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Distinct Proteins (distinct + protein)
Selected AbstractsPinocchio, a novel protein expressed in the antenna, contributes to olfactory behavior in Drosophila melanogasterDEVELOPMENTAL NEUROBIOLOGY, Issue 2 2005Stephanie M. Rollmann Abstract Most organisms depend on chemoreception for survival and reproduction. In Drosophila melanogaster multigene families of chemosensory receptors and putative odorant binding proteins have been identified. Here, we introduce an additional distinct protein, encoded by the CG4710 gene, that contributes to olfactory behavior. Previously, we identified through P[lArB] -element mutagenesis a smell impaired (smi) mutant, smi21F, with odorant-specific defects in avoidance responses. Here, we show that the smi21F mutant also exhibits reduced attractant responses to some, but not all, of a select group of odorants. Furthermore, electroantennogram amplitudes are increased in smi21F flies. Characterization of flanking sequences of the P[lArB] insertion site, complementation mapping, phenotypic reversion through P -element excision, and expression analysis implicate a predicted gene, CG4710, as the candidate smi gene. CG4710 produces two transcripts that encode proteins that contain conserved cysteines and which are reduced in the smi21F mutant. Furthermore, in situ hybridization reveals CG4710 expression in the third antennal segment. We have named this gene of previously unknown function and its product "Pinocchio (Pino)". © 2005 Wiley Periodicals, Inc. J Neurobiol., 2005 [source] Co-option of endocytic functions of cellular caveolae by pathogensIMMUNOLOGY, Issue 1 2001J.-S. Shin Summary It is increasingly becoming clear that various immune cells are infected by the very pathogens that they are supposed to attack. Although many mechanisms for microbial entry exist, it appears that a common route of entry shared by certain bacteria, viruses and parasites involves cellular lipid-rich microdomains sometimes called caveolae. These cellular entities, which are characterized by their preferential accumulation of glycosylphosphatidylinositol (GPI)-anchored molecules, cholesterol and various glycolipids, and a distinct protein (caveolin), are present in many effector cells of the immune system including neutrophils, macrophages, mast cells and dendritic cells. These structures have an innate capacity to endocytoze various ligands and traffic them to different intracellular sites and sometimes, back to the extracellular cell surface. Because caveolae do not typically fuse with lysosomes, the ligands borne by caveolar vesicles are essentially intact, which is in marked contrast to ligands endocytozed via the classical endosome,lysosome pathway. A number of microbes or their exotoxins co-opt the unique features of caveolae to enter and traffic, without any apparent loss of viability and function, to different sites within immune and other host cells. In spite of their wide disparity in size and other structural attributes, we predict that a common feature among caveolae-utilizing pathogens and toxins is that their cognate receptor(s) are localized within plasmalemmal caveolae of the host cell. [source] Characterization of Vorticella convallaria calcium-binding centrin proteinsTHE JOURNAL OF EUKARYOTIC MICROBIOLOGY, Issue 2 2005KATARZYNA KONIOR The stalked ciliate, Vorticella convallaria, is a good model system to study mechanochemical motility because its contractile organelles (spasmoneme and myonemes) use a mode of contraction that differs from most other eukaryotic motile systems. Since calcium triggers this contraction, we have undertaken the molecular characterization of the calcium-binding proteins associated with these organelles. We have isolated and identified seven unique centrin-like cDNAs from V. convallaria. Each encodes an acidic protein of approximately 20-kDa, containing a unique N-terminus and four potential calcium-binding domains. We predict that each centrin has a distinct function within the cell. To define these functions, we have initiated immunofluorescence localization studies utilizing various anti-centrin antibodies. Western analysis indicates that each antibody recognizes a distinct protein or subset of proteins in Vorticella. Using these antibodies, we have localized centrin to various structures within the cell; myonemes, spasmoneme, and the oral apparatus. Because each of these antibodies recognizes a different protein on Westerern analysis, we conclude that a number of calcium-binding proteins are associated with the contractile organelles. To further characterize this gene family, we have initiated immunolocalization at the