Showing papers in "Cellular and Molecular Life Sciences in 2001"

HuR and mRNA stability

Abstract: An important mechanism of posttranscriptional gene regulation in mammalian cells is the rapid degradation of messenger RNAs (mRNAs) signaled by AU-rich elements (AREs) in their 3′ untranslated regions. HuR, a ubiquitously expressed member of the Hu family of RNA-binding proteins related to Drosophila ELAV, selectively binds AREs and stabilizes ARE-containing mRNAs when overexpressed in cultured cells. This review discusses mRNA decay as a general form of gene regulation, decay signaled by AREs, and the role of HuR and its Hu-family relatives in antagonizing this mRNA degradation pathway. The influence of newly identified protein ligands to HuR on HuR function in both normal and stressed cells may explain how ARE-mediated mRNA decay is regulated in response to environmental change.

The glutathione peroxidases.

Abstract: There are several proteins in mammalian cells that can metabolize hydrogen peroxide and lipid hydroperoxides. These proteins include four selenium-containing glutathione peroxidases that are found in different cell fractions and tissues of the body. This review considers the structure and distribution of the selenoperoxidases and how this relates to their biological function. The functions of the selenoperoxidases were originally studied in systems where their activity was manipulated by changing dietary selenium levels. More recently, molecular techniques have allowed overexpression of selenoperoxidases in cell lines and animals. Additionally, cellular glutathione peroxidase knockout mice have been used to investigate the functions of this protein. From this work it is clear that the selenoperoxidases are involved in cell antioxidant systems. However, they also have more subtle functions in ensuring the regulation and formation of arachadonic acid metabolites that are derived from hydroperoxide intermediates. The range of biological processes, which are potentially dependent on optimal selenoperoxidase activity in mammals, emphasises the importance of achieving adequate selenium intake in the diet.

Polyamines in cell growth and cell death: molecular mechanisms and therapeutic applications.

Abstract: Polyamines are aliphatic cations with multiple functions and are essential for life. Cellular polyamine levels are regulated by multiple pathways such as synthesis from amino acid precursors, cellular uptake mechanisms that salvage polyamines from diet and intestinal microorganisms, as well as stepwise degradation and efflux. Investigations using polyamine biosynthetic inhibitors indicate that alterations in cellular polyamine levels modulate normal and cancer cell growth. Studies using transgenic mice overexpressing polyamine biosynthetic enzymes support a role of polyamines in carcinogenesis. Many, if not all, signal transduction pathways intersect with polyamine biosynthetic pathways and the regulation of intracellular polyamine levels. Direct binding of polyamines to DNA and their ability to modulate DNA-protein interactions appear to be important in the molecular mechanisms of polyamine action in cell proliferation. Consistent with the role of polyamines as facilitators of cell growth, several studies have shown their ability to protect cells from apoptosis. However, polyamines also have a role in facilitating cell death. The basis of these diverse cellular responses is currently not known. Cell death response might be partly mediated by the production of hydrogen peroxide during polyamine catabolism. In addition, the ability of polyamines to alter DNA-protein and protein-protein interactions might be disruptive to cellular functions, when abnormally high levels are accumulated due to defects in polyamine catabolic or efflux pathways. A large body of data indicates that polyamine pathway can be a molecular target for therapeutic intervention in several types cancers. Inhibitors of biosynthesis, polyamine analogues as well as oligonucleotide/polyamine analogue combinations are promising drug candidates for chemoprevention and/or treatment of cancer.

Cytochromes P450 and metabolism of xenobiotics.

Abstract: Cytochromes P450 (henceforth P450s) are involved in a variety of metabolic and biosynthetic processes. The number of known P450 enzymes exceeds 1000, while the endogenous substrates of most of them remain unknown. All P450 enzymes exhibit similarity in their structure and general mechanism of action; however, there are significant differences in the detailed function of individual enzymes as well as in the structures and properties of their active sites. This review discusses the properties of the most important P450 enzymes taking part in drug metabolism in humans. P450 3A4 is of paramount importance, because it is the most abundant P450 in the human liver and is known to metabolize the majority of drugs whose biotransformation is known. Genetically dependent variabilities of individual P450 activities and levels are described, documenting the importance of pharmacogenetics aimed at explaining differences in the response of the organism to various drugs.

