Analyses of artificial dicentric chromosomes suggested that the activity of the centromere is regulated epigenetically; on some stably maintained dicentric chromosomes, one of the centromeres no longer functions as a platform for kinetochore formation, although the DNA sequence remains intact. Such epigenetic centromere inactivation occurs in cells of various eukaryotes harbouring ‘regional centromeres’, such as those of maize, fission yeast and humans, suggesting that the position of the active centromere is determined by epigenetic markers on a chromosome rather than the nucleotide sequence. Our recent findings in fission yeast revealed that epigenetic centromere inactivation consists of two steps: disassembly of the kinetochore initiates inactivation and subsequent heterochromatinization prevents revival of the inactivated centromere. Kinetochore disassembly followed by heterochromatinization is also observed in normal senescent human cells. Thus epigenetic centromere inactivation may not only stabilize abnormally generated dicentric chromosomes, but also be part of an intrinsic mechanism regulating cell proliferation.
Emerging insights into the mechanistic link between α-synuclein and glucocerebrosidase in Parkinson's disease
Authors: McGlinchey Ryan P. , Lee Jennifer C.
Published: August 2015

Mutations in the GBA1 gene, encoding the enzyme glucocerebrosidase, cause the lysosomal storage disorder GD (Gaucher’s disease), and are associated with the development of PD (Parkinson's disease) and other Lewy body disorders. Interestingly, GBA1 variants are the most common genetic risk factor associated with PD. Although clinical studies argue a strong case towards a link between GBA1 mutations and the development of PD, mechanistic insights have been lacking. In the present article, we review recent findings that have provided some biochemical evidence to bridge this relationship, focusing on the molecular link between two proteins, α-synuclein and glucocerebrosidase, involved in PD and GD respectively.
Streptococcus zooepidemicus and Streptococcus equi evolution: the role of CRISPRs
Authors: Waller Andrew S. , Robinson Carl
Published: August 2015

The host-restricted bacterium Streptococcus equi is the causative agent of equine strangles, the most frequently diagnosed infectious disease of horses worldwide. The disease is characterized by abscessation of the lymph nodes of the head and neck, leading to significant welfare and economic cost. S. equi is believed to have evolved from an ancestral strain of Streptococcus zooepidemicus, an opportunistic pathogen of horses and other animals. Comparison of the genome of S. equi strain 4047 with those of S. zooepidemicus identified examples of gene loss due to mutation and deletion, and gene gain through the acquisition of mobile genetic elements that have probably shaped the pathogenic specialization of S. equi. In particular, deletion of the CRISPR (clustered regularly interspaced short palindromic repeats) locus in the ancestor of S. equi may have predisposed the bacterium to acquire and incorporate new genetic material into its genome. These include four prophages and a novel integrative conjugative element. The virulence cargo carried by these mobile genetic elements is believed to have shaped the ability of S. equi to cause strangles. Further sequencing of S. zooepidemicus has highlighted the diversity of this opportunistic pathogen. Again, CRISPRs are postulated to influence evolution, balancing the need for gene gain over genome stability. Analysis of spacer sequences suggest that these pathogens may be susceptible to a limited range of phages and provide further evidence of cross-species exchange of genetic material among Streptococcus pyogenes, Streptococcus agalactiae and Streptococcus dysgalactiae.
Gliotransmission: focus on exocytotic release of L-glutamate and D-serine from astrocytes
Authors: Martineau Magalie
Published: August 2015

The release of neuromodulators, called gliotransmitters, by astrocytes is proposed to modulate neurotransmission and synaptic plasticity, and thereby cognitive functions; but they are also proposed to have a role in diverse neurological disorders. Two main routes have been proposed to ensure gliotransmitter release: non-exocytotic release from cytosolic pools through plasma membrane proteins, and Ca2+-regulated exocytosis through the fusion of gliotransmitter-storing secretory organelles. Regulated Ca2+-dependent glial exocytosis has received much attention and is appealing since its existence endows astrocytes with some of the basic properties thought to be exclusive to neurons and neuroendocrine cells. The present review summarizes recent findings regarding the exocytotic mechanisms underlying the release of two excitatory amino acids, L-glutamate and D-serine.
Checkpoint regulation of replication forks: global or local?
Authors: Iyer Divya Ramalingam , Rhind Nicholas
Published: August 2015

