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991.
The renewal and differentiation of Isl1+ cardiovascular progenitors are controlled by a Wnt/beta-catenin pathway 总被引:3,自引:0,他引:3
Qyang Y Martin-Puig S Chiravuri M Chen S Xu H Bu L Jiang X Lin L Granger A Moretti A Caron L Wu X Clarke J Taketo MM Laugwitz KL Moon RT Gruber P Evans SM Ding S Chien KR 《Cell Stem Cell》2007,1(2):165-179
Isl1(+) cardiovascular progenitors and their downstream progeny play a pivotal role in cardiogenesis and lineage diversification of the heart. The mechanisms that control their renewal and differentiation are largely unknown. Herein, we show that the Wnt/beta-catenin pathway is a major component by which cardiac mesenchymal cells modulate the prespecification, renewal, and differentiation of isl1(+) cardiovascular progenitors. This microenvironment can be reconstituted by a Wnt3a-secreting feeder layer with ES cell-derived, embryonic, and postnatal isl1(+) cardiovascular progenitors. In vivo activation of beta-catenin signaling in isl1(+) progenitors of the secondary heart field leads to their massive accumulation, inhibition of differentiation, and outflow tract (OFT) morphogenic defects. In addition, the mitosis rate in OFT myocytes is significantly reduced following beta-catenin deletion in isl1(+) precursors. Agents that manipulate Wnt signals can markedly expand isl1(+) progenitors from human neonatal hearts, a key advance toward the cloning of human isl1(+) heart progenitors. 相似文献
992.
Eshete M Marchbank MT Deutscher SL Sproat B Leszczynska G Malkiewicz A Agris PF 《The protein journal》2007,26(1):61-73
Protein recognition of RNA has been studied using Peptide Phage Display Libraries, but in the absence of RNA modifications.
Peptides from two libraries, selected for binding the modified anticodon stem and loop (ASL) of human tRNALys3 having 2-thiouridine (s2U34) and pseudouridine (Ψ39), bound the modified human ASLLys3(s2U34;Ψ39) preferentially and had significant homology with RNA binding proteins. Selected peptides were narrowed to a manageable number
using a less sensitive, but inexpensive assay before conducting intensive characterization. The affinity and specificity of
the best binding peptide (with an N-terminal fluorescein) were characterized by fluorescence spectrophotometry. The peptide
exhibited the highest binding affinity for ASLLys3(s2U34;Ψ39), followed by the hypermodified ASLLys3 (mcm5s2U34;ms2t6A37) and the unmodified ASLLys3, but bound poorly to singly modified ASLLys3 constructs (Ψ39, ms2t6A37,
s2U34), ASLLys1,2
(t6A37) and Escherichia coli ASLGlu (s2U34). Thus, RNA modifications are potentially important recognition elements for proteins and can be targets for selective recognition
by peptides. 相似文献
993.
Deletion of complement factor H-related genes CFHR1 and CFHR3 is associated with atypical hemolytic uremic syndrome 下载免费PDF全文
Zipfel PF Edey M Heinen S Józsi M Richter H Misselwitz J Hoppe B Routledge D Strain L Hughes AE Goodship JA Licht C Goodship TH Skerka C 《PLoS genetics》2007,3(3):e41
Atypical hemolytic uremic syndrome (aHUS) is associated with defective complement regulation. Disease-associated mutations have been described in the genes encoding the complement regulators complement factor H, membrane cofactor protein, factor B, and factor I. In this study, we show in two independent cohorts of aHUS patients that deletion of two closely related genes, complement factor H-related 1 (CFHR1) and complement factor H-related 3 (CFHR3), increases the risk of aHUS. Amplification analysis and sequencing of genomic DNA of three affected individuals revealed a chromosomal deletion of approximately 84 kb in the RCA gene cluster, resulting in loss of the genes coding for CFHR1 and CFHR3, but leaving the genomic structure of factor H intact. The CFHR1 and CFHR3 genes are flanked by long homologous repeats with long interspersed nuclear elements (retrotransposons) and we suggest that nonallelic homologous recombination between these repeats results in the loss of the two genes. Impaired protection of erythrocytes from complement activation is observed in the serum of aHUS patients deficient in CFHR1 and CFHR3, thus suggesting a regulatory role for CFHR1 and CFHR3 in complement activation. The identification of CFHR1/CFHR3 deficiency in aHUS patients may lead to the design of new diagnostic approaches, such as enhanced testing for these genes. 相似文献
994.