Multifunctional alpha-enolase: its role in diseases.

Abstract: Enolase, a key glycolytic enzyme, belongs to a novel class of surface proteins which do not possess classical machinery for surface transport, yet through an unknown mechanism are transported on the cell surface. Enolase is a multifunctional protein, and its ability to serve as a plasminogen receptor on the surface of a variety of hematopoetic, epithelial and endothelial cells suggests that it may play an important role in the intravascular and pericellular fibrinolytic system. Its role in systemic and invasive autoimmune disorders was recognized only very recently. In addition to this property, its ability to function as a heat-shock protein and to bind cytoskeletal and chromatin structures indicate that enolase may play a crucial role in transcription and a variety of pathophysiological processes.

The CD40/CD154 receptor/ligand dyad.

Abstract: Until recently, the expression and primary function of the cell surface receptor CD40 and its ligand CD154 were considered restricted to B and T lymphocytes, and their interactions required for the thymus-dependent humoral response. However, current work from several groups challenges this view of the CD40/CD154 dyad as a mere mediator of lymphocyte communication. A variety of non-lymphocytic cell types express both receptor and ligand, including hematopoetic and non-hematopoetic cells, such as monocytes, basophils, eosinophils, dendritic cells, fibroblasts, smooth muscle, and endothelial cells. Accordingly, ligation of CD40 mediates a broad variety of immune and inflammatory responses, such as the expression of adhesion molecules, cytokines, matrix-degrading enzymes, prothrombotic activities, and apoptotic mediators. Consequently, CD40 signaling has been associated with pathogenic processes of chronic inflammatory diseases, such as autoimmune diseases, neurodegenerative disorders, graft-versus-host disease, cancer, and atherosclerosis. This review focuses on the synthesis and structure of CD40 and outlines CD154/CD40 signaling pathways, and emphasizes the previously unexpected importance of the CD40/CD154 receptor/ligand dyad in a spectrum of immunoregulatory processes and prevalent human diseases.

From MDR to MXR: new understanding of multidrug resistance systems, their properties and clinical significance.

Abstract: The ATP binding cassette (ABC) superfamily of membrane transporters is one of the largest protein classes known, and counts numerous proteins involved in the trafficking of biological molecules across cell membranes. The first known human ABC transporter was P-glycoprotein (P-gp), which confers multidrug resistance (MDR) to anticancer drugs. In recent years, we have obtained an increased understanding of the mechanism of action of P-gp as its ATPase activity, substrate specificity and pharmacokinetic interactions have been investigated. This review focuses on the functional characterization of P-gp, as well as other ABC transporters involved in MDR: the family of multidrug-resistance-associated proteins (MRP1-7), and the recently discovered ABC half-transporter MXR (also known as BCRP, ABCP and ABCG2). We describe recent progress in the analysis of protein structure-function relationships, and consider the conceptual problem of defining and identifying substrates and inhibitors of MDR. An in-depth discussion follows of how coupling of nucleotide hydrolysis to substrate transport takes place, and we propose a scheme for the mechanism of P-gp function. Finally, the clinical correlations, both for reversal of MDR in cancer and for drug delivery, are discussed.

Bromelain: biochemistry, pharmacology and medical use.