ultrastructural level. This will permit subcellular localization of all Vorticella centrins and enable us to dissect the function of this multi-gene family. [source] Phytanic Acid Accumulation Is Associated with Conduction Delay and Sudden Cardiac Death in Sterol Carrier Protein-2/Sterol Carrier Protein-x Deficient MiceJOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 11 2004GEROLD MÖNNIG M.D. Introduction: The sterol carrier protein-2 gene encodes two functionally distinct proteins: sterol carrier protein-2 (SCP2, a peroxisomal lipid carrier) and sterol carrier protein-x (SCPx, a peroxisomal thiolase known as peroxisomal thiolase-2), which is involved in peroxisomal metabolism of bile acids and branched-chain fatty acids. We show in this study that mice deficient in SCP2 and SCPx (SCP2null) develop a cardiac phenotype leading to a high sudden cardiac death rate if mice are maintained on diets enriched for phytol (a metabolic precursor of branched-chain fatty acids). Methods and Results: In 210 surface and 305 telemetric ECGs recorded in wild-type (C57BL/6; wt; n = 40) and SCP2 null mice (n = 40), no difference was observed at baseline. However, on diet, cycle lengths were prolonged in SCP2 null mice (262.9 ± 190 vs 146.3 ± 43 msec), AV conduction was prolonged (58.3 ± 17 vs 42.6 ± 4 ms), and QRS complexes were wider (19.1 ± 5 vs 14.0 ± 4 ms). In 11 gene-targeted Langendorff-perfused hearts isolated from SCP2 null mice after dietary challenge, complete AV blocks (n = 5/11) or impaired AV conduction (Wenckebach point 132 ± 27 vs 92 ± 10 msec; P < 0.05) could be confirmed. Monophasic action potentials were not different between the two genotypes. Left ventricular function studied by echocardiography was similar in both strains. Phytanic acid but not pristanic acid accumulated in the phospholipid fraction of myocardial membranes isolated from SCP2 null mice. Conclusion: Accumulation of phytanic acid in myocardial phospholipid membranes is associated with bradycardia and impaired AV nodal and intraventricular impulse conduction, which could provide an explanation for sudden cardiac death in this model. [source] Basal and stimulated lactate fluxes in primary cultures of astrocytes are differentially controlled by distinct proteinsJOURNAL OF NEUROCHEMISTRY, Issue 3 2008Fumihiko Maekawa Abstract Lactate release by astrocytes is postulated to be of importance for neuroenergetics but its regulation is poorly understood. Basigin, a chaperone protein for specific monocarboxylate transporters (MCTs), represents a putatively important regulatory element for lactate fluxes. Indeed, basigin knockdown by RNA interference in primary cultures of astrocytes partially reduced both proton-driven lactate influx and efflux. But more strikingly, enhancement of lactate efflux induced by glutamate was prevented while the effect of sodium azide was significantly reduced by treatment of cultured astrocytes with anti-basigin small interfering RNA. Enhancement of glucose utilization was unaffected under the same conditions. Basal lactate uptake and release were significantly reduced by MCT1 knockdown, even more so than with basigin knockdown, whereas glutamate-driven or sodium azide-induced enhancement of lactate release was not inhibited by either MCT1, 2, or 4 small interfering RNAs. In conclusion, MCT1 plays a pivotal role in the control of basal proton-driven lactate flux in astrocytes while basigin is only partly involved, most likely via its interaction with MCT1. In contrast, basigin appears to critically regulate the enhancement of lactate release caused by glutamate (or sodium azide) but via an effect on another unidentified transporter at least present in astrocytes in vitro. [source] Mitochondrial gene diversity in the common vole Microtus arvalis shaped by historical divergence and local adaptationsMOLECULAR ECOLOGY, Issue 11 2004SABINE FINK Abstract The phylogeography of the common vole (Microtus arvalis) was examined by analysing mitochondrial DNA (mtDNA) sequence variation in 1044 base pairs (bp) of the cytochrome b (cytb) gene and in 322 bp of the control region (ctr) among 106 individuals from 58 locations. The geographical distribution of four previously recognized cytb evolutionary lineages in Europe was refined and a new lineage was found in southern Germany. All lineages were distributed allopatrically, except in one sample that was probably located in a contact zone. The occurrence of several lineages in the Alps is in keeping with their recent recolonization from distinct sources. The translation of 84 cytb DNA sequences produced 33 distinct proteins with relationships that differed from those of the DNA haplotypes, suggesting that