Identification of proteins that associate with integrin α2 by proteomic analysis in human fibrosarcoma HT-1080 cells

Integrins are adhesion receptors for components of the extracellular matrix (ECMs) that regulate multiple cellular functions, such as migration, invasion, proliferation, and survival by mediating bidirectional signal transmission. Even though many proteins have been reported to associate with integrins both on and in cells, systemic analyses of the adhesome have not been carried out. In previous studies, we identified proteins associating with a membrane-type protease, MT1-MMP, using nano-flow liquid chromatography/tandem mass spectrometry (nano-LC/MS/MS) of associated proteins prepared by optimized conditions for cell lysis and purification. Since integrins were identified as MT1-MMP-associated proteins, we next applied this method to analyze integrin-associated proteins. In this study, we expressed integrin α2 fused at the C terminus to a FLAG peptide in HT1080 cells. Cells stably expressing the chimeric protein were lysed with 1% Brij-98 and affinity purified using anti-FLAG antibody. Integrin β1 co-purified with integrin α2 confirming the specificity of the purification procedure. Analysis of the purified mixture by nano-LC/MS/MS identified 70 proteins. Nineteen of these were membrane proteins, including adhesion proteins, receptors, transporters, proteinases, and ion-channel receptors, and the balance were cytoplasmic. Interestingly, eight of the proteins had previously been shown to associate with MT1-MMP. We believe the present study provides a platform to facilitate the study of the mechanisms of cell adhesion, migration, and invasion.     

Underexpression of mitochondrial-DNA encoded ATP synthesis-related genes and DNA repair genes in systemic lupus erythematosus

Introduction  

Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease characterized by various systemic symptoms and multiple organ damage. We clarify biological and functional abnormalities in SLE by comparing the gene expression profiles of SLE patients with those of healthy individuals.     

Histochemical demonstration of different types of poly-N-acetyllactosamine structures in human thyroid neoplasms using lectins and endo-β-galactosidase digestion

Summary Blood-group-related antigens expressed in papillary carcinomas and other types of neoplasm of the human thyroid glands have been shown to be carried by poly-N-acetyllactosamines containing a linear domain susceptible to endo-β-galactosidase digestion. To make clear more precisely the backbone poly-N-acetyllactosamine structures, labelled lectins specific to different types of these structures and specific to core structures with β1-6GlcNAc branching of N- and O-linked glycoproteins were employed in conjunction with prior endo-β-galactosidase digestion on formalin-fixed, paraffin-embedded neoplasms of the human thyroid glands. In papillary carcinomas,Datura stramonium agglutinin (DSA) and succinyl wheat germ agglutinin (Suc-WGA) reacted most consistently and frequently with papillary carcinomas from all the individuals examined. Pokeweed mitogen (PWM) likewise stained the cells of papillary carcinomas from all the individuals examined, but in some individuals the number of lectin-reactive cells were very small.Lycoperscion esculentum aggultinin (LEA),Solanum tuberosum agglutinin (STA),Phaseolus vulgaris agglutinin L (PHA-L) andArtocarpus integrifolia agglutinin (jacalin) similarly bound to the cancer cells from most of the individuals, and in these cases the number of reactive cells was usually much more restricted than was the case with DSA or PWM. In adenoma and other types of carcinoma, such as follicular carcinomas, these lectins specific to poly-N-acetyllactosamine exhibited slight or no reactivity with the cells, whereas PHA-L and jacalin similarly bound to the cells of adenomas and carcinomas from most of the individuals examined. Prior digestion with endo-β-galactosidase completely eliminated or markedly reduced the reactivity with PWM and LEA in papillary carcinomas. Reactivity with DSA, Suc-WGA, STA, PHA-L and jacalin was slightly reduced or not at all affected by enzyme digestion. These results confirmed that poly-N-acetyllactosamine species found in papillary carcinomas are quite different from those in other types of thyroid neoplasm, suggesting that at least three different types of poly-N-acetyllactosamine, that is, linear unbranched short and long sequences and highly branched ones are produced in these cells.     

The Caenorhabditis elegans ADAMTS family gene adt-1 is necessary for morphogenesis of the male copulatory organs.

Remodeling of the extracellular matrix (ECM) is pivotal for various biological processes, including organ morphology and development. The Caenorhabditis elegans male tail has male-specific copulatory organs, the rays and the fan. Ray morphogenesis, which involves a rapid remodeling of the ECM, is an important model of morphogenesis, although its mechanism is poorly understood. ADAMTS (a disintegrin-like and metalloproteinase with thrombospondin type I motifs) is a novel metalloproteinase family that is thought to be an important regulator for ECM remodeling during development and pathological states. We report here that a new C. elegans ADAMTS family gene, adt-1, plays an important regulatory role in ray morphogenesis. Inactivation of the adt-1 gene resulted in morphological changes in the rays as well as the appearance of abnormal protuberances around the rays. In addition, mating ability was remarkably impaired in adt-1 deletion mutant males. Furthermore, we found that the green fluorescent protein reporter driven by the adt-1 promoter was specifically expressed throughout the rays in the male tail. We hypothesize that ADT-1 controls the ray extension process via remodeling of the ECM in the cuticle.     

Myosin VI-Dependent Actin Cages Encapsulate Parkin-Positive Damaged Mitochondria

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A 570-kb DNA Sequence of the Escherichia coli K-12 Genome Corresponding to the 28.0-40.1 min Region on the Linkage Map

The 569,750 base pair sequence corresponding to the 28.0–40.1min region on the genetic map of Escherichia coli K-12 (W3110)was determined. This region includes the replication terminusregion and contained at least 549 potential open reading frames.Among them, 160 (29%) were previously reported, 174 (32%) werehomologous to other known genes, 102 (18%) were identical orsimilar to hypothetical genes registered in databases, and theremaining 113 (21%) did not show a significant similarity toany other gene. Of interest was the finding of a large numberof genes and gene clusters in andnear the replication terminationregion which had been thought to be genetically silent. Thoseincludeda cluster of genes for fatty acid ß-oxidation,the third copy of the pot (spermidine/putrescine transport system)gene cluster, the second dpp (dipeptide transport system) operon,the second dsm (anaerobic dimethyl sulfoxide reductase) operon,a cluster of fim (fimbrial) genes anda DNA helicase-like genewith a high molecular weight. In addition, we found the dnaC-and dnaT-like genes in the cryptic prophage, Rac, anda numberof genes originated probably from plasmids.