Abstract: Bromelain is a crude extract from the pineapple that contains, among other components, various closely related proteinases, demonstrating, in vitro and in vivo, antiedematous, antiinflammatory, antithrombotic and fibrinolytic activities. The active factors involved are biochemically characterized only in part. Due to its efficacy after oral administration, its safety and lack of undesired side effects, bromelain has earned growing acceptance and compliance among patients as a phytotherapeutical drug. A wide range of therapeutic benefits has been claimed for bromelain, such as reversible inhibition of platelet aggregation, angina pectoris, bronchitis, sinusitis, surgical traumas, thrombophlebitis, pyelonephritis and enhanced absorption of drugs, particularly of antibiotics. Biochemical experiments indicate that these pharmacological properties depend on the proteolytic activity only partly, suggesting the presence of nonprotein factors in bromelain. Recent results from preclinical and pharmacological studies recommend bromelain as an orally given drug for complementary tumor therapy: bromelain acts as an immunomodulator by raising the impaired immunocytotoxicity of monocytes against tumor cells from patients and by inducing the production of distinct cytokines such as tumor necrosis factor-α, interleukin (Il)-1β, Il-6, and Il-8. In a recent clinical study with mammary tumor patients, these findings could be partially confirmed. Especially promising are reports on animal experiments claiming an antimetastatic efficacy and inhibition of metastasis-associated platelet aggregation as well as inhibition of growth and invasiveness of tumor cells. Apparently, the antiinvasive activity does not depend on the proteolytic activity. This is also true for bromelain effects on the modulation of immune functions, its potential to eliminate burn debris and to accelerate wound healing. Whether bromelain will gain wide acceptance as a drug that inhibits platelet aggregation, is antimetastatic and facilitates skin debridement, among other indications, will be determined by further clinical trials. The claim that bromelain cannot be effective after oral administration is definitely refuted at this time.

Arabinogalactan-proteins: structure, expression and function.

Abstract: Arabinogalactan-proteins (AGPs) are a family of extensively glycosylated hydroxyproline-rich glycoproteins that are thought to have important roles in various aspects of plant growth and development. After a brief introduction to AGPs highlighting the problems associated with defining and classifying this diverse family of glycoproteins, AGP structure is described in terms of the protein component (including data from molecular cloning), carbohydrate component, processing of AGPs (including recent data on glycosylphosphatidylinositol membrane anchors) and overall molecular shape. Next, the expression of AGPs is examined at several different levels, from the whole plant to the cellular levels, using a variety of experimental techniques and tools. Finally, AGP function is considered. Although the existing functional evidence is not incontrovertible, it does clearly point to roles for AGPs in vegetative, reproductive, and cellular growth and development as well as programmed cell death and social control. In addition and most likely inextricably linked to their functions, AGPs are presumably involved in molecular interactions and cellular signaling at the cell surface. Some likely scenarios are discussed in this context. AGPs also have functions of real or potential commercial value, most notably as emulsifiers in the food industry and as potential immunological regulators for human health. Several important questions remain to be answered with respect to AGPs. Clearly, elucidating the unequivocal functions of particular AGPs and relating these functions to their respective structures and modes of action remain as major challenges in the years ahead.

Extracellular electron transfer

Abstract: Results from several laboratories indicate that extracellular electron transfer may be a general mechanism whereby microoorganisms generate energy for cell growth and/or maintenance. Specifically, bacteria can use redox-active organic small molecules, generated outside or inside the cells, to shuttle electrons between reduced and oxidized compounds. Electron shuttling has now been reported for several different bacterial species, and exchanges of shuttling compounds may even syntrophically link diverse organisms in nature. Biofilm systems in both geological and clinical settings are likely to be important environments for metabolisms that employ extracellular electron transfer. Both structural and functional analyses suggest that electron shuttles and some virulence factors may be related to one another.

The glycinergic inhibitory synapse.

Abstract: Glycine is one of the most important inhibitory neurotransmitters in the spinal cord and the brainstem, and glycinergic synapses have a well-established role in the regulation of locomotor behavior. Research over the last 15 years has yielded new insights on glycine neurotransmission. Glycinergic synapses are now known not to be restricted to the spinal cord and the brainstem. Presynaptic machinery for glycine release and uptake, the structure and function of postsynaptic receptors and the factors (both pre- and postsynaptic) which control the strength of glycinergic inhibition have been extensively studied. It is now established that glycinergic synapses can be excitatory in the immature brain and that some inhibitory synapses can corelease gamma-aminobutyric acid (GABA) and glycine. Moreover, the presence of glycine transporters on glial cells and the capacity of these cells to release glycine suggest that glycine may also act as a neuromodulator. Extensive molecular studies have revealed the presence of distinct subtypes of postsynaptic glycine receptors with different functional properties. Mechanisms of glycine receptors aggregation at postsynaptic sites during development are better understood and functional implications of variation in receptor number between postsynaptic sites are partly elucidated. Mutations of glycine receptor subunits have been shown to underly some human locomotor disorders, including the startle disease. Clearly, recent work on glycine receptor channels and the synapses at which they mediate inhibitory signalling in both young and adult animals necessitates an update of our vision of glycinergic inhibitory transmission.

