enrichment-extracellular-matrix-proteins-from-tissues-digestion-into

Enrichment of Extracellular Matrix Proteins from Tissues and Digestion into Peptides for Mass Spectrometry Analysis

The extracellular matrix (ECM) is a complex meshwork of cross-linked proteins that provides biophysical and biochemical cues that are major regulators of ...

Massachusetts Institute of Technology

The remodeling of epithelial monolayers induced by hepatocyte growth factor (HGF) results in the reorganization of actin cytoskeleton and cellular junctions. We previously showed that the membrane-cytoskeleton linker ezrin plays a major role in HGF-induced morphogenic effects. Here we identified a novel partner of phosphorylated ezrin, the Fes kinase, that acts downstream of ezrin in HGF-mediated cell scattering. We found that Fes interacts directly, through its SH2 domain, with ezrin phosphorylated at tyrosine 477. We show that in epithelial cells, activated Fes localizes either to focal adhesions or cell-cell contacts depending on cell confluency. The recruitment and the activation of Fes to the cell-cell contacts in confluent cells depend on its interaction with ezrin. When this interaction is impaired, Fes remains in focal adhesions and as a consequence the cells show defective spreading and scattering in response to HGF stimulation. Altogether, these results provide a novel mechanism whereby ezrin/Fes interaction at cell-cell contacts plays an essential role in HGF-induced cell scattering and implicates Fes in the cross-talk between cell-cell and cell-matrix adhesion.

Spatial recruitment and activation of the Fes kinase by ezrin promotes HGF-induced cell scattering.

The model of erythroleukemia caused by Spi-1/PU.1 transgenesis in mice is a multistage disease. A preleukemic step is characterized by an acute proliferation of proerythroblasts due to the arrest of differentiation provoked by Spi-1/PU.1. Later on, a blastic crisis occurs associated with somatic oncogenic mutations in the stem cell factor (SCF) receptor kit. To gain insights into the mechanisms of the leukemic progression, we performed proteomic profiling analyses of proerythroblasts isolated at the 2 stages of the disease. Our results indicate that the level of ezrin, a membrane cytoskeletal crosslinker, is increased in the leukemic cells. We show that Kit oncogenic forms are responsible for ezrin phosphorylation and that phosphorylation rather than overexpression is essential in the leukemic proerythroblasts. Using expression of dominant-negative forms of ezrin, we show that phosphorylation of ezrin on residue Y353 participates in apoptosis resistance, whereas phosphorylation on residue Y145 promotes proliferation of the leukemic cells in vitro and in vivo. Another recurrent oncogenic form of tyrosine kinases (Flt3) most frequently involved in human myeloid leukemia was also able to phosphorylate ezrin. These findings point to a new role for ezrin as signaling player in the development of leukemia, being a downstream effector of oncogenic tyrosine kinases in leukemic blasts.

Ezrin is a target for oncogenic Kit mutants in murine erythroleukemia.

Gefitinib inhibits epidermal growth factor-independent angiogenesis, but the molecular mechanism underlying this inhibition has yet to be defined. Here we show that gefitinib dose-dependently inhibited chemotaxis of endothelial cells toward fibroblast growth factor-2 (FGF-2), but not toward vascular endothelial growth factor-A (VEGF-A). Gefitinib inhibited lamellipodium formation by endothelial cells induced by FGF-2, but not by VEGF-A. Gefitinib at 10 microM did not inhibit autophosphorylation of FGF receptor 1 or VEGF receptor 2. A non-receptor protein tyrosine kinase, Fes, has two coiled-coil domains (CCDs) in its N-terminal region. Fes is activated by trans-autophosphorylation through CCD functions. An inactivating mutation in the second CCD abolished FGF-2 activation of Fes, indicating involvement of this CCD in FGF-2-induced Fes activation. Gefitinib-treatment decreased both CCD-independent and FGF-2- or VEGF-A-promoted Fes activity with a maximal decrease at 1 microM. The same results were observed in cells stably expressing kinase-inactive Fes; a dominant negative effect was observed in cells treated with FGF-2, but not with VEGF-A. Taken together, these results indicate that FGF-2 activates Fes via the second CCD, leading to lamellipodium formation and chemotaxis by endothelial cells, and gefitinib may act through Fes as an inhibitor of FGF-2-driven angiogenesis.

Inhibition of endothelial cell chemotaxis toward FGF-2 by gefitinib associates with downregulation of Fes activity.

The highly related ERM (Ezrin, Radixin, Moesin) proteins provide a regulated linkage between the membrane and the underlying actin cytoskeleton. They also provide a platform for the transmission of signals in responses to extracellular cues. Studies in different model organisms and in cultured cells have highlighted the importance of ERM proteins in the generation and maintenance of specific domains of the plasma membrane. A central question is how do ERM proteins coordinate actin filament organization and membrane protein transport/stability with signal transduction pathways to build up complex structures? Through their interaction with numerous partners including membrane proteins, actin cytoskeleton and signaling molecules, ERM proteins have the ability to organize multiprotein complexes in specific cellular compartments. Likewise, ERM proteins participate in diverse functions including cell morphogenesis, endocytosis/exocytosis, adhesion and migration. This review focuses on aspects still poorly understood related to the function of ERM proteins in epithelial cell adhesion and migration.

Emerging role for ERM proteins in cell adhesion and migration.

Completion of genome sequences for many organisms allows a reasonably complete definition of the complement of extracellular matrix (ECM) proteins. In mammals this "core matrisome" comprises ∼300 proteins. In addition there are large numbers of ECM-modifying enzymes, ECM-binding growth factors, and other ECM-associated proteins. These different categories of ECM and ECM-associated proteins cooperate to assemble and remodel extracellular matrices and bind to cells through ECM receptors. Together with receptors for ECM-bound growth factors, they provide multiple inputs into cells to control survival, proliferation, differentiation, shape, polarity, and motility of cells. The evolution of ECM proteins was key in the transition to multicellularity, the arrangement of cells into tissue layers, and the elaboration of novel structures during vertebrate evolution. This key role of ECM is reflected in the diversity of ECM proteins and the modular domain structures of ECM proteins both allow their multiple interactions and, during evolution, development of novel protein architectures.

Overview of the matrisome--an inventory of extracellular matrix constituents and functions.