Cell-cycle checkpoints are generally global in nature: one unattached kinetochore prevents the segregation of all chromosomes; stalled replication forks inhibit late origin firing throughout the genome. A potential exception to this rule is the regulation of replication fork progression by the S-phase DNA damage checkpoint. In this case, it is possible that the checkpoint is global, and it slows all replication forks in the genome. However, it is also possible that the checkpoint acts locally at sites of DNA damage, and only slows those forks that encounter DNA damage. Whether the checkpoint regulates forks globally or locally has important mechanistic implications for how replication forks deal with damaged DNA during S-phase.
The serine shuttle between glia and neurons: implications for neurotransmission and neurodegeneration
Authors: Wolosker Herman , Radzishevsky Inna
Published: August 2015

D-Serine is a physiological co-agonist of NMDARs (N-methyl-D-aspartate receptors) required for neurotransmission, synaptic plasticity and neurotoxicity. There is no consensus, however, on the relative roles of neurons and astrocytes in D-serine signalling. The effects of D-serine had been attributed to its role as a gliotransmitter specifically produced and released by astrocytes. In contrast, recent studies indicate that neurons regulate their own NMDARs by releasing D-serine via plasma membrane transporters and depolarization-sensitive pathways. Only a minority of astrocytes contain authentic D-serine, whereas neuronal D-serine accounts for up to 90% of the total D-serine pool. Neuronal and glial D-serine production requires astrocytic L-serine generated by a 3-phosphoglycerate dehydrogenase-dependent pathway. These findings support a model whereby astrocyte-derived L-serine shuttles to neurons to fuel the synthesis of D-serine by serine racemase. We incorporate these new findings in a revised model of serine dynamics, called the glia–neuron serine shuttle, which highlights the role of glia–neuron cross-talk for optimal NMDAR activity and brain development.
Mitochondrial trafficking in neurons and the role of the Miro family of GTPase proteins
Authors: Birsa Nicol , Norkett Rosalind , Higgs Nathalie , Lopez-Domenech Guillermo , Kittler Josef T.
Published: August 2015

Correct mitochondrial dynamics are essential to neuronal function. These dynamics include mitochondrial trafficking and quality-control systems that maintain a precisely distributed and healthy mitochondrial network, so that local energy demands or Ca2+-buffering requirements within the intricate architecture of the neuron can be met. Mitochondria make use of molecular machinery that couples these organelles to microtubule-based transport via kinesin and dynein motors, facilitating the required long-range movements. These motors in turn are associated with a variety of adaptor proteins allowing additional regulation of the complex dynamics demonstrated by these organelles. Over recent years, a number of new motor and adaptor proteins have been added to a growing list of components implicated in mitochondrial trafficking and distribution. Yet, there are major questions that remain to be addressed about the regulation of mitochondrial transport complexes. One of the core components of this machinery, the mitochondrial Rho GTPases Miro1 (mitochondrial Rho 1) and Miro2 have received special attention due to their Ca2+-sensing and GTPase abilities, marking Miro an exceptional candidate for co-ordinating mitochondrial dynamics and intracellular signalling pathways. In the present paper, we discuss the wealth of literature regarding Miro-mediated mitochondrial transport in neurons and recently highlighted involvement of Miro proteins in mitochondrial turnover, emerging as a key process affected in neurodegeneration.
Dynein, microtubule and cargo: a ménage à trois
Authors: Pavin Nenad , Tolić-Nørrelykke Iva M.
Published: August 2015

To exert forces, motor proteins bind with one end to cytoskeletal filaments, such as microtubules and actin, and with the other end to the cell cortex, a vesicle or another motor. A general question is how motors search for sites in the cell where both motor ends can bind to their respective binding partners. In the present review, we focus on cytoplasmic dynein, which is required for a myriad of cellular functions in interphase, mitosis and meiosis, ranging from transport of organelles and functioning of the mitotic spindle to chromosome movements in meiotic prophase. We discuss how dynein targets sites where it can exert a pulling force on the microtubule to transport cargo inside the cell.
Astrocyte glutamine synthetase: pivotal in health and disease
Authors: Rose Christopher F. , Verkhratsky Alexei , Parpura Vladimir
Published: August 2015

The multifunctional properties of astrocytes signify their importance in brain physiology and neurological function. In addition to defining the brain architecture, astrocytes are primary elements of brain ion, pH and neurotransmitter homoeostasis. GS (glutamine synthetase), which catalyses the ATP-dependent condensation of ammonia and glutamate to form glutamine, is an enzyme particularly found in astrocytes. GS plays a pivotal role in glutamate and glutamine homoeostasis, orchestrating astrocyte glutamate uptake/release and the glutamate–glutamine cycle. Furthermore, astrocytes bear the brunt of clearing ammonia in the brain, preventing neurotoxicity. The present review depicts the central function of astrocytes, concentrating on the importance of GS in glutamate/glutamine metabolism and ammonia detoxification in health and disease.
The membranes of Gram-negative bacteria: progress in molecular modelling and simulation
Authors: Khalid Syma , Berglund Nils A. , Holdbrook Daniel A. , Leung Yuk M. , Parkin Jamie
Published: August 2015