WD-repeat proteins: structure characteristics, biological function, and their involvement in human diseases.

Abstract: Defined by the presence of four or more repeating units containing a conserved core of approximately 40 amino acids that usually ending with tryptophan-aspartic acid (WD), WD-repeat proteins belong to a large and fast-expanding conservative protein family. As demonstrated by the crystal structure of the G protein β subunit, all WD-repeat proteins are speculated to form a circularized β propeller structure. The importance of these proteins is not only demonstrated by their critical roles in many essential biological functions ranging from signal transduction, transcription regulation, to apoptosis, but is also recognized by their association with several human diseases. Defining the function of a WD-repeat protein is the current challenge. It is, however, paramount to uncover the function of individual WD-repeat proteins, explore the protein interaction mechanism through WD-repeat domains and, ultimately, understand the complex biological processes and organisms themselves.

How do thermophilic proteins deal with heat

Abstract: Recent years have witnessed an explosion of sequence and structural information for proteins from hyperthermophilic and thermophilic organisms. Complete genome sequences are available for many hyperthermophilic archaeons. Here, we review some recent studies on protein thermostability along with work from our laboratory. A large number of sequence and structural factors are thought to contribute toward higher intrinsic thermal stability of proteins from these organisms. The most consistent are surface loop deletion, increased occurrence of hydrophobic residues with branched side chains and an increased proportion of charged residues at the expense of uncharged polar residues. The energetic contribution of electrostatic interactions such as salt bridges and their networks toward protein stability can be stabilizing or destabilizing. For hyperthermophilic proteins, the contribution is mostly stabilizing. Macroscopically, improvement in electrostatic interactions and strengthening of hydrophobic cores by branched apolar residues increase the enthalpy change between the folded and unfolded states of a thermophilic protein. At the same time, surface loop deletion contributes to decreased conformational entropy and decreased heat capacity change between the folded and unfolded states of the protein.

Functional, biochemical and genetic diversity of prokaryotic nitrate reductases

Abstract: Prokaryotic nitrate reduction can serve a number of physiological roles and can be catalysed by a number of biochemically distinct nitrate reductases. Three distinct nitrate reductase classes can be indentified in prokaryotes, NAS, NAR and NAP. NAS is located in the cytoplasmic compartment and participates in nitrogen assimilation. NAR is usually a three-subunit complex anchored to the cytoplasmic face of the membrane with its active site located in the cytoplasmic compartment and is involved in anaerobic nitrate respiration. NAP is a two-subunit complex, located in the periplasmic compartment, that is coupled to quinol oxidation via a membrane anchored tetraheme cytochrome. It shows considerable functional flexibility by participating in anaerobic respiration or redox energy dissipation depending on the organism in which it is found. The members of all three classes of enzymes bind the bis-molybdopterin guanine dinucleotide cofactor at the active site, but they differ markedly in the number and nature of cofactors used to transfer electrons to this site. Analysis of prokaryotic genome sequences available at the time of writing reveals that the different nitrate reductases are phylogenetically widespread.

Three classes of C2H2 zinc finger proteins.

Abstract: C2H2 zinc finger proteins probably comprise the largest family of regulatory proteins in mammals. Most zinc fingers bind to a cognate DNA. In addition to DNA, many of the proteins also bind to RNA or protein, and some bind to RNA only. The binding properties depend on the amino acid sequence of the finger domains and of the linker between fingers, as well as on the higher-order structures and the number of fingers. C2H2 zinc finger proteins contain from 1 to more than 30 figures. Based on the number and the pattern of the fingers, most of the proteins can be classified into one of three groups: triple-C2H2, multiple-adjacent-C2H2, and separated-paired-C2H2 finger proteins. In contrast to proteins with triple-C2H2 fingers, proteins with multiple-adjacent-C2H2 fingers can bind multiple, different ligands. Proteins with a number of separated-paired fingers bind to the target by means of only a single pair.