the mtDNA lineages did not diverge in response to selection. In comparison with M. agrestis, a neutrality test detected no overall evidence for selection in the cytb gene, but a closer examination of a structural model showed that evolutionarily conserved and functionally important positions were often affected. A new phylogeographical test of random accumulation of nonsynonymous mutations generated significant results in three lineages. We therefore conclude that the molecular diversity of cytb in M. arvalis is overall the result of the demographic history of the populations, but that there have been several episodes of local adaptation to peculiar environments. [source] Imprinting on chromosome 20: Tissue-specific imprinting and imprinting mutations in the GNAS locus,AMERICAN JOURNAL OF MEDICAL GENETICS, Issue 3 2010Gavin Kelsey Abstract The GNAS locus on chromosome 20q13.11 is the archetypal complex imprinted locus. It comprises a bewildering array of alternative transcripts determined by differentially imprinted promoters which encode distinct proteins. It also provides the classic example of tissue-specific imprinted gene expression, in which the canonical GNAS transcript coding for Gs, is expressed predominantly from the maternal allele in a set of seemingly unrelated tissues. Functionally, this rather obscure imprinting is nevertheless of considerable clinical significance, as it dictates the nature of the disease caused by inactivating mutations in Gs,, with end organ hormone resistance specifically on maternal transmission (pseudohypoparathyroidism type 1a, PHP1a). In addition, there is a bona fide imprinting disorder, PHP1b, which is caused specifically by DNA methylation defects in the differentially methylated regions (DMRs) that determine tissue-specific monoallelic expression of GNAS. Although the genetic defect in PHP1a and the disrupted imprinting in PHP1b both essentially result in profound reduction of Gs, activity in tissues with monoallelic GNAS expression, and despite a growing awareness of the overlap in these two conditions, there are important pathophysiological differences between the two whose basis is not fully understood. PHP1b is one of the only imprinted gene syndromes in which cis -acting mutations have been discovered that disrupt methylation of germline-derived imprint marks; such imprinting mutations in GNAS are helping to provide important new insights into the mechanisms of imprinting establishment generally. © 2010 Wiley-Liss, Inc. [source] Insights into the anthrax lethal factor,substrate interaction and selectivity using docking and molecular dynamics simulationsPROTEIN SCIENCE, Issue 8 2009Georgios A. Dalkas Abstract The anthrax toxin of the bacterium Bacillus anthracis consists of three distinct proteins, one of which is the anthrax lethal factor (LF). LF is a gluzincin Zn-dependent, highly specific metalloprotease with a molecular mass of ,90 kDa that cleaves most isoforms of the family of mitogen-activated protein kinase kinases (MEKs/MKKs) close to their amino termini, resulting in the inhibition of one or more signaling pathways. Previous studies on the crystal structures of uncomplexed LF and LF complexed with the substrate MEK2 or a MKK-based synthetic peptide provided structure-activity correlations and the basis for the rational design of efficient inhibitors. However, in the crystallographic structures, the substrate peptide was not properly oriented in the active site because of the absence of the catalytic zinc atom. In the current study, docking and molecular dynamics calculations were employed to examine the LF-MEK/MKK interaction along the catalytic channel up to a distance of 20 Å from the zinc atom. This residue-specific view of the enzyme-substrate interaction provides valuable information about: (i) the substrate selectivity of LF and its inactivation of MEKs/MKKs (an issue highly important not only to anthrax infection but also to the pathogenesis of cancer), and (ii) the discovery of new, previously unexploited, hot-spots of the LF catalytic channel that are important in the enzyme/substrate binding and interaction. [source] Impact of the transcriptional regulator, Ace2, on the Candida glabrata secretomePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 2 2010David A. Stead Abstract Candida glabrata is a major fungal pathogen of humans, and the virulence of C. glabrata is increased by inactivation of the transcription factor, Ace2. Our previous examination of the effects of Ace2 inactivation upon the intracellular proteome suggested that the hypervirulence of C. glabrata ace2 mutants might be caused by differences in the secretome. Therefore in this study we