&#x7EC6;&#x80DE;&#x5916;&#x57FA;&#x8D28; (ECM) &#x662F;&#x4EA4;&#x8054;&#x86CB;&#x767D;&#x8D28;&#x63D0;&#x4F9B;&#x751F;&#x7269;&#x7269;&#x7406;&#x548C;&#x751F;&#x7269;&#x5316;&#x5B66;&#x7684;&#x63D0;&#x793A;&#xFF0C;&#x6709;&#x7684;&#x7EC6;&#x80DE;&#x589E;&#x6B96;&#x3001; &#x5B58;&#x6D3B;&#x3001; &#x5206;&#x5316;&#x548C;&#x8FC1;&#x79FB;&#x7684;&#x91CD;&#x8981;&#x76D1;&#x7BA1;&#x673A;&#x6784;&#x7684;&#x590D;&#x6742;&#x7F51;&#x7EDC;&#x3002;&#x6211;&#x4EEC;&#x5728;&#x5EA7;&#x5236;&#x5B9A;&#x5B9A;&#x6027;&#x7684;&#x4F53;&#x5185; ECM &#x7EC4;&#x6210;&#x6B63;&#x5E38;&#x7EC4;&#x7EC7;&#x548C;&#x4F7F;&#x7528;&#x6D53;&#x7F29;&#x86CB;&#x767D;&#x7684;&#x80BF;&#x7624;&#x86CB;&#x767D;&#x8D28;&#x7EC4;&#x5B66;&#x7814;&#x7A76;&#x6218;&#x7565;&#xFF0C;ECM &#x7EC4;&#x4EF6;&#x53CA;&#x968F;&#x540E;&#x7684;&#x5206;&#x6790;&#x63D0;&#x53D6;&#x8D28;&#x8C31;&#x6CD5;&#x3002;&#x540C;&#x65F6;&#xFF0C;&#x6211;&#x4EEC;&#x53D1;&#x5C55;&#x751F;&#x7269;&#x4FE1;&#x606F;&#x5B66;&#x65B9;&#x6CD5;&#x6765;&#x9884;&#x6D4B;&#x4E2D;&#x7684;&#x7845; &Prime;matrisome&Prime; &#x5B9A;&#x4E49;&#x4E3A; ECM &#x86CB;&#x767D;&#x7684;&#x5408;&#x594F;&#x548C;&#x76F8;&#x5173;&#x56E0;&#x7D20;&#x3002;&#x6211;&#x4EEC;&#x62A5;&#x544A;&#x7EC6;&#x80DE;&#x5916;&#x57FA;&#x8D28;&#x5C0F;&#x9F20;&#x80BA;&#x764C;&#x548C;&#x7ED3;&#x80A0;&#x764C;&#x3001; &#x6BCF;&#x961F;&#x8D85;&#x8FC7; 100 ECM &#x86CB;&#x767D;&#x548C;&#x6BCF;&#x4ECB;&#x7ECD;&#x4E00;&#x4E2A;&#x7279;&#x6027;&#x7B7E;&#x540D;&#x7684;&#x8868;&#x5F81;&#x3002;&#x6B64;&#x5916;&#xFF0C;&#x6211;&#x4EEC;&#x5728;&#x5C0F;&#x9F20;&#x4E2D;&#x4F7F;&#x7528;&#x4EBA;&#x7C7B;&#x80BF;&#x7624;&#x5F02;&#x79CD;&#x79FB;&#x690D;&#xFF0C;&#x663E;&#x793A;&#x80BF;&#x7624;&#x7EC6;&#x80DE;&#x548C;&#x95F4;&#x8D28;&#x7EC6;&#x80DE;&#x6709;&#x52A9;&#x4E8E;&#x751F;&#x4EA7;&#x7684;&#x80BF;&#x7624;&#x77E9;&#x9635;&#x548C;&#x80BF;&#x7624;&#x7684;&#x4E0D;&#x540C;&#x8F6C;&#x79FB;&#x6F5C;&#x80FD;&#x4E0D;&#x540C;&#x80BF;&#x7624;-&#x548C;&#x57FA;&#x8D28;&#x6D3E;&#x751F;&#x7684; ECM &#x7EC4;&#x4EF6;&#x3002;&#x53EF;&#x4EE5;&#x5E7F;&#x6CDB;&#x5E94;&#x7528;&#x7684;&#x7B56;&#x7565;&#xFF0C;&#x6211;&#x4EEC;&#x63CF;&#x8FF0;&#x548C;&#x8BF4;&#x660E;&#x5728;&#x8FD9;&#x91CC;&#x548C;&#x5176;&#x4ED6;&#x4FBF;&#x5229;&#x8FD9;&#x4E9B;&#x65B9;&#x6CD5;&#x4E2D;&#x7684;&#x5E94;&#x7528;&#xFF0C;&#x6211;&#x4EEC;&#x63D0;&#x4F9B;&#x8D44;&#x6E90;&#x5305;&#x62EC;&#x5B9E;&#x9A8C;&#x5BA4;&#x534F;&#x8BAE;&#xFF0C;ECM &#x57DF;&#x548C;&#x86CB;&#x767D;&#x8D28;&#x548C; bioinformatically &#x6D3E;&#x751F;&#x4EBA;&#x8BF4;&#x660E;&#x7684;&#x6E05;&#x5355;&#x548C;&#x9F20;&#x6807; matrisome&#x3002;

Matrisome&#xFF1A; &#x5728;&#x7845;&#x5B9A;&#x4E49;&#x548C;&#x6B63;&#x5E38;&#x548C;&#x80BF;&#x7624;&#x7EC6;&#x80DE;&#x5916;&#x57FA;&#x8D28;&#x7684;&#x86CB;&#x767D;&#x8D28;&#x7EC4;&#x5B66;&#x5728;&#x4F53;&#x5185;&#x8868;&#x5F81;&#x3002;

The extracellular matrix (ECM) is a complex meshwork of cross-linked proteins providing both biophysical and biochemical cues that are important regulators of cell proliferation, survival, differentiation and migration. We present here a proteomic strategy developed to characterize the in vivo ECM composition of normal tissues and tumors using enrichment of protein extracts for ECM components and subsequent analysis by mass spectrometry. In parallel, we have developed a bioinformatic approach to predict the in silico â€³matrisomeâ€³ defined as the ensemble of ECM proteins and associated factors. We report the characterization of the extracellular matrices of murine lung and colon, each comprising more than 100 ECM proteins and each presenting a characteristic signature. Moreover, using human tumor xenografts in mice, we show that both tumor cells and stromal cells contribute to the production of the tumor matrix and that tumors of differing metastatic potential differ in both the tumor- and the stroma-derived ECM components. The strategy we describe and illustrate here can be broadly applied and, to facilitate application of these methods by others, we provide resources including laboratory protocols, inventories of ECM domains and proteins, and instructions for bioinformatically deriving the human and mouse matrisome.

The matrisome: in silico definition and in vivo characterization by proteomics of normal and tumor extracellular matrices.

La matrice extracellulaire (ECM) est un r&eacute;seau complex de prot&eacute;ines r&eacute;ticul&eacute;es fournissant des indices biochimiques et biophysiques qui sont d&#39;importants r&eacute;gulateurs de la survie, la prolif&eacute;ration cellulaire, la diff&eacute;renciation et la migration. Nous pr&eacute;sentons ici une strat&eacute;gie prot&eacute;omiques &eacute;labor&eacute;e afin de caract&eacute;riser la composition ECM in vivo des tissus normaux et des tumeurs &agrave; l&#39;aide de l&#39;enrichissement de la prot&eacute;ine extraits pour composantes d&#39;ECM et de leur analyse par spectrom&eacute;trie de masse. En parall&egrave;le, nous avons d&eacute;velopp&eacute; une approche bioinformatique pour pr&eacute;dire l&#39;in silico &Prime;matrisome&Prime; d&eacute;fini comme l&#39;ensemble des prot&eacute;ines ECM et facteurs connexes. Les auteurs rapportent la caract&eacute;risation des matrices extracellulaires de murin poumon et du c&ocirc;lon, chacun comprenant plus de 100 prot&eacute;ines ECM et chacune pr&eacute;sentant une signature caract&eacute;ristique. En outre, &agrave; l&#39;aide de x&eacute;nogreffes tumorales humaines chez la souris, nous montrons que les cellules tumorales et des cellules stromales contribuent &agrave; la production de la matrice de la tumeur et que des diff&eacute;rence de potentiel m&eacute;tastatique des tumeurs diff&egrave;rent dans la tumeur - tant les composants d&eacute;riv&eacute;s de stroma de ECM. La strat&eacute;gie, les auteurs d&eacute;crivent et illustrent ici peut &ecirc;tre largement appliqu&eacute;e et, pour faciliter l&#39;application de ces m&eacute;thodes par d&#39;autres, nous fournissons des ressources, y compris les protocoles de laboratoire, inventaires des domaines de l&#39;ECM et prot&eacute;ines et des instructions pour bioinformatically d&eacute;rivant de l&#39;homme et la souris matrisome.