Histone acetylation and disease

Abstract: Differential acetylation of histones and transcription factors plays an important regulatory role in developmental processes, proliferation and differentiation. Aberrant acetylation or deacetylation leads to such diverse disorders as leukemia, epithelial cancers, fragile X syndrome and Rubinstein-Taybi syndrome. The various groups of histone acetyltransferases (CBP/p300, GNAT, MYST, nuclear receptor coactivators and TAFII250) and histone deacetylases are surveyed with regard to their possible or known involvement in cancer progression and human developmental disorders. Current treatment strategies are discussed, which are still mostly limited to histone deacetylase inhibitors such as trichostatin A and butyrate.

Neurotrophin signalling pathways regulating neuronal apoptosis.

Abstract: Recent evidence indicates that naturally occurring neuronal death in mammals is regulated by the interplay between receptor-mediated prosurvival and proapoptotic signals. The neurotrophins, a family of growth factors best known for their positive effects on neuronal biology, have now been shown to mediate both positive and negative survival signals, by signalling through the Trk and p75 neurotrophin receptors, respectively. The mechanisms whereby these two neurotrophin receptors interact to determine neuronal survival have been difficult to decipher, largely because both can signal independently or coincidentally, depending upon the cell or developmental context. Nonetheless, the past several years have seen significant advances in our understanding of this receptor signalling system. In this review, we focus on the proapoptotic actions of the p75 neurotrophin receptor (p75NTR), and on the interplay between Trk and p75NTR that determines neuronal survival.

Apoptotic and necrotic cell death induced by death domain receptors.

Abstract: Apoptosis and necrosis are two distinct forms of cell death. Caspases are indispensable as initiators and effectors of apoptotic cell death and are involved in many of the morphological and biochemical features of apoptosis. Major changes in mitochondrial membrane integrity and release of proapoptotic factors, such as cytochrome c from the mitochondrial intermembrane space, play an important sensor and amplifying role during apoptotic cell death. In vitro studies of cell death in cell lines have revealed that inhibition of the classical caspase-dependent apoptotic pathway leads in several cases to necrotic cell death. Thus, the same cell death stimulus can result either in apoptotic or necrotic cell death, depending on the availability of activated caspase. Therefore, death domain receptors may initiate an active caspase-independent necrotic signaling pathway. In this review, we describe what is known about the apoptotic and necrotic cell death pathways. Principal elements of necrosis include mitochondrial oxidative phosphorylation, reactive oxygen production, and non-caspase proteolytic cascades depending on serine proteases, calpains, or cathepsins.

Sphingolipids in mammalian cell signalling.

Abstract: Sphingolipids and their metabolites, ceramide, sphingosine and sphingosine-1-phosphate, are involved in a variety of cellular processes including differentiation, cellular senescence, apoptosis and proliferation Ceramide is the main second messenger, and is produced by sphingomyelinase-induced hydrolysis of sphingomyelin and by de novo synthesis Many stimuli, eg growth factors, cytokines, G protein-coupled receptor agonists and stress (UV irradiation) increase cellular ceramide levels Sphingomyelin in the plasma membrane is located primarily in the outer (extracellular) leaflet of the bilayer, whilst sphingomyelinases are found at the inner (cytosolic) face and within lysosomes/endosomes Such cellular compartmentalisation restricts the site of ceramide production and subsequent interaction with target proteins Glycosphingolipids and sphingomyelin together with cholesterol are major components of specialised membrane microdomains known as lipid rafts, which are involved in receptor aggregation and immune responses Many signalling molecules, for example Src family tyrosine kinases and glycosylinositolphosphate-anchored proteins, are associated with rafts, and disruption of these domains affects cellular responses such as apoptosis Sphingosine and sphingosine-1-phosphate derived from ceramide are also signalling molecules In particular, sphingosine-1-phosphate is involved in proliferation, differentiation and apoptosis Sphingosine-1-phosphate can act both extracellularly through endothelial-differentiating gene (EDG) family G protein-coupled receptors and intracellularly through direct interactions with target proteins The importance of sphingolipid signalling in cardiovascular development has been reinforced by recent reports implicating EDG receptors in the regulation of embryonic cardiac and vascular morphogenesis

The suppressors of cytokine signalling (SOCS).