have characterised the C. glabrata secretome and examined the effects of Ace2 inactivation upon this extracellular proteome. We have identified 31 distinct proteins in the secretome of wild-type C. glabrata cells by MS/MS of proteins that were precipitated from the growth medium and enriched by affinity chromatography on concanavalin A. Most of these proteins are predicted to be cell wall proteins, cell wall modifying enzymes and aspartyl proteinases. The endochitinase Cts1 and the endoglucanase Egt2 were not detected in the C. glabrata secretome following Ace2 inactivation. This can account for the cell separation defect of C. glabrata ace2 cells. Ace2 inactivation also resulted in the detection of new proteins in the C. glabrata secretome. The release of such proteins might contribute to the hypervirulence of ace2 cells. [source] In vivo proteome dynamics during early bovine myogenesisPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 20 2008Thibault Chaze Abstract Myogenesis is a complex process of which the underlying mechanisms are conserved between species, including birds and mammals. Despite a good understanding of the stages of myogenesis, many of the mechanisms involved in the regulation of proliferation of the successive myoblast generations, the cellular transitions cell proliferation/alignment of myoblasts/fusion of myoblasts into myotubes/differentiation of myofibres and the control of total myofibre number still remain unknown. An in vivo proteomic analysis of the semitendinosus muscle from Charolais foetuses, at three specific stages of myogenesis (60, 110 and 180,days postconception), was conducted using 2-DE and MS. Expression profiles of more than 170 proteins were revealed and analysed using two way hierarchical clustering and statistical analysis. Our studies identify, for the first time, distinct proteins of varied biological functions and protein clusters with myogenic processes, such as the control of cell cycle activity and apoptosis, the establishment of cellular metabolism and muscle contractile properties and muscle cell reorganisation. These results are of fundamental interest to the field of myogenesis in general, and more specifically to the control of muscle development in meat producing animals. [source] Proteomic analysis of exoproteins expressed by enterotoxigenic Staphylococcus aureus strainsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 12 2008Gabriella Pocsfalvi Dr. Abstract Pathogenic bacteria excrete a variety of virulence factors into extracellular medium and to the cell surface which have essential roles in the colonization and insurrection of the host cells, and thus reflect the degree of bacterial pathogenicity. For the exploration of virulence factors expressed in the secreted proteome fraction, different Staphylococcus aureus strains were analyzed using gel-based bottom-up proteomic approach. A total of 119 distinct proteins were identified for the enterotoxin gene cluster (egc) negative and seb gene positive S. aureus American Type Culture Collection (ATCC) 14458 strain by the use of one- and 2-DE based proteomics. Detailed analysis of enterotoxin region of the 2-D map confirmed, beside the highly expressed staphylococcal enterotoxin B (SEB), the presence of enterotoxin-like proteins SElK and SElQ previously predicted by genotyping (Sergeev et al.., J. Clin. Microbiol. 2004, 42, 2134,2143). Exoprotein patterns at the late-exponential (7,h) and stationary (24,h) phases of cellular growth show a high-level similarity in this region. Comparative analysis of enterotoxin region of five S. aureus strains including two clinical isolates (RIMD 31092 and A900322), a food derived strain (AB-8802) with highly prevalent egc positive operon and a nonenterotoxigenic reference strain (ROS) revealed the presence of different known enterotoxins and other virulence factors along with a number of core exoproteins. In addition, production of SElL (RIMD 31092) and SElP (A900322) was demonstrated for the first time at the protein level. Under the experimental conditions applied none of the enterotoxins encoded by the genes of egc operon was identified. [source] Exploring snake venom proteomes: multifaceted analyses for complex toxin mixturesPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 4 2008Jay W. Fox Dr. Abstract Snake venom proteomes are complex mixtures of a large number of distinct proteins. In a sense, the field of snake venom proteomics has been under investigation since the very earliest biochemical studies on venoms where peptides and proteins were isolated and structurally and biologically characterized. With the recent developments in mass spectrometry for the identification