Le matrisome : in silico d&eacute;finition et caract&eacute;risation in vivo par prot&eacute;omique des matrices extracellulaires normales et tumorales.

Die extrazellul&auml;re Matrix (ECM) ist ein komplexes Netzwerk von querverbundenen Proteinen bietet sowohl biophysikalische und biochemische Hinweise, die wichtige Regulatoren der Zellproliferation, &uuml;berleben, Differenzierung und Migration sind. Wir pr&auml;sentieren hier eine Proteomic-Strategie entwickelt, um die in-vivo ECM-Zusammensetzung des normalen Geweben und Tumoren mit Anreicherung des Proteins charakterisieren Extrakte f&uuml;r ECM-Komponenten und anschlie&szlig;ende Analyse durch Massenspektrometrie. Parallel haben wir einen Bioinformatic Ansatz zur Vorhersage der in Silico entwickelt &Prime;matrisome&Prime; definiert als Ensemble von ECM-Proteinen und verbundenen Faktoren. Wir berichten die Charakterisierung der extrazellularen Matrizen murine Lung und Doppelpunkt, jeweils mehr als 100 ECM-Proteine und jeweils eine charakteristische Signatur pr&auml;sentiert. Dar&uuml;ber hinaus zeigen mit menschlichen Tumor Xenografts in M&auml;use, wir, dass Tumorzellen und stromal Zellen die Produktion der Tumor Matrix beitragen und Tumoren unterschiedlicher metastasierendem Potenzial in den Tumor- und die Stroma abgeleitete ECM-Komponenten unterscheiden. Die Strategie, die wir beschreiben und illustrieren hier weitgehend angewendet werden kann, und um die Anwendung dieser Methoden von anderen zu erleichtern, bieten wir Ressourcen einschlie&szlig;lich Laborprotokolle, Vorr&auml;te von ECM-Dom&auml;nen und Proteinen und Anweisungen f&uuml;r die Bioinformatically Ableitung der Mensch und Maus Matrisome.

Die Matrisome: in Silico-Definition und in-vivo Charakterisierung von Proteomics von normalen und Tumor extrazellularen Matrizen.

La matrice extracellulare (ECM) &egrave; un complesso reticolo di proteine reticolate fornendo spunti biofisici e biochimici che sono importanti regolatori della proliferazione cellulare, la sopravvivenza, differenziazione e migrazione. Vi presentiamo qui una strategia proteomic sviluppata per caratterizzare la composizione di ECM in vivo dei tessuti normali e tumori mediante arricchimento della proteina estratti per componenti di ECM e successiva analisi di spettrometria di massa. In parallelo, abbiamo sviluppato un approccio di bioinformatic per predire in silico &Prime;matrisome&Prime; definito come l&#39;insieme delle proteine di ECM e associati fattori. Riportiamo la caratterizzazione delle matrici extracellulari del polmone murino e colon, ciascuno composto da pi&ugrave; di 100 proteine ECM e ognuno presenta una firma caratteristica. Inoltre, utilizzando xenotrapianti tumorali umane in topi, mostrano che sia le cellule tumorali e cellule stromali contribuiscono alla produzione della matrice tumorale e che i tumori di diverso potenziale metastatico differiscono nel tumore - sia derivato da stroma ECM componenti. La strategia si descrivono e illustrano qui pu&ograve; essere applicata in generale e, per agevolare l&#39;applicazione di questi metodi da altri, mettiamo a disposizione risorse compresi i protocolli di laboratorio, gli inventari di ECM domini e proteine e le istruzioni per bioinformatically di derivazione umana e matrisome del mouse.

Il matrisome: in silico definizione e caratterizzazione in vivo di proteomica delle matrici extracellulari normali e tumorali.

&#x7D30;&#x80DE;&#x5916;&#x30DE;&#x30C8;&#x30EA;&#x30C3;&#x30AF;&#x30B9; (ECM) &#x306F;&#x3001;&#x8907;&#x96D1;&#x306A;&#x7DB2;&#x76EE;&#x306E;&#x67B6;&#x6A4B;&#x86CB;&#x767D;&#x8CEA;&#x3092;&#x7D30;&#x80DE;&#x5897;&#x6B96;&#x3001;&#x751F;&#x5B58;&#x7387;, &#x5206;&#x5316;&#x304A;&#x3088;&#x3073;&#x79FB;&#x884C;&#x306E;&#x91CD;&#x8981;&#x306A;&#x30EC;&#x30AE;&#x30E5;&#x30EC;&#x30FC;&#x30BF;&#x3067;&#x3059;&#x5E74;&#x4F1A;&#x3068;&#x751F;&#x5316;&#x5B66;&#x306E;&#x624B;&#x304C;&#x304B;&#x308A;&#x3092;&#x63D0;&#x4F9B;&#x3059;&#x308B;&#x306E;&#x3067;&#x3059;&#x3002;&#x3053;&#x3053;&#x3067;&#x30A8;&#x30AD;&#x30B9; ECM &#x30B3;&#x30F3;&#x30DD;&#x30FC;&#x30CD;&#x30F3;&#x30C8;&#x304A;&#x3088;&#x3073;&#x305D;&#x306E;&#x5F8C;&#x306E;&#x5206;&#x6790;&#x7528;&#x8CEA;&#x91CF;&#x5206;&#x6790;&#x6CD5;&#x306B;&#x3088;&#x308B; in vivo &#x3067;&#x306E; ECM &#x7D44;&#x6210;&#x306E;&#x6B63;&#x5E38;&#x7D44;&#x7E54;&#x3068;&#x816B;&#x760D;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x6FC3;&#x7E2E;&#x3092;&#x7279;&#x5FB4;&#x4ED8;&#x3051;&#x308B;&#x305F;&#x3081;&#x306B;&#x958B;&#x767A;&#x30D7;&#x30ED;&#x30C6;&#x30AA;&#x30FC;&#x30E0;&#x6226;&#x7565;&#x3092;&#x63D0;&#x793A;&#x3057;&#x307E;&#x3059;&#x3002;&#x4E26;&#x884C;&#x3057;&#x3066;&#x3001;&#x6211;&#x3005; &#x306F; in silico &#x4E88;&#x6E2C;&#x3059;&#x308B; bioinformatic &#x30A2;&#x30D7;&#x30ED;&#x30FC;&#x30C1;&#x3092;&#x958B;&#x767A;&#x3057;&#x305F; &Prime;matrisome&Prime; ECM &#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x30A2;&#x30F3;&#x30B5;&#x30F3;&#x30D6;&#x30EB;&#x3068;&#x3057;&#x3066;&#x5B9A;&#x7FA9;&#x3055;&#x308C;&#x3066;&#x3044;&#x308B;&#x8981;&#x56E0;&#x3068;&#x306E;&#x95A2;&#x9023;&#x3002;3H1000 &#x30DE;&#x30A6;&#x30B9;&#x80BA;&#x3001;&#x30B3;&#x30ED;&#x30F3;&#x3001;&#x305D;&#x308C;&#x305E;&#x308C; 100 &#x4EE5;&#x4E0A;&#x306E; ECM &#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x3092;&#x69CB;&#x6210;&#x3057;&#x3066;&#x305D;&#x308C;&#x305E;&#x308C;&#x7279;&#x5FB4;&#x7684;&#x306A;&#x7F72;&#x540D;&#x3092;&#x63D0;&#x793A;&#x306E;&#x7D30;&#x80DE;&#x5916;&#x30DE;&#x30C8;&#x30EA;&#x30C3;&#x30AF;&#x30B9;&#x306E;&#x30AD;&#x30E3;&#x30E9;&#x30AF;&#x30BF;&#x30EA;&#x30BC;&#x30FC;&#x30B7;&#x30E7;&#x30F3;&#x3092;&#x5831;&#x544A;&#x3057;&#x307E;&#x3059;&#x3002;&#x307E;&#x305F;&#x3001;&#x816B;&#x760D;&#x306E;&#x30DE;&#x30A6;&#x30B9;&#x3092;&#x4F7F;&#x7528;&#x3057;&#x3066;&#x3001;&#x816B;&#x760D;&#x7D30;&#x80DE;&#x3068;&#x9593;&#x8CEA;&#x7D30;&#x80DE;&#x816B;&#x760D;&#x30DE;&#x30C8;&#x30EA;&#x30C3;&#x30AF;&#x30B9;&#x306E;&#x751F;&#x7523;&#x306B;&#x8CA2;&#x732E;&#x3059;&#x308B;&#x3053;&#x3068;&#x3068;&#x3001;&#x816B;&#x760D; - &#x3068;&#x9593;&#x8CEA;&#x7531;&#x6765; ECM &#x30B3;&#x30F3;&#x30DD;&#x30FC;&#x30CD;&#x30F3;&#x30C8;&#x306E;&#x4E21;&#x65B9;&#x3067;&#x7570;&#x306A;&#x308B;&#x8EE2;&#x79FB;&#x80FD;&#x306E;&#x816B;&#x760D;&#x3068;&#x306F;&#x7570;&#x306A;&#x308B;&#x3053;&#x3068;&#x793A;&#x3059;.&#x6211;&#x3005; &#x306B;&#x3064;&#x3044;&#x3066;&#x8AAC;&#x660E;&#x3057;&#x3001;&#x3053;&#x3053;&#x3067;&#x8AAC;&#x660E;&#x6226;&#x7565;&#x3092;&#x5E83;&#x304F;&#x9069;&#x7528;&#x3059;&#x308B;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x3001;&#x4ED6;&#x306E;&#x4EBA;&#x304C;&#x3053;&#x308C;&#x3089;&#x306E;&#x30E1;&#x30BD;&#x30C3;&#x30C9;&#x306E;&#x30A2;&#x30D7;&#x30EA;&#x30B1;&#x30FC;&#x30B7;&#x30E7;&#x30F3;&#x3092;&#x5BB9;&#x6613;&#x306B;&#x3059;&#x308B;&#x306B;&#x306F;&#x3001;&#x5B9F;&#x9A13;&#x5BA4;&#x306E;&#x30D7;&#x30ED;&#x30C8;&#x30B3;&#x30EB;&#x3001;ECM &#x30C9;&#x30E1;&#x30A4;&#x30F3;&#x304A;&#x3088;&#x3073;&#x624B;&#x9806;&#x306B;&#x3064;&#x3044;&#x3066;&#x306F;&#x3001;&#x4EBA;&#x9593;&#x306E;&#x6D3E;&#x751F; bioinformatically &#x3068;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x5728;&#x5EAB;&#x306A;&#x3069;&#x306E;&#x30EA;&#x30BD;&#x30FC;&#x30B9;&#x3092;&#x63D0;&#x4F9B;&#x3057;&#x3066;&#x30DE;&#x30A6;&#x30B9; matrisome&#x3002;