Abstract: Members of the SOCS (suppressor of cytokine signalling) family of proteins play key roles in the negative regulation of cytokine signal transduction. A series of elegant biochemical and molecular biological studies has revealed that these proteins act in a negative feedback loop, inhibiting the cytokine-activated Janus kinase/signal transducers and activators of transcription (JAK/ STAT) signalling pathway to modulate cellular responses. Although structurally related, the precise mechanisms of SOCS-1, SOCS-3 and cytokine-inducible SH2-containing protein (CIS) action vary. Direct interaction of SOCS SH2 domains with the JAK kinases or cytokine receptors allows their recruitment to the signalling complex, where they inhibit JAK catalytic activity or block access of the STATs to receptor binding sites. The defining feature of the family, the C-terminal SOCS box domain, appears dispensable for these actions but is likely to play a key role in negative regulation of signalling by targeting molecules associated with the SOCS proteins for degradation. The relevance of SOCS-mediated regulation of cytokine responses has been brought into sharp focus by the dramatic phenotypes of mice lacking these regulators. Indispensable roles for members of this family have been identified in the regulation of interferon gamma, growth hormone and erythropoietin, and the absence of SOCS-1 or SOCS-3 is lethal in mice.

The role of mammalian antimicrobial peptides and proteins in awakening of innate host defenses and adaptive immunity.

Abstract: Since we live in a dirty environment, we have developed many host defenses to contend with microorganisms. The epithelial lining of our skin, gastrointestinal tract and bronchial tree produces a number of antibacterial peptides, and our phagocytic neutrophils rapidly ingest and enzymatically degrade invading organisms, as well as produce peptides and enzymes with antimicrobial activities. Some of these antimicrobial moieties also appear to alert host cells involved in both innate host defense and adaptive immune responses. The epithelial cells are a source of constitutively produced beta defensin (HBD1) and proinflammatory cytokine-inducible beta defensins (HBD2 and -3) and cathelicidin (LL37). The neutrophils-derived antimicrobial peptides are released on demand from their cytoplasmic granules. They include the enzymes cathepsin G and chymase, azurocidin, a defensins and cathelicidin. In contrast, C5a and C3b are produced by activation of the serum complement cascade. The antimicrobial moieties direct the migration and activate target cells by interacting with selected G-protein-coupled seven-transmembrane receptors (GPCRs) on cell surfaces. The beta defensins interact with the CCR6 chemokine GPCRs, whereas cathelicidins interact with the low-affinity FPRL-1 receptors. The neutrophil-derived cathepsin G acts on the high-affinity FMLP receptor (GPCR) known as FPR, while the receptors for chymase and azurocidin have not been identified as yet. The serum-derived C5a uses a GPCR known as C5aR to mediate its chemotactic and cell-activating effects. Consequently, all these ligand-receptor interactions in addition to mediating chemotaxis also activate receptor-expressing cells to produce other mediators of inflammation.

Transthyretin: a review from a structural perspective.

Abstract: Transthyretin (formerly called prealbumin) plays important physiological roles as a transporter of thyroxine and retinol-binding protein. X-ray structural studies have provided information on the active conformation of the protein and the site of binding of both ligands. Transthyretin is also one of the precursor proteins commonly found in amyloid deposits. Both wild-type and single-amino-acid-substituted variants have been identified in amyloid deposits, the variants being more amyloidogenic. Sequencing of the gene and the resulting production of a transgenic mouse model have resulted in progress toward solving the mechanism of amyloid formation and detecting the variant gene in individuals at risk.

Crosstalk between cell cycle regulators and the myogenic factor MyoD in skeletal myoblasts.