of proteins, coupled with venom gland transcriptomes, has the field of snake venom proteomics began to flourish. These developments have led to exciting insights into the protein composition of venoms and subsequently their pathological activities. In this review, we will discuss the state of art of snake venom proteomics. Although we have not reached the ultimate goal of characterizing and quantifying all unique proteins in a venom proteome, current technologies have opened many opportunities for high-throughput proteomic studies that have gone beyond simple protein identification to analyzing various functional aspects, such as post-translational modifications, proteolytic processing and toxin-target interactions. In this review, we will discuss the technological approaches used in the study of venom proteomics highlighting the advances made and future directions. [source] Insights into human CD34+ hematopoietic stem/progenitor cells through a systematically proteomic survey coupled with transcriptomePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 9 2006Feng Liu Abstract Hematopoietic stem cells are capable of self-renewal and differentiation into different hematopoietic lineages. To gain a comprehensive understanding of hematopoietic stem/progenitor cells, a systematic proteomic survey of human CD34+ cells collected from human umbilical cord blood was performed, in which the proteins were separated by 1- and 2-DE, as well as by nano-LC, and subsequently identified by MS. A total of 370,distinct proteins identified from those cells provided new insights into the potential of the stem/progenitor cells because the nerve, gonad, and eye-associated proteins were reliably identified. Interestingly, the transcripts of 133 (35.9%) identified proteins were not found by the prevalent transcriptome approaches, although several selected transcripts could be detected by RT-PCR. Moreover, the heterogeneity of 33,proteins identified from 2-DE was attributable primarily to post-translational processes rather than to alternative splicing at transcriptional level. Furthermore, the biosyntheses of 15,proteins identified in this study appears not to be completely interrupted in spite of the fact that corresponding antisense RNAs were found in the existing transcriptome data. The integrated proteomic and transcriptomic analyses employed here provided a unique view of the human stem/progenitor cells. [source] Analysis of chicken serum proteome and differential protein expression during development in single-comb White Leghorn hensPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 7 2006San-Yuan Huang Abstract Serum is believed to harbor thousands of distinct proteins that are either actively secreted or leak from various blood cells or tissues. Exploring protein composition in serum may accelerate the discovery of novel protein biomarkers for specific economic traits in livestock species. This study analyzed serum protein composition to establish a 2-DE reference map, and monitored protein dynamics of single-comb White Leghorn hens at 8, 19 and 23,weeks after hatching. A total of 119,CBB-stained and 315,silver-stained serum protein spots were analyzed by MALDI-TOF,MS. Of these, 98,CBB-stained and 94,silver-stained protein spots were significantly matched to existing chicken proteins. The identified spots represented 30,distinctive proteins in the serum of laying hens. To compare protein expression during development, expression levels of 47,protein spots were quantified by relative spot volume with Melanie,3 software. Ten protein spots increased and 3,protein spots decreased as hen age increased. Previous research has suggested that some of these proteins play critical roles in egg production. The differentially expressed proteins with unknown identities will be valuable candidates for further explorations of their roles in egg production of laying hens. [source] Large-scale analysis of the human ubiquitin-related proteomePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 16 2005Masaki Matsumoto Abstract Protein ubiquitylation contributes to the regulation of many cellular processes including protein degradation, receptor internalization, and repair of DNA damage. We now present a comprehensive characterization of ubiquitin-conjugated and ubiquitin-associated proteins in human cells. The proteins were purified by immunoaffinity chromatography under denaturing or native conditions. They were then digested with trypsin, and the resulting peptides were analyzed by 2-D LC and MS/MS. A total of 670 distinct proteins were identified; 345 proteins (51%) were classified as Urp-D (ubiquitin-related proteome under the denaturing