Molecular modelling and simulations have been employed to study the membranes of Gram-negative bacteria for over 20 years. Proteins native to these membranes, as well as antimicrobial peptides and drug molecules have been studied using molecular dynamics simulations in simple models of membranes, usually only comprising one lipid species. Thus, traditionally, the simulations have reflected the majority of in vitro membrane experimental setups, enabling observations from the latter to be rationalized at the molecular level.

In the last few years, the sophistication and complexity of membrane models have improved considerably, such that the heterogeneity of the lipid and protein composition of the membranes can now be considered both at the atomistic and coarse-grain levels of granularity. Importantly this means relevant biology is now being retained in the models, thereby linking the in silico and in vivo scenarios. We discuss recent progress in simulations of proteins in simple lipid bilayers, more complex membrane models and finally describe some efforts to overcome timescale limitations of atomistic molecular dynamics simulations of bacterial membranes.
How many lives does CLIMP-63 have?
Authors: Sandoz Patrick A. , van der Goot F. Gisou
Published: August 2015

In 1995, in the Biochemical Society Transactions, Mundy published the first review on CLIMP-63 (cytoskeleton-linking membrane protein 63) or CKPA4 (cytoskeleton-associated protein 4), initially just p63 [1]. Here we review the following 20 years of research on this still mysterious protein. CLIMP-63 is a type II transmembrane protein, the cytosolic domain of which has the capacity to bind microtubules whereas the luminal domain can form homo-oligomeric complexes, not only with neighbouring molecules but also, in trans, with CLIMP-63 molecules on the other side of the endoplasmic reticulum (ER) lumen, thus promoting the formation of ER sheets. CLIMP-63 however also appears to have a life at the cell surface where it acts as a ligand-activated receptor. The still rudimentary information of how CLIMP-63 fulfills these different roles, what these are exactly and how post-translational modifications control them, will be discussed.
Engineering activated protein C to maximize therapeutic efficacy
Authors: Quinn Louise M. , Drakeford Clive , O’Donnell James S. , Preston Roger J.S.
Published: August 2015

The anticoagulant-activated protein C (APC) acts not solely as a crucial regulator of thrombus formation following vascular injury, but also as a potent signalling enzyme with important functions in the control of both acute and chronic inflammatory disease. These properties have been exploited to therapeutic effect in diverse animal models of inflammatory disease, wherein recombinant APC administration has proven to effectively limit disease progression. Subsequent clinical trials led to the use of recombinant APC (Xigris) for the treatment of severe sepsis. Although originally deemed successful, Xigris was ultimately withdrawn due to lack of efficacy and an unacceptable bleeding risk. Despite this apparent failure, the problems that beset Xigris usage may be tractable using protein engineering approaches. In this review, we detail the protein engineering approaches that have been utilized to improve the therapeutic characteristics of recombinant APC, from early studies in which the distinct anti-coagulant and signalling activities of APC were separated to reduce bleeding risk, to current attempts to enhance APC cytoprotective signalling output for increased therapeutic efficacy at lower APC dosage. These novel engineered variants represent the next stage in the development of safer, more efficacious APC therapy in disease settings in which APC plays a protective role.
Dual regulation of transcription factor Nrf2 by Keap1 and by the combined actions of β-TrCP and GSK-3
Authors: Hayes John D. , Chowdhry Sudhir , Dinkova-Kostova Albena T. , Sutherland Calum
Published: August 2015