Marko-Varga G Ogiwara A Nishimura T Kawamura T Fujii K Kawakami T Kyono Y Tu HK Anyoji H Kanazawa M Akimoto S Hirano T Tsuboi M Nishio K Hada S Jiang H Fukuoka M Nakata K Nishiwaki Y Kunito H Peers IS Harbron CG South MC Higenbottam T Nyberg F Kudoh S Kato H 《Journal of proteome research》2007,6(8):2925-2935
Personalized medicine allows the selection of treatments best suited to an individual patient and disease phenotype. To implement personalized medicine, effective tests predictive of response to treatment or susceptibility to adverse events are needed, and to develop a personalized medicine test, both high quality samples and reliable data are required. We review key features of state-of-the-art proteomic profiling and introduce further analytic developments to build a proteomic toolkit for use in personalized medicine approaches. The combination of novel analytical approaches in proteomic data generation, alignment and comparison permit translation of identified biomarkers into practical assays. We further propose an expanded statistical analysis to understand the sources of variability between individuals in terms of both protein expression and clinical variables and utilize this understanding in a predictive test. 相似文献
995.
Bouyssié D Gonzalez de Peredo A Mouton E Albigot R Roussel L Ortega N Cayrol C Burlet-Schiltz O Girard JP Monsarrat B 《Molecular & cellular proteomics : MCP》2007,6(9):1621-1637
Proteomics strategies based on nanoflow (nano-) LC-MS/MS allow the identification of hundreds to thousands of proteins in complex mixtures. When combined with protein isotopic labeling, quantitative comparison of the proteome from different samples can be achieved using these approaches. However, bioinformatics analysis of the data remains a bottleneck in large scale quantitative proteomics studies. Here we present a new software named Mascot File Parsing and Quantification (MFPaQ) that easily processes the results of the Mascot search engine and performs protein quantification in the case of isotopic labeling experiments using either the ICAT or SILAC (stable isotope labeling with amino acids in cell culture) method. This new tool provides a convenient interface to retrieve Mascot protein lists; sort them according to Mascot scoring or to user-defined criteria based on the number, the score, and the rank of identified peptides; and to validate the results. Moreover the software extracts quantitative data from raw files obtained by nano-LC-MS/MS, calculates peptide ratios, and generates a non-redundant list of proteins identified in a multisearch experiment with their calculated averaged and normalized ratio. Here we apply this software to the proteomics analysis of membrane proteins from primary human endothelial cells (ECs), a cell type involved in many physiological and pathological processes including chronic inflammatory diseases such as rheumatoid arthritis. We analyzed the EC membrane proteome and set up methods for quantitative analysis of this proteome by ICAT labeling. EC microsomal proteins were fractionated and analyzed by nano-LC-MS/MS, and database searches were performed with Mascot. Data validation and clustering of proteins were performed with MFPaQ, which allowed identification of more than 600 unique proteins. The software was also successfully used in a quantitative differential proteomics analysis of the EC membrane proteome after stimulation with a combination of proinflammatory mediators (tumor necrosis factor-alpha, interferon-gamma, and lymphotoxin alpha/beta) that resulted in the identification of a full spectrum of EC membrane proteins regulated by inflammation. 相似文献
996.
Van Leene J Stals H Eeckhout D Persiau G Van De Slijke E Van Isterdael G De Clercq A Bonnet E Laukens K Remmerie N Henderickx K De Vijlder T Abdelkrim A Pharazyn A Van Onckelen H Inzé D Witters E De Jaeger G 《Molecular & cellular proteomics : MCP》2007,6(7):1226-1238
Defining protein complexes is critical to virtually all aspects of cell biology because many cellular processes are regulated by stable protein complexes, and their identification often provides insights into their function. We describe the development and application of a high throughput tandem affinity purification/mass spectrometry platform for cell suspension cultures to analyze cell cycle-related protein complexes in Arabidopsis thaliana. Elucidation of this protein-protein interaction network is essential to fully understand the functional differences between the highly redundant cyclin-dependent kinase/cyclin modules, which are generally accepted to play a central role in cell cycle control, in all eukaryotes. Cell suspension cultures were chosen because they provide an unlimited supply of protein extracts of actively dividing and undifferentiated cells, which is crucial for a systematic study of the cell cycle interactome in the absence of plant development. Here we report the mapping of a protein interaction network around six known core cell cycle proteins by an integrated approach comprising generic Gateway-based vectors with high cloning flexibility, the fast generation of transgenic suspension cultures, tandem affinity purification adapted for plant cells, matrix-assisted laser desorption ionization tandem mass spectrometry, data analysis, and functional assays. We identified 28 new molecular associations and confirmed 14 previously described interactions. This systemic approach provides new insights into the basic cell cycle control mechanisms and is generally applicable to other pathways in plants. 相似文献
997.