condition) and comprised ubiquitin-conjugated molecules, whereas 325 proteins (49%) were classified as Urp-N (ubiquitin-related proteome only under the native condition) and included molecules that associated with ubiquitylated proteins. The proportions of proteins in various functional categories differed substantially between Urp-D and Urp-N. Many ribosomal subunits were detected in the Urp-D group of proteins and several of these subunits were directly shown to be ubiquitylated by mass spectrometric analysis, suggesting that ubiquitylation might play an important role in the regulation and/or quality control of ribosomal proteins. Our results demonstrate the potential of proteomics analysis of protein ubiquitylation to provide important insight into the regulation of protein stability and other ubiquitin-related cellular functions. [source] Reference maps of mouse serum acute-phase proteins: Changes with LPS-induced inflammation and apolipoprotein,A-I and A-II transgenesPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 16 2005Robin Wait Abstract We present reference maps of the mouse serum proteome (run under reducing and non-reducing conditions), from control animals, from mice injected with lipopolysaccharide (LPS) to induce systemic inflammation, and from mice transgenic for human apolipoproteins,A-I and A-II. Seventy-seven spots/spot chains from the reducing gels were identified by HPLC MS/MS, representing 28,distinct proteins, including a species-specific protease inhibitor, contrapsin, and high levels of carboxylesterase. The concentrations of acute-phase reactants were monitored for 96,h after LPS challenge. The greatest changes (four-fold 48,h after LPS administration) were observed for haptoglobin and hemopexin. Orosomucoid/,1 -acid glycoprotein and apolipoprotein,A-I increased steadily, to 50,60% above baseline at 96,h from stimulation. In mice transgenic for human apolipoprotein,A-I the levels of expression of orosomucoid/,1 -acid glycoprotein, ,1 -macroglobulin, esterase, kininogen and contrapsin were altered compared to knockout mice lacking apolipoprotein,A-I. In contrast, except for the presence of apolipoprotein,A-II, no statistically significant difference was observed in mice transgenic for human apolipoprotein,A-II. [source] Differential protein expression in human gliomas and molecular insightsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 4 2005Vaibhav C. Chumbalkar Abstract Gliomas are the most common of the primary intracranial tumors with astrocytomas constituting about 40%. Using clinically and histologically assessed astrocytomas, we have studied their protein profiles using a two-dimensional gel electrophoresis-mass spectrometry approach and identified differentially expressed proteins which may be useful molecular indicators to understand these tumors. Examination of the protein profiles of 27,astrocytoma samples of different grades revealed 72,distinct, differentially expressed proteins belonging to various functional groups such as cytoskeleton and intermediate filament proteins, heat shock proteins (HSPs), enzymes and regulatory proteins. Based on the consistency of their differential expression, 29,distinct proteins could be short-listed and may have a role in the pathology of astrocytomas. Some were found to be differentially expressed in both Grade,III and IV astrocytomas while others were associated with a particular grade. A notable observation was underexpression of Prohibitin, a potential tumor suppressor protein, Rho-GDP dissociation inhibitor, Rho-GDI, a regulator of Rho GTPases and HSPs as well as destabilization of glial fibrillary acidic protein, GFAP, major protein of the glial filaments, in Grade,III malignant tumors. We attempt to explain glioma malignancy and progression in terms of their combined role. [source] Anatomic site-specific proteomic signatures of gastrointestinal stromal tumorsPROTEOMICS - CLINICAL APPLICATIONS, Issue 5 2009Yoshiyuki Suehara Abstract The gastrointestinal stromal tumor (GIST) is the most common mesenchymal malignancy of the gastrointestinal tract. Its clinical course ranges widely from a curable disorder to a highly malignant disease. Although its clinical and molecular characteristics depend on the anatomic site of origin, the molecular background of GIST arising in different anatomical site has not been studied yet. To investigate the proteomic background of GIST, we examined the proteomic features corresponding to the anatomic site of tumor origin. Comparison of the proteomic profile of gastric (23 cases) and small intestinal (9 cases) GIST by 2-DE revealed 105 protein spots with significantly different intensity (p <0.01) between the two