Abstract: During the early process of skeletal muscle differentiation, myogenic factors are not only involved in muscle-specific gene induction but also in regulating the transition from the proliferative stage, when MyoD and Myf5 are already expressed, to the orderly exit from the cell division cycle. This key step in skeletal muscle differentiation involves the down-regulation of cell cycle activators such as cyclins and cdks, and up-regulation of cell cycle inhibitors such as Rb, p21, p27, and p57. In particular, Rb and p21 have been shown to play an important role in the growth arrest of differentiating myoblasts. Their level and/or activity, while being negatively controlled by growth factors, appear to be positively linked with the myogenic factor MyoD, which plays a cooperative role in the induction of growth arrest. MyoD can block proliferation independently of its transcriptional activity. Therefore, the interplay between G1 cyclins and cdk inhibitors, on the one hand, and MyoD and its cofactors, on the other, plays a critical role in myoblast cell cycle withdrawal. Accurate synchronization of dividing myoblasts revealed that MyoD and Myf5 are themselves subject to specific cell cycle-dependent regulation, with MyoD at its highest level in early G1 and its lowest level at the G1 to S phase transition. The time-window when cells exit their cycle into differentiation is in G1, when MyoD is maximal and Myf5 is down. In contrast, quiescent non-differentiating myoblasts (i.e., in G0) present an opposite pattern for the two factors: high Myf5 and no MyoD. Several recent studies have focused on MyoD phosphorylation and its potential role in ubiquitination-mediated degradation of the protein. Linking this phosphorylation to the cell cycle-dependent drop in MyoD protein before S phase leads, to a mechanism implying cdk2-cyclin E and its inhibitors (p57kip and p21cip) in the tight control of MyoD levels and subsequent myoblast cell cycle progression or exit into differentiation.

The GPI-anchor and protein sorting.

Abstract: Glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) are a diverse class of proteins that are anchored to the membrane solely via means of a post-translational lipid modification, the GPI-moiety. Since their discovery in the late 1970s, years of research have provided significant insight into the functions of this ubiquitous modification. In addition to the structure and biosynthesis of the GPI-moiety, perhaps the best-studied feature of this glycolipid is its ability to impart characteristic membrane-trafficking properties to the proteins that it anchors. Study of the mechanism of sorting of GPI-APs has brought to light the importance of lateral heterogeneities in cell membranes, termed rafts, in biological sorting processes. The focus of this review is to examine the emerging role of the GPI-anchor and mechanisms involved in GPI-AP sorting in the context of intracellular trafficking pathways.

The presynaptic cytomatrix of brain synapses.

Abstract: Synapses are principal sites for communication between neurons via chemical messengers called neurotransmitters. Neurotransmitters are released from presynaptic nerve terminals at the active zone, a restricted area of the cell membrane situated exactly opposite to the postsynaptic neurotransmitter reception apparatus. At the active zone neurotransmitter-containing synaptic vesicles (SVs) dock, fuse, release their content and are recycled in a strictly regulated manner. The cytoskeletal matrix at the active zone (CAZ) is thought to play an essential role in the organization of this SV cycle. Several multi-domain cytoskeleton-associated proteins, including RIM, Bassoon, Piccolo/Aczonin and Munc-13, have been identified, which are specifically localized at the active zone and thus are putative molecular components of the CAZ. This review will summarize our present knowledge about the structure and function of these CAZ-specific proteins. Moreover, we will review our present view of how the exocytotic and endocytic machineries at the site of neurotransmitter release are linked to and organized by the presynaptic cytoskeleton. Finally, we will summarize recent progress that has been made in understanding how active zones are assembled during nervous system development.

Protein oxidation and 20S proteasome-dependent proteolysis in mammalian cells.

Abstract: The generation of reactive oxygen species is an inevitable aspect of aerobic life. In addition to being exposed to free radicals in the environment, aerobic organisms must also deal with oxygen radicals generated as byproducts of a number of physiological mechanisms for example, by the mitochondrial and endoplasmic reticulum electron transport chains, and by cells of the immune system. Although most organisms are equipped with several lines of defense against oxidative stress, these defensive mechanisms are not 100% effective, and oxidatively modified forms of proteins accumulate during aging, and in many pathological conditions. Oxidatively modified proteins can form large aggregates due to covalent cross-linking or increased surface hydrophobicity. Unless repaired or removed from cells, these oxidized proteins are often toxic and can threaten cell viability. Mammalian cells exhibit only limited direct repair mechanisms, and oxidatively damaged proteins appear to undergo selective proteolysis, primarily by the major cytosolic proteinase, the proteasome. Interestingly, it appears that the 20S 'core' proteasome conducts the recognition and elimination of oxidized proteins in an ATP-independent and ubiquitin-independent pathway.