Matrisome&#xFF1A; silico &#x5B9A;&#x7FA9;&#x3068; in vivo &#x306E;&#x30AD;&#x30E3;&#x30E9;&#x30AF;&#x30BF;&#x30EA;&#x30BC;&#x30FC;&#x30B7;&#x30E7;&#x30F3; &#x30D7;&#x30ED;&#x30C6;&#x30AA;&#x30DF;&#x30AF;&#x30B9;&#x306B;&#x3088;&#x308B;&#x6B63;&#x5E38;&#x304A;&#x3088;&#x3073;&#x816B;&#x760D;&#x7D30;&#x80DE;&#x5916;&#x30DE;&#x30C8;&#x30EA;&#x30C3;&#x30AF;&#x30B9;&#x306E;&#x3067;&#x3002;

&#xAE30;&#xC9C8; (ECM) &#xC138;&#xD3EC; &#xC99D;&#xC2DD;, &#xC0DD;&#xC874;, &#xBD84;&#xD654; &#xBC0F; &#xB9C8;&#xC774;&#xADF8;&#xB808;&#xC774;&#xC158;&#xC758; &#xC911;&#xC694; &#xD55C; &#xB808; &#xADE4; &#xB808;&#xC774; &#xD130;&#xB294; &#xC0DD;&#xBB3C; &#xB9AC; &#xBC0F; &#xC0DD; &#xD654; &#xD655; &#xC801;&#xC778; &#xB2E8;&#xC11C;&#xB97C; &#xC81C;&#xACF5; &#xD558;&#xB294; &#xAD50;&#xCC28; &#xC5F0;&#xACB0; &#xB41C; &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xBCF5;&#xC7A1; &#xD55C; &#xCC44;&#xB110; &#xC774;&#xB2E4;. &#xC5EC;&#xAE30;&#xC5D0; &#xB098;&#xC624;&#xB294; &#xC815;&#xC0C1; &#xC870;&#xC9C1;&#xACFC; &#xC885;&#xC591; &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xB18D;&#xCD95;&#xC744; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; in vivo ECM &#xAD6C;&#xC131;&#xC758; &#xD2B9;&#xC131;&#xC744; &#xAC1C;&#xBC1C; proteomic &#xC804;&#xB7B5; ECM &#xAD6C;&#xC131; &#xC694;&#xC18C; &#xBC0F; &#xD6C4;&#xC18D; &#xBD84;&#xC11D;&#xC744; &#xC704;&#xD55C; &#xCD94;&#xCD9C; &#xBB3C; &#xC9C8;&#xB7C9; &#xBD84;&#xC11D; &#xD558; &#xC5EC;. &#xBCD1;&#xB82C;, &#xC2DC;&#xB0B4; silico &#xC608;&#xCE21; bioinformatic &#xC811;&#xADFC; &#xBC29;&#xC2DD;&#xC744; &#xAC1C;&#xBC1C; &#xD588;&#xC2B5;&#xB2C8;&#xB2E4; &Prime;matrisome&Prime; ECM &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xC559;&#xC0C1;&#xBE14;&#xC73C;&#xB85C; &#xC815;&#xC758; &#xD558; &#xACE0; &#xAD00;&#xB828; &#xC694;&#xC778;. &#xC6B0;&#xB9AC; murine &#xD3D0;&#xC640; &#xCF5C;&#xB860;, 100 &#xAC1C; &#xC774;&#xC0C1;&#xC758; ECM &#xB2E8;&#xBC31;&#xC9C8;&#xC73C;&#xB85C; &#xAD6C;&#xC131; &#xB41C; &#xAC01;&#xAC01; &#xBC0F; &#xD2B9;&#xC131; &#xC11C;&#xBA85;&#xC744; &#xC81C;&#xC2DC; &#xD558; &#xACE0; &#xAC01;&#xAC01;&#xC758; &#xC138;&#xD3EC; &#xC678; &#xB9E4;&#xD2B8;&#xB9AD;&#xC2A4;&#xC758; &#xD2B9;&#xC131;&#xC744; &#xBCF4;&#xACE0; &#xD569;&#xB2C8;&#xB2E4;. &#xB610;&#xD55C;, &#xC885;&#xC591; &#xC138;&#xD3EC;&#xC640; &#xAE30;&#xC9C8; &#xC138;&#xD3EC; &#xC885;&#xC591; &#xB9E4;&#xD2B8;&#xB9AD;&#xC2A4;&#xC758; &#xC0DD;&#xC0B0;&#xC5D0; &#xAE30;&#xC5EC; &#xD558; &#xACE0; &#xC11C;&#xB85C; &#xB2E4;&#xB978; &#xC7A0;&#xC7AC;&#xB825;&#xC744; &#xC804;&#xC774;&#xC131; &#xC885;&#xC591; &#xC885;&#xC591;-&#xC640; ECM &#xAE30;&#xC9C8; &#xD30C;&#xC0DD; &#xB41C; &#xAD6C;&#xC131; &#xC694;&#xC18C;&#xC5D0; &#xB530;&#xB77C; &#xB2E4;&#xB97C; &#xBCF4;&#xC5EC; &#xC778;&#xAC04;&#xC758; &#xC885;&#xC591; xenografts &#xB9C8;&#xC6B0;&#xC2A4; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC;, &#xC6B0;&#xB9AC;. &#xC6B0;&#xB9AC;&#xAC00; &#xC124;&#xBA85; &#xD558; &#xACE0; &#xC5EC;&#xAE30;&#xC11C; &#xC124;&#xBA85; &#xD558;&#xB294; &#xC804;&#xB7B5;&#xC744; &#xAD11;&#xBC94;&#xC704; &#xD558; &#xAC8C; &#xC801;&#xC6A9; &#xB420; &#xC218; &#xC788;&#xB2E4; &#xACE0; &#xB2E4;&#xB978; &#xC0AC;&#xB78C;&#xC5D0; &#xC758;&#xD574;&#xC774; &#xBC29;&#xBC95;&#xC758; &#xC751;&#xC6A9; &#xD504;&#xB85C;&#xADF8;&#xB7A8;&#xC744; &#xCD09;&#xC9C4; &#xD558;&#xAE30; &#xC704;&#xD558;&#xC5EC; &#xC6B0;&#xB9AC; &#xC2E4;&#xD5D8;&#xC2E4; &#xD504;&#xB85C;&#xD1A0;&#xCF5C;, ECM &#xB3C4;&#xBA54;&#xC778; &#xBC0F; &#xB2E8;&#xBC31;&#xC9C8;&#xACFC; bioinformatically &#xC778;&#xAC04; &#xD30C;&#xC0DD;&#xC5D0; &#xB300; &#xD55C; &#xC9C0;&#xCE68;&#xC758; &#xC7AC;&#xACE0;&#xB97C; &#xD3EC;&#xD568; &#xD55C; &#xB9AC;&#xC18C;&#xC2A4;&#xB97C; &#xC81C;&#xACF5; &#xD558; &#xACE0; &#xB9C8;&#xC6B0;&#xC2A4; matrisome.