groups. Mass spectrometric study identified 68 distinct proteins for these 105 protein spots, including cancer-associated ones such as prohibitin, pigment epithelium-derived factor, and alpha-actinin 4. The intensity of 37/105 (35.2%) protein spots was significantly concordant with the corresponding mRNA levels (p <0.01). Although both 2-D DIGE and microarray experiments showed significant up-regulation of vimentin expression in small intestinal GIST, Western blotting did not show a significant difference between the two groups. In conclusion, our study demonstrates the proteins specially expressed in GIST depending on their site of origin, as well as the unique advantage offered by use of proteomics to acquire such data. The identified proteins may provide clues to understanding the different characteristics of GIST depending on their site of origin. [source] Identification of differentially expressed proteins in papillary thyroid carcinomas with V600E mutation of BRAFPROTEOMICS - CLINICAL APPLICATIONS, Issue 7 2007Efisio Puxeddu Abstract BRAF, a serine/threonine kinase of the RAF family, is a downstream transducer of the RAS-regulated MAPK pathway. V600E mutation of BRAF protein is the most common genetic alteration occurring in papillary thyroid carcinomas and is prognostic of poor clinicopathological outcomes. Protein expression in the subclass of PTC bearing the BRAFV600E mutation was investigated by using 2-DE and MS/MS techniques and compared to that of matched normal thyroid tissues from seven patients. 2-D gel image analysis revealed that the expression of eight polypeptide spots, corresponding to five proteins, were significantly underexpressed in PTC bearing BRAFV600E mutation whereas 25 polypeptides, representing 19 distinct proteins, were significantly upregulated in tumour tissue, as compared to normal thyroid. Among the differentially expressed polypeptides, mitochondrial proteins, ROS-scavenger enzymes, apoptosis-related proteins as well as proteins involved in tumour cell proliferation were identified. Although dissimilarities between the present results and those previously reported can be ascribed to the use of different 2-DE techniques, the possibility that BRAFV600E mutation is responsible for changes in protein expression distinct from those induced by other oncogenes cannot be ruled out. [source] Mammalian Sperm Energy Resources Management and Survival during Conservation in RefrigerationREPRODUCTION IN DOMESTIC ANIMALS, Issue 2006JE Rodriguez-Gil Contents The present review has as its main aim to present an overview regarding the mechanisms utilized by mammalian sperm to manage its intracellular energy levels. This management will strongly influence the sperm's ability to maintain its overall function during its entire life span. Thus, the precise knowledge of these mechanisms will be of the utmost interest to optimize the systems utilized to conserve mammalian sperm for a medium-to-long time-lapse. Briefly, utilization of hexoses as energy substrates by mammalian sperm is very finely regulated from the very first step of its metabolization. Furthermore, the equilibrium among the separate, monosaccharide metabolization pathways in mammalian sperm depends on many factors. This prevents the possibility to draw a general vision of sperm energy utilization, which explains the results of all mammalian species in all points of the sperm life-cycle. To complicate the matter further, there are separate energy phenotypes among mammalian spermatozoa. The precise knowledge of these phenotypes is of the greatest importance in order to optimize the design of new extenders for sperm conservation in refrigerated conditions. Moreover, sugars can act on sperm not only as passive metabolic substrates, but also as direct function activators through mechanisms like specific changes in the tyrosine phosphorylation status of distinct proteins. Finally, mammalian sperm utilizes non-glucidic substrates like citrate and lactate to obtain energy in a regular form. This utilization is also finely regulated and of importance to maintain overall sperm function. This implies that the exact proportion of glucidic and non-glucidic energy substrates could be very important to optimize the survival ability of these cells in conservation. [source] Visualization of protein interactions in living plant cells using bimolecular fluorescence complementationTHE PLANT JOURNAL, Issue 3 2004Michael Walter Summary Dynamic networks of protein,protein interactions regulate numerous cellular processes and determine the ability to respond appropriately to environmental stimuli. However, the investigation of protein complex formation in living plant cells