Trans-Golgi network sorting.

Abstract: The trans-Golgi network (TGN) is a major secretory pathway sorting station that directs newly synthesized proteins to different subcellular destinations. The TGN also receives extracellular materials and recycled molecules from endocytic compartments. In this review, we summarize recent progress on understanding TGN structure and the dynamics of trafficking to and from this compartment. Protein sorting into different transport vesicles requires specific interactions between sorting motifs on the cargo molecules and vesicle coat components that recognize these motifs. Current understanding of the various targeting signals and vesicle coat components that are involved in TGN sorting are discussed, as well as the molecules that participate in retrieval to this compartment in both yeast and mammalian cells. Besides proteins, lipids and lipid-modifying enzymes also participate actively in the formation of secretory vesicles. The possible mechanisms of action of these lipid hydrolases and lipid kinases are discussed. Finally, we summarize the fundamentally different apical and basolateral cell surface delivery mechanisms and the current facts and hypotheses on protein sorting from the TGN into the regulated secretory pathway in neuroendocrine cells.

Emerging connections between DNA methylation and histone acetylation

Abstract: Modifications of both DNA and chromatin can affect gene expression and lead to gene silencing. Evidence of links between DNA methylation and histone hypoacetylation is accumulating. Several proteins that specifically bind to methylated DNA are associated with complexes that include histone deacetylases (HDACs). In addition, DNA methyltransferases of mammals appear to interact with HDACs. Experiments with animal cells have shown that HDACs are responsible for part of the repressive effect of DNA methylation. Evidence was found in Neurospora that protein acetylation can in some cases affect DNA methylation. The available data suggest that the roles of DNA methylation and histone hypoacetylation, and their relationship with each other, can vary, even within an organism. Some open questions in this emerging field that should be answered in the near future are discussed.

Signalling roles of mammalian phospholipase D1 and D2.

Abstract: Phospholipase D (PLD) catalyses the hydrolysis of phosphatidylcholine to generate the lipid second messenger, phosphatidate (PA) and choline. PLD activity in mammalian cells is low and is transiently stimulated upon activation by G-protein-coupled and receptor tyrosine kinase cell surface receptors. Two mammalian PLD enzymes (PLD1 and PLD2) have been cloned and their intracellular regulators identified as ARF and Rho proteins, protein kinase Calpha as well as the lipid, phosphatidylinositol bisphosphate (PIP2). I discuss the regulation of these enzymes by cell surface receptors, their cellular localisation and the potential function of PA as a second messenger. Evidence is presented for a role of PA in regulating the lipid kinase activity of PIP 5-kinase, an enzyme that synthesises PIP2. A signalling role of phospholipase D via PA and indirectly via PIP2 in regulating membrane traffic and actin dynamics is indicated by the available data.

Glycine-rich proteins as structural components of plant cell walls.

Abstract: Glycine-rich proteins (GRPs) have been found in the cell walls of many higher plants and form a third group of structural protein components of the wall in addition to extensins and proline-rich proteins. The primary sequences of GRPs contain more than 60% glycine. GRPs are localized mainly in the vascular tissue of the plant, and their coding genes provide an excellent system to analyze the molecular basis of vascular-specific gene expression. In French bean, the major cell wall GRP has been localized at the ultrastructural level in the modified primary cell wall of protoxylem. Immunological studies showed that it forms a major part of these highly extensible and specialized cell walls. Specific digestion of GRP1.8 from bean by collagenase suggests that it shares structural similarities with collagen. The protein is synthesized by living protoxylem cells as well as xylem parenchyma cells. After cell death, GRPs are exported from neighboring xylem parenchyma cells to the protoxylem wall, a rare example of protein transport between cells in plants. We propose that GRPs are part of a repair system of the plant during the stretching phase of protoxylem.