Matrisome: silico &#xC815;&#xC758; &#xBC0F; &#xC815;&#xC0C1; &#xBC0F; &#xC885;&#xC591; &#xC138;&#xD3EC; &#xC678; &#xB9E4;&#xD2B8;&#xB9AD;&#xC2A4;&#xC758; proteomics&#xC5D0; &#xC758;&#xD574; vivo&#xC5D0;&#xC11C; &#xD2B9;&#xC131;&#xD654;.

La matriz extracelular (ECM) es una red compleja de prote&iacute;nas reticuladas proporcionando se&ntilde;ales tanto biof&iacute;sicos y bioqu&iacute;micos que son importantes reguladores de la proliferaci&oacute;n celular, supervivencia, diferenciaci&oacute;n y migraci&oacute;n. Presentamos aqu&iacute; una estrategia prote&oacute;mica desarrollada para caracterizar la composici&oacute;n de ECM en vivo de los tejidos normales y tumores con enriquecimiento de prote&iacute;na extractos para componentes de ECM y posterior an&aacute;lisis por espectrometr&iacute;a de masas. Paralelamente, hemos desarrollado un enfoque de bioinform&aacute;tica para predecir la silico en &Prime;matrisome&Prime; se define como el conjunto de prote&iacute;nas ECM y asociados factores. Divulgamos la caracterizaci&oacute;n de las matrices extracelulares de murino pulm&oacute;n y colon, cada uno compuesto por m&aacute;s de 100 prote&iacute;nas ECM y cada uno presenta una firma caracter&iacute;stica. Por otra parte, con xenoinjertos tumorales humanas en ratones, mostramos que tanto las c&eacute;lulas del tumor y las c&eacute;lulas estromales contribuyen a la producci&oacute;n de la matriz del tumor y que tumores de potencial metast&aacute;tico diferente difieren en el tumor - y los componentes derivados del estroma de ECM. La estrategia describen e ilustran aqu&iacute; puede aplicarse ampliamente y, para facilitar la aplicaci&oacute;n de estos m&eacute;todos por otros, proveemos recursos incluyendo los protocolos de laboratorio, inventarios de dominios de ECM y prote&iacute;nas y las instrucciones para bioinformatically que el ser humano y rat&oacute;n matrisome.

El matrisome: en definici&oacute;n de silico y caracterizaci&oacute;n in vivo por prote&oacute;mica de matrices extracelulares normal y tumoral.