by methods such as fluorescence resonance energy transfer has remained experimentally difficult, time consuming and requires sophisticated technical equipment. Here, we report the implementation of a bimolecular fluorescence complementation (BiFC) technique for visualization of protein,protein interactions in plant cells. This approach relies on the formation of a fluorescent complex by two non-fluorescent fragments of the yellow fluorescent protein brought together by association of interacting proteins fused to these fragments (Hu et al., 2002). To enable BiFC analyses in plant cells, we generated different complementary sets of expression vectors, which enable protein interaction studies in transiently or stably transformed cells. These vectors were used to investigate and visualize homodimerization of the basic leucine zipper (bZIP) transcription factor bZIP63 and the zinc finger protein lesion simulating disease 1 (LSD1) from Arabidopsis as well as the dimer formation of the tobacco 14-3-3 protein T14-3c. The interaction analyses of these model proteins established the feasibility of BiFC analyses for efficient visualization of structurally distinct proteins in different cellular compartments. Our investigations revealed a remarkable signal fluorescence intensity of interacting protein complexes as well as a high reproducibility and technical simplicity of the method in different plant systems. Consequently, the BiFC approach should significantly facilitate the visualization of the subcellular sites of protein interactions under conditions that closely reflect the normal physiological environment. [source] The spliceosome: the most complex macromolecular machine in the cell?BIOESSAYS, Issue 12 2003Timothy W. Nilsen The primary transcripts, pre-mRNAs, of almost all protein-coding genes in higher eukaryotes contain multiple non-coding intervening sequences, introns, which must be precisely removed to yield translatable mRNAs. The process of intron excision, splicing, takes place in a massive ribonucleoprotein complex known as the spliceosome. Extensive studies, both genetic and biochemical, in a variety of systems have revealed that essential components of the spliceosome include five small RNAs,U1, U2, U4, U5 and U6, each of which functions as a RNA, protein complex called an snRNP (small nuclear ribonucleoprotein). In addition to snRNPs, splicing requires many non-snRNP protein factors, the exact nature and number of which has been unclear. Technical advances, including new affinity purification methods and improved mass spectrometry techniques, coupled with the completion of many genome sequences, have now permitted a number of proteomic analyses of purified spliceosomes. These studies, recently reviewed by Jurica and Moore,1 reveal that the spliceosome is composed of as many as 300 distinct proteins and five RNAs, making it among the most complex macromolecular machines known. BioEssays 25:1147,1149, 2003. © 2003 Wiley Periodicals, Inc. [source] The granin family of uniquely acidic proteins of the diffuse neuroendocrine system: comparative and functional aspectsBIOLOGICAL REVIEWS, Issue 4 2004Karen B. Helle ABSTRACT The chromogranins A (CgA) and B (CgB) and secretogranin II (SgII) constitute the main members of a family of uniquely acidic secretory proteins in elements of the diffuse neuroendocrine system. These genetically distinct proteins, CgA, CgB, SgII and the less well known secretogranins III,VII are collectively referred to as,granins'and characterised by numerous pairs of basic amino acids as potential cleavage sites for processing by the co-stored prohormone converting enzymes PC 1/3 and PC2. This review is directed towards comparative and functional aspects of the granins with emphasis on their phylogenetically conserved sequences. Recent developments provide ample evidence of widely different effects and targets for the intact granins and their derived peptides, intracellularly in the directed trafficking of storage components during granule maturation and extracellularly in autocrine, paracrine and endocrine interactions. Most of the effects assigned to the granin derived peptides fit into patterns of direct or indirect inhibitory modulations of major functions. So far, peptides derived from CgA (vasostatins, chromacin, pancreastatin, WE-14, catestatin and parastatin), CgB (secretolytin) and SgII (secretoneurin) are the most likely candidates for granin-derived regulatory peptides, of postulated relevance not only for homeostatic processes, but also for tissue assembly and repair, inflammatory responses and the first line of defence against invading microorganisms. [source] | ||||||||||||||||||||||||||||