Marmagne A Ferro M Meinnel T Bruley C Kuhn L Garin J Barbier-Brygoo H Ephritikhine G 《Molecular & cellular proteomics : MCP》2007,6(11):1980-1996
The proteomics of plasma membrane has brought to date only scarce and partial information on the actual protein repertoire. In this work, the plant plasma membrane proteome of Arabidopsis thaliana was investigated. A highly purified plasma membrane fraction was washed by NaCl and Na2CO3 salts, and the insoluble fractions were further analyzed by nano-LC-MS/MS. With 446 proteins identified, we hereby describe the largest plasma membrane proteome diversity reported so far. Half of the proteins were predicted to display transmembrane domains and/or to be anchored to the membrane, validating a posteriori the pertinence of the approach. A fine analysis highlighted two main specific and novel features. First, the main functional category is represented by a majority of as yet unreported signaling proteins, including 11% receptor-like kinases. Second, 16% of the identified proteins are predicted to be lipid-modified, specifically involving double lipid linkage through N-terminal myristoylation, S-palmitoylation, C-terminal prenylation, or glycosylphosphatidylinositol anchors. Thus, our approach led for the first time to the identification of a large number of peripheral proteins as part of the plasma membrane and allowed the functionality of the plasma membrane in the cell context to be reconsidered. 相似文献
998.
Despite recent advances in developmental biology, and the sequencing and annotation of genomes, key questions regarding the organisation of cells into embryos remain. One possibility is that uncharacterised genes having nonstandard coding arrangements and functions could provide some of the answers. Here we present the characterisation of tarsal-less (tal), a new type of noncanonical gene that had been previously classified as a putative noncoding RNA. We show that tal controls gene expression and tissue folding in Drosophila, thus acting as a link between patterning and morphogenesis. tal function is mediated by several 33-nucleotide-long open reading frames (ORFs), which are translated into 11-amino-acid-long peptides. These are the shortest functional ORFs described to date, and therefore tal defines two novel paradigms in eukaryotic coding genes: the existence of short, unprocessed peptides with key biological functions, and their arrangement in polycistronic messengers. Our discovery of tal-related short ORFs in other species defines an ancient and noncanonical gene family in metazoans that represents a new class of eukaryotic genes. Our results open a new avenue for the annotation and functional analysis of genes and sequenced genomes, in which thousands of short ORFs are still uncharacterised. 相似文献
999.
Targeted inhibition of miRNA maturation with morpholinos reveals a role for miR-375 in pancreatic islet development 总被引:5,自引:0,他引:5 下载免费PDF全文
Several vertebrate microRNAs (miRNAs) have been implicated in cellular processes such as muscle differentiation, synapse function, and insulin secretion. In addition, analysis of Dicer null mutants has shown that miRNAs play a role in tissue morphogenesis. Nonetheless, only a few loss-of-function phenotypes for individual miRNAs have been described to date. Here, we introduce a quick and versatile method to interfere with miRNA function during zebrafish embryonic development. Morpholino oligonucleotides targeting the mature miRNA or the miRNA precursor specifically and temporally knock down miRNAs. Morpholinos can block processing of the primary miRNA (pri-miRNA) or the pre-miRNA, and they can inhibit the activity of the mature miRNA. We used this strategy to knock down 13 miRNAs conserved between zebrafish and mammals. For most miRNAs, this does not result in visible defects, but knockdown of miR-375 causes defects in the morphology of the pancreatic islet. Although the islet is still intact at 24 hours postfertilization, in later stages the islet cells become scattered. This phenotype can be recapitulated by independent control morpholinos targeting other sequences in the miR-375 precursor, excluding off-target effects as cause of the phenotype. The aberrant formation of the endocrine pancreas, caused by miR-375 knockdown, is one of the first loss-of-function phenotypes for an individual miRNA in vertebrate development. The miRNA knockdown strategy presented here will be widely used to unravel miRNA function in zebrafish. 相似文献
1000.
Hughes AE Orr N Patterson C Esfandiary H Hogg R McConnell V Silvestri G Chakravarthy U 《PLoS medicine》2007,4(12):e355