Nuclear factor-erythroid 2 p45 (NF-E2 p45)-related factor 2 (Nrf2) is a master regulator of redox homoeostasis that allows cells to adapt to oxidative stress and also promotes cell proliferation. In this review, we describe the molecular mechanisms by which oxidants/electrophilic agents and growth factors increase Nrf2 activity. In the former case, oxidants/electrophiles increase the stability of Nrf2 by antagonizing the ability of Kelch-like ECH-associated protein 1 (Keap1) to target the transcription factor for proteasomal degradation via the cullin-3 (Cul3)–RING ubiquitin ligase CRLKeap1. In the latter case, we speculate that growth factors increase the stability of Nrf2 by stimulating phosphoinositide 3-kinase (PI3K)−protein kinase B (PKB)/Akt signalling, which in turn results in inhibitory phosphorylation of glycogen synthase kinase-3 (GSK-3) and in doing so prevents the formation of a DSGIS motif-containing phosphodegron in Nrf2 that is recognized by the β-transducin repeat-containing protein (β-TrCP) Cul1-based E3 ubiquitin ligase complex SCFβ-TrCP. We present data showing that in the absence of Keap1, the electrophile tert-butyl hydroquinone (tBHQ) can stimulate Nrf2 activity and induce the Nrf2-target gene NAD(P)H:quinone oxidoreductase-1 (NQO1), whilst simultaneously causing inhibitory phosphorylation of GSK-3β at Ser9. Together, these observations suggest that tBHQ can suppress the ability of SCFβ-TrCP to target Nrf2 for proteasomal degradation by increasing PI3K−PKB/Akt signalling. We also propose a scheme that explains how other protein kinases that inhibit GSK-3 could stimulate induction of Nrf2-target genes by preventing formation of the DSGIS motif-containing phosphodegron in Nrf2.
Redox-based regulation of neural stem cell function and Nrf2
Authors: Madhavan Lalitha
Published: August 2015

Neural stem cells (NSCs) play vital roles in the development and maintenance of brain tissues throughout life. They can also potentially act as powerful sources of regeneration and repair during pathology to replace degenerating cells and counteract deleterious changes in the tissue microenvironment. However, both aging and neurodegeneration involve an up-regulation of processes, such as oxidative stress, inflammation, somatic mutations, and reduction in growth factors in neural tissues, which threaten the robust functioning of NSCs. Nevertheless, recent evidence also indicates that NSCs may possess the intrinsic capability to cope with such stressors in their microenvironment. Whereas the mechanisms governing the responses of NSCs to stress are diverse, a common theme that is emerging suggests that underlying changes in intracellular redox status are crucial. Here we discuss such redox-based regulation of NSCs, particularly in relation to nuclear erythroid factor 2-like 2 (Nrf2), which is a key cellular stress resistance factor, and its implications for successfully harnessing NSC therapeutic potential towards developing cell-based therapeutics for nervous system disorders.
Targeting mitochondrial energy metabolism with TSPO ligands
Authors: Gut Philipp
Published: August 2015

The translocator protein (18 kDa) (TSPO) resides on the outer mitochondrial membrane where it is believed to participate in cholesterol transport and steroid hormone synthesis. Although it is almost ubiquitously expressed, what TSPO does in non-steroidogenic tissues is largely unexplored. Recent studies report changes in glucose homoeostasis and cellular energy production when TSPO function is modulated by selective ligands or by genetic loss-of-function. This review summarizes findings that connect TSPO function with the regulation of mitochondrial energy metabolism. The juxtaposition of TSPO at the cytosolic/mitochondrial interface and the existence of endogenous ligands that are regulated by metabolism suggest that TSPO functions to adapt mitochondrial to cellular metabolism. From a pharmacological perspective the specific up-regulation of TSPO in neuro-inflammatory and injury-induced conditions make TSPO an interesting, druggable target of mitochondrial metabolism.
Sustained NRF2 activation in hereditary leiomyomatosis and renal cell cancer (HLRCC) and in hereditary tyrosinemia type 1 (HT1)
Authors: Sandhu Ivraj Singh , Maksim Nicholas James , Amouzougan Eva Alice , Gallion Bryce Wilson , Raviele Anthony L.J. , Ooi Aikseng
Published: August 2015

The nuclear erythroid 2-like 2 transcription factor (NRF2), is a major regulator of cellular redox balance. Although NRF2 activation is generally regarded as beneficial to human health, recent studies have identified that sustained NRF2 activation is over-represented in many cancers. This raises the question regarding the role of NRF2 activation in the development and progression of those cancers. This review focuses on the mechanisms and the effects of NRF2 activation in two hereditary cancer predisposition syndromes: hereditary leiomyomatosis and renal cell cancer (HLRCC) and hereditary tyrosinemia type 1 (HT1). Because the cancer initiating mutations in these hereditary syndromes are well defined, they offer a unique opportunity to explore the roles of NRF2 activation in the early stages of carcinogenesis. Over the years, a variety of approaches have been utilized to study the biology of HLRCC and HT1. In HLRCC, in vitro studies have demonstrated the importance of NRF2 activation in sustaining cancer cell proliferation. In the mouse model of HT1 however, NRF2 activation seems to protect cells from malignant transformation. In both HT1 and HLRCC, NRF2 activation promotes the clearance of electrophilic metabolites, enabling cells to survive cancer-initiating mutations. Biological insights gained from the hereditary syndromes’ studies may shed light on to the roles of NRF2 activation in sporadic tumours.