&#x89C4;&#x7BA1;&#x5237;&#x8FB9;&#x5883;&#x7ED2;&#x6BDB;&#x4E0A;&#x76AE;&#x7EC6;&#x80DE;&#x7684;&#x5F62;&#x6001;&#x53D1;&#x751F;&#x5BFC;&#x81F4;&#x7684;&#x808C;&#x52A8;&#x86CB;&#x767D;&#x4E1D;&#x805A;&#x5408;&#x7684;&#x673A;&#x5236;&#x4ECD;&#x7136;&#x672A;&#x77E5;&#x3002;Eps8&#xFF0C;&#x5927;&#x5BB6;&#x5EAD;&#x7684;&#x86CB;&#x767D;&#x8D28;&#x80FD;&#x591F;&#x76D6;&#x548C;&#x6346;&#x7ED1;&#x808C;&#x52A8;&#x86CB;&#x767D;&#x4E1D;&#x539F;&#x578B;&#x8868;&#x660E;&#x8981;&#x6346;&#x7ED1; microvillar &#x808C;&#x52A8;&#x86CB;&#x767D;&#x4E1D;&#x3002;&#x6211;&#x4EEC;&#x7684;&#x62A5;&#x544A;&#xFF0C;Eps8L1a&#xFF0C;Eps8 &#x5BB6;&#x65CF;&#x6210;&#x5458;&#x7684;&#x548C;&#x65B0;&#x578B;&#x7684; ezrin &#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x5408;&#x4F5C;&#x4F19;&#x4F34;&#xFF0C;&#x901A;&#x8FC7;&#x5176;&#x4E0A;&#x9650;&#x8BBE;&#x7F6E;&#x6D3B;&#x52A8;&#x63A7;&#x4EF6;&#x6CBC;&#x867E;&#x957F;&#x5EA6;&#x3002;Eps8L1a &#x7684;&#x67AF;&#x7AED;&#x5BFC;&#x81F4;&#x5F62;&#x6210;&#x7684;&#x957F;&#x7ED2;&#x6BDB;&#xFF0C;&#x800C;&#x5176;&#x8FC7;&#x5EA6;&#x8868;&#x8FBE;&#x4E86;&#x76F8;&#x53CD;&#x7684;&#x6548;&#x679C;&#x3002;&#x6211;&#x4EEC;&#x8BC1;&#x660E;&#x8FD9; ezrin &#x5DEE;&#x5F02;&#x8C03;&#x8282;&#x6CBC;&#x867E;&#x5927;&#x4F1A;&#x671F;&#x95F4;&#x7684; Eps8 &#x548C; Eps8L1a &#x7684;&#x808C;&#x52A8;&#x86CB;&#x767D;&#x4E0A;&#x9650;&#x548C;&#x6346;&#x7ED1;&#x6D3B;&#x52A8;&#x3002;Eps8 &#x4E0E; ezrin &#x7684;&#x5171;&#x8868;&#x8FBE;&#x4FC3;&#x8FDB;&#x5F62;&#x6210;&#x7684;&#x819C;&#x8936;&#x8FB9;&#x548C;&#x5854;&#x592B;&#x8328;&#x7684;&#x7ED2;&#x6BDB;&#xFF0C;&#x800C; ezrin &#x548C; Eps8L1a &#x7684;&#x8868;&#x8FBE;&#x8BF1;&#x5BFC;&#x7684;&#x805A;&#x7C7B;&#x5206;&#x6790;&#x808C;&#x52A8;&#x86CB;&#x767D;&#x542B;&#x5728;&#x7EC6;&#x80DE;&#x8868;&#x9762;&#x7684;&#x7ED3;&#x6784;&#x3002;&#x8FD9;&#x4E9B;&#x72EC;&#x7279;&#x7684;&#x5F62;&#x6001;&#x53D8;&#x5316;&#x65E2;&#x4E0D;&#x662F;&#x89C2;&#x5BDF;&#x5230;&#x7A81;&#x53D8;&#x4F53;&#x7684; ezrin &#x7F3A;&#x9677;&#x5728;&#x5176;&#x7ED1;&#x5B9A;&#x5230; Eps8/Eps8L1a &#x4E0E; Eps8 &#x6216; Eps8L1a &#x7684; coexpressed &#x65F6;&#xFF0C;&#x4E5F;&#x4E0D;&#x9075;&#x5B88; ezrin &#x4E0E;&#x808C;&#x52A8;&#x86CB;&#x767D;&#x6346;&#x7ED1;&#x5728;&#x4E00;&#x8D77;&#x7684; Eps8 &#x6216; Eps8L1a &#x6709;&#x7F3A;&#x9677;&#x7A81;&#x53D8;&#x4F53;&#x7684;&#x8868;&#x793A;&#x65F6;&#x6216;-&#x5C01;&#x9876;&#x6D3B;&#x52A8;&#xFF0C;&#x5206;&#x522B;&#x3002;&#x6211;&#x4EEC;&#x7684;&#x6570;&#x636E;&#x663E;&#x793A;&#x4E2D;&#x7684;&#x7A0B;&#x5E8F;&#x96C6;&#x548C;&#x808C;&#x52A8;&#x86CB;&#x767D; microvillar &#x4E1D;&#x7EC4;&#x7EC7; ezrin &#x548C; Eps8 &#x86CB;&#x767D;&#x7684;&#x534F;&#x540C;&#x4F5C;&#x7528;&#x3002;

Ezrin &#x7684; Eps8 &#x86CB;&#x767D;&#x8D28;&#x7684;&#x4E0D;&#x540C;&#x6D3B;&#x52A8;&#xFF0C;&#x4EE5;&#x4FC3;&#x8FDB;&#x89C4;&#x8303;&#x6CBC;&#x867E;&#x5F62;&#x6001;&#x53D1;&#x751F;&#x3002;

The mechanisms that regulate actin filament polymerization resulting in the morphogenesis of the brush border microvilli in epithelial cells remain unknown. Eps8, the prototype of a family of proteins capable of capping and bundling actin filaments, has been shown to bundle the microvillar actin filaments. We report that Eps8L1a, a member of the Eps8 family and a novel ezrin-interacting partner, controls microvillus length through its capping activity. Depletion of Eps8L1a leads to the formation of long microvilli, whereas its overexpression has the opposite effect. We demonstrate that ezrin differentially modulates the actin-capping and -bundling activities of Eps8 and Eps8L1a during microvillus assembly. Coexpression of ezrin with Eps8 promotes the formation of membrane ruffles and tufts of microvilli, whereas expression of ezrin and Eps8L1a induces the clustering of actin-containing structures at the cell surface. These distinct morphological changes are neither observed when a mutant of ezrin defective in its binding to Eps8/Eps8L1a is coexpressed with Eps8 or Eps8L1a nor observed when ezrin is expressed with mutants of Eps8 or Eps8L1a defective in the actin-bundling or -capping activities, respectively. Our data show a synergistic effect of ezrin and Eps8 proteins in the assembly and organization of actin microvillar filaments.

Ezrin regulates microvillus morphogenesis by promoting distinct activities of Eps8 proteins.

Les m&eacute;canismes qui r&eacute;gulent la polym&eacute;risation de l&#39;actine filament r&eacute;sultant dans la morphogen&egrave;se de des microvillosit&eacute;s de fronti&egrave;re de brosse dans les cellules &eacute;pith&eacute;liales restent inconnus. Eps8, le prototype d&#39;une famille de prot&eacute;ines capables de bouchage et de groupement des filaments d&#39;actine, a d&eacute;montr&eacute; que les filaments d&#39;actine microvillosit&eacute;s de grouper. Nous rapportons que Eps8L1a, un membre de la famille Eps8 et un partenaire d&#39;ezrin interagissant avec roman, longueur de microvillosit&eacute;s contr&ocirc;les par le biais de son activit&eacute; de bouchage. Appauvrissement de la couche de Eps8L1a conduit &agrave; la formation des microvillosit&eacute;s longues, alors que sa surexpression a l&#39;effet inverse. Nous d&eacute;montrons qu&#39;ezrin module diff&eacute;remment les activit&eacute;s actine-capsulage et - groupement de Eps8 et Eps8L1a lors de l&#39;assemblage des microvillosit&eacute;s. Coexpression d&#39;ezrin avec Eps8 favorise la formation de la membrane volants et des touffes de microvillosit&eacute;s, tandis que l&#39;expression d&#39;ezrin et Eps8L1a induit la clusterisation actine contenant des structures &agrave; la surface cellulaire. Ces changements morphologiques distincts sont observ&eacute;es quand un mutant d&#39;ezrin d&eacute;fectueux &agrave; sa liaison &agrave; Eps8/Eps8L1a est coexpriment avec Eps8 ou Eps8L1a ni observ&eacute;es quand Bob ezrin est exprim&eacute;e avec des mutants de d&eacute;fectueux dans l&#39;actine-groupement Eps8 ou Eps8L1a ou - bouchage des activit&eacute;s, respectivement. Nos r&eacute;sultats montrent un effet synergique d&#39;ezrin et prot&eacute;ines Eps8 par l&#39;Assembl&eacute;e et l&#39;Organisation des filaments d&#39;actine microvillosit&eacute;s.

Ezrin r&eacute;glemente la morphogen&egrave;se des microvillosit&eacute;s en promouvant des activit&eacute;s distinctes de prot&eacute;ines Eps8.

Die Mechanismen, die Aktin-Filament-Polymerisation f&uuml;hrt die Morphogenese der Pinsel Grenze Mikrovilli Epithelzellen Regeln bleiben unbekannt. Eps8, der Prototyp einer Familie von Proteinen, die Lage, Begrenzung und B&uuml;ndelung von Aktin-Filamenten, hat sich gezeigt, die microvillar Aktin-Filamente zu b&uuml;ndeln. Wir berichten, dass Eps8L1a, ein Mitglied aus der Familie der Eps8 und eine neuartige Ezrin-Interaktion Partner, Steuerelemente Mikrovilli L&auml;nge durch seine Abdeckfl&auml;che Aktivit&auml;t. Ersch&ouml;pfung der Eps8L1a f&uuml;hrt zur Bildung von langen Mikrovilli, w&auml;hrend seine &Uuml;berexpression den gegenteiligen Effekt hat. Wir zeigen, dass Ezrin differentiell die Aktin-capping und -b&uuml;ndeln Aktivit&auml;ten von Eps8 und Eps8L1a bei der Montage der Mikrovilli moduliert. Coexpression von Ezrin mit Eps8 f&ouml;rdert die Bildung von Membran-R&uuml;schen und Haarb&uuml;schel Mikrovilli, w&auml;hrend Ausdruck von Bob Ezrin und Eps8L1a induziert die B&uuml;ndelung von Aktin-haltigen Strukturen auf der Zelloberfl&auml;che. Diese unterschiedlichen morphologischen Ver&auml;nderungen sind eingehalten, wenn eine Mutante von Ezrin Defekt in seiner Bindung an Eps8/Eps8L1a mit Eps8 oder Eps8L1a coexpressed ist weder beobachtet, wenn Bob Ezrin mit Mutanten von Eps8 oder Eps8L1a Defekt in der Aktin-B&uuml;ndelung bemessen ist oder - Aktivit&auml;ten, bzw. Kappung. Unsere Daten zeigen eine synergistische Wirkung von Bob Ezrin und Eps8 Proteine in die Assembly und die Organisation der microvillar Aktin-Filamenten.

Bob Ezrin reguliert Mikrovilli Morphogenese durch eigene Aktivit&auml;ten der Eps8 Proteine zu f&ouml;rdern.

I meccanismi che regolano la polimerizzazione di filamenti di actina con conseguente la morfogenesi dei microvilli di confine pennello nelle cellule epiteliali rimangono sconosciuti. Eps8, il prototipo di una famiglia di proteine in grado di tappatura e impacchettare i filamenti di actina, ha dimostrato di impacchettare i filamenti di actina microvillare. Segnaliamo che Eps8L1a, un membro della famiglia Eps8 e un romanzo partner interagenti ezrin, lunghezza microvillo controlli attraverso la sua attivit&agrave; tappatura. Deplezione di Eps8L1a conduce alla formazione di lunghi microvilli, mentre la sua sovraespressione ha l&#39;effetto opposto. Dimostriamo che ezrin differenzialmente modula l&#39;actina-tappatura - impacchettante attivit&agrave; ed Eps8 e Eps8L1a durante il montaggio microvillo. Coexpression di ezrin con Eps8 promuove la formazione di increspature di membrana e ciuffi di microvilli, espressione di ezrin ed Eps8L1a induce il clustering di actina contenente strutture alla superficie delle cellule. Questi cambiamenti morfologici distinti non sono osservati quando un mutante di ezrin difettoso nella sua associazione a Eps8/Eps8L1a &egrave; coespressi con Eps8 o Eps8L1a, n&eacute; osservati quando ezrin &egrave; espresso con mutanti di Eps8 o Eps8L1a difettoso in actina-impacchettare o - tappatura attivit&agrave;, rispettivamente. I nostri dati mostrano un effetto sinergico di ezrin ed Eps8 proteine nell&#39;assembly e organizzazione di filamenti di actina microvillare.

Ezrin regola la morfogenesi microvillo promuovendo attivit&agrave; distinte delle proteine Eps8.

&#x30D6;&#x30E9;&#x30B7;&#x306E;&#x56FD;&#x5883;&#x5FAE;&#x7D68;&#x6BDB;&#x4E0A;&#x76AE;&#x7D30;&#x80DE;&#x306E;&#x5F62;&#x614B;&#x5F62;&#x6210;&#x306B;&#x304A;&#x3051;&#x308B;&#x7D50;&#x679C;&#x30A2;&#x30AF;&#x30C1;&#x30F3; &#x30D5;&#x30A3;&#x30E9;&#x30E1;&#x30F3;&#x30C8;&#x91CD;&#x5408;&#x3092;&#x8ABF;&#x7BC0;&#x3059;&#x308B;&#x30E1;&#x30AB;&#x30CB;&#x30BA;&#x30E0;&#x306F;&#x4E0D;&#x660E;&#x306E;&#x307E;&#x307E;&#x3002;Eps8&#x3001;&#x30AD;&#x30E3;&#x30C3;&#x30D4;&#x30F3;&#x30B0; &#x30A2;&#x30AF;&#x30C1;&#x30F3; &#x30D5;&#x30A3;&#x30E9;&#x30E1;&#x30F3;&#x30C8;&#x3092;&#x675F;&#x306D;&#x308B;&#x3053;&#x3068;&#x304C;&#x3067;&#x304D;&#x308B;&#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x5BB6;&#x65CF;&#x306E;&#x30D7;&#x30ED;&#x30C8;&#x30BF;&#x30A4;&#x30D7;&#x306F; microvillar &#x306E;&#x30A2;&#x30AF;&#x30C1;&#x30F3; &#x30D5;&#x30A3;&#x30E9;&#x30E1;&#x30F3;&#x30C8;&#x3092;&#x30D0;&#x30F3;&#x30C9;&#x30EB;&#x3059;&#x308B;&#x793A;&#x3055;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x305D;&#x306E; Eps8L1a&#x3001;Eps8 &#x306E;&#x5BB6;&#x65CF;&#x306E;&#x4E00;&#x54E1;&#x3068;&#x5C0F;&#x8AAC;&#x30A8;&#x30BA;&#x30EA;&#x30F3;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x306E;&#x30D1;&#x30FC;&#x30C8;&#x30CA;&#x30FC;&#x306B;&#x30B3;&#x30F3;&#x30C8;&#x30ED;&#x30FC;&#x30EB;&#x3057;&#x305F;&#x5FAE;&#x7D68;&#x6BDB;&#x306E;&#x9577;&#x3055;&#x305D;&#x306E;&#x30AD;&#x30E3;&#x30C3;&#x30D4;&#x30F3;&#x30B0;&#x6D3B;&#x52D5;&#x3092;&#x901A;&#x3057;&#x3066;&#x5831;&#x544A;&#x3057;&#x307E;&#x3059;&#x3002;&#x305D;&#x306E;&#x904E;&#x5270;&#x767A;&#x73FE;&#x306F;&#x9006;&#x306E;&#x52B9;&#x679C;&#x304C;&#x3042;&#x308B;&#x4E00;&#x65B9; Eps8L1a &#x306E;&#x67AF;&#x6E07;&#x9577;&#x3044;&#x5FAE;&#x7D68;&#x6BDB;&#x306E;&#x5F62;&#x6210;&#x306B; &#x3057;&#x307E;&#x3059;&#x3002;&#x6211;&#x3005; &#x306F;&#x305D;&#x306E;&#x30A8;&#x30BA;&#x30EA;&#x30F3;&#x5DEE;&#x52D5;&#x306E; Eps8 &#x3068; Eps8L1a &#x306E;&#x30A2;&#x30AF;&#x30C1;&#x30F3; &#x30AD;&#x30E3;&#x30C3;&#x30D4;&#x30F3;&#x30B0;&#x3068;&#x30D0;&#x30F3;&#x30C9;&#x30EA;&#x30F3;&#x30B0;&#x6D3B;&#x52D5;&#x3057;&#x305F;&#x5FAE;&#x7D68;&#x6BDB;&#x30A2;&#x30BB;&#x30F3;&#x30D6;&#x30EA;&#x4E2D;&#x306B;&#x5909;&#x8ABF;&#x3092;&#x793A;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x30A8;&#x30BA;&#x30EA;&#x30F3; Eps8 &#x3068;&#x306E;&#x5171;&#x767A;&#x73FE;&#x4FC3;&#x9032;&#x819C;&#x30D5;&#x30EA;&#x30EB;&#x306E;&#x5F62;&#x6210;&#x3068;&#x5FAE;&#x7D68;&#x6BDB;&#x3001;&#x30BF;&#x30D5;&#x30C4;&#x306E;&#x30A2;&#x30AF;&#x30C1;&#x30F3;&#x3092;&#x542B;&#x3080;&#x30AF;&#x30E9;&#x30B9;&#x30BF; &#x30EA;&#x30F3;&#x30B0; &#x30A8;&#x30BA;&#x30EA;&#x30F3;&#x304A;&#x3088;&#x3073; Eps8L1a &#x306E;&#x767A;&#x73FE;&#x3092;&#x8A98;&#x5C0E;&#x3059;&#x308B;&#x306E;&#x306B;&#x5BFE;&#x3057;&#x3001;&#x7D30;&#x80DE;&#x8868;&#x9762;&#x306E;&#x69CB;&#x9020;&#x3002;&#x3053;&#x308C;&#x3089;&#x306E;&#x7570;&#x306A;&#x308B;&#x5F62;&#x614B;&#x306E;&#x5909;&#x5316;&#x306F; Eps8 &#x307E;&#x305F;&#x306F; Eps8L1a &#x306E;&#x30A8;&#x30BA;&#x30EA;&#x30F3;&#x6B20;&#x9665;&#x3068;&#x306E;&#x7D50;&#x5408;&#x306B; Eps8/Eps8L1a &#x3067;&#x306E;&#x7A81;&#x7136;&#x5909;&#x7570;&#x4F53;&#x306E;&#xFF09; &#x3068;&#x304D;&#x89B3;&#x5BDF;&#x3082;&#x30A8;&#x30BA;&#x30EA;&#x30F3;&#x5909;&#x7570;&#x30A2;&#x30AF;&#x30C1;&#x30F3;&#x675F;&#x3067; Eps8 &#x307E;&#x305F;&#x306F; Eps8L1a &#x306E;&#x6B20;&#x9665;&#x306E;&#x8868;&#x73FE;&#x304C;&#x89B3;&#x5BDF;&#x307E;&#x305F;&#x306F; - &#x305D;&#x308C;&#x305E;&#x308C;&#x306E;&#x6D3B;&#x52D5;&#x3092;&#x30AD;&#x30E3;&#x30C3;&#x30D4;&#x30F3;&#x30B0;&#x3057;&#x307E;&#x3059;&#x3002;&#x6211;&#x3005; &#x306E;&#x30C7;&#x30FC;&#x30BF; &#x30A2;&#x30BB;&#x30F3;&#x30D6;&#x30EA;&#x3068; microvillar &#x306E;&#x30A2;&#x30AF;&#x30C1;&#x30F3;&#x306E;&#x7D44;&#x7E54;&#x3067;&#x30A8;&#x30BA;&#x30EA;&#x30F3;&#x3068; Eps8 &#x86CB;&#x767D;&#x8CEA;&#x306E;&#x76F8;&#x4E57;&#x52B9;&#x679C;&#x3092;&#x793A;&#x3057;&#x307E;&#x3059;&#x3002;

&#x30A8;&#x30BA;&#x30EA;&#x30F3; Eps8 &#x30BF;&#x30F3;&#x30D1;&#x30AF;&#x8CEA;&#x306E;&#x500B;&#x5225;&#x306E;&#x6D3B;&#x52D5;&#x3092;&#x63A8;&#x9032;&#x3057;&#x305F;&#x5FAE;&#x7D68;&#x6BDB;&#x306E;&#x5F62;&#x614B;&#x5F62;&#x6210;&#x3092;&#x8ABF;&#x7BC0;&#x3057;&#x307E;&#x3059;&#x3002;

&#xC0C1;&#xD53C; &#xC138;&#xD3EC;&#xC5D0;&#xC11C; &#xBE0C;&#xB7EC;&#xC2DC; &#xD14C;&#xB450;&#xB9AC; microvilli morphogenesis &#xACB0;&#xACFC; &#xB9D0;&#xB77C; &#xD544; &#xB77C; &#xBA58; &#xD2B8; &#xC911; &#xD569;&#xC744; &#xC870;&#xC808; &#xD558;&#xB294; &#xBA54;&#xCEE4;&#xB2C8;&#xC998;&#xC744; &#xC54C; &#xC218; &#xC5C6;&#xB294; &#xB0A8;&#xC544;. Eps8, &#xC0C1;&#xD55C; &#xACE0; &#xBC88;&#xB4E4; &#xB9D0;&#xB77C; &#xD544; &#xB77C; &#xBA58; &#xD2B8;, &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xAC00;&#xC871;&#xC758; &#xD504;&#xB85C;&#xD1A0; &#xD0C0;&#xC785; &#xBC88;&#xB4E4; microvillar &#xB9D0;&#xB77C; &#xD544; &#xB77C; &#xBA58; &#xD2B8;&#xB85C; &#xD45C;&#xC2DC; &#xB418;&#xC5C8;&#xC2B5;&#xB2C8;&#xB2E4;. &#xC6B0;&#xB9AC; &#xADF8; Eps8L1a, Eps8 &#xC81C;&#xD488;&#xAD70; &#xBC0F; &#xC18C;&#xC124; ezrin &#xC791;&#xC6A9; &#xD30C;&#xD2B8;&#xB108;&#xC758; &#xC0C1;&#xD55C; &#xD65C;&#xB3D9;&#xC744; &#xD1B5;&#xD574; &#xCEE8;&#xD2B8;&#xB864; microvillus &#xAE38;&#xC774; &#xBCF4;&#xACE0;. &#xBC18;&#xBA74; &#xBC18;&#xB300;&#xC758; &#xD6A8;&#xACFC;&#xAC00; &#xC790;&#xC0AC;&#xC758; overexpression eps8l1a&#xC758; &#xACE0;&#xAC08; &#xAE34; microvilli&#xC758; &#xD615;&#xC131;&#xC5D0; &#xC774;&#xB974;&#xAC8C;. &#xC6B0;&#xB9AC;&#xB294; &#xADF8; ezrin microvillus &#xC870;&#xB9BD; &#xD558;&#xB294; &#xB3D9;&#xC548; Eps8 &#xBC0F; eps8l1a&#xC758; &#xB9D0;&#xB77C; &#xC0C1;&#xD55C; &#xBC0F;-&#xBC88;&#xB4E4; &#xD65C;&#xB3D9;&#xC744; &#xCC28;&#xB3D9; &#xBCC0;&#xC870; &#xBC29;&#xBC95;&#xC744; &#xBCF4;&#xC5EC; &#xC90D;&#xB2C8;&#xB2E4;. &#xBC18;&#xBA74; ezrin &#xBC0F; eps8l1a&#xC758; &#xC2DD; &#xC720;&#xB3C4; &#xD3EC;&#xD568; &#xD558;&#xB294; &#xB9D0;&#xB77C; &#xD074;&#xB7EC;&#xC2A4;&#xD130;&#xB9C1; ezrin eps8&#xC640; coexpression &#xB9C9; &#xC8FC;&#xB984; &#xD615;&#xC131;&#xACFC; microvilli, tufts &#xCD09;&#xC9C4; &#xC138;&#xD3EC; &#xD45C;&#xBA74;&#xC5D0;&#xC11C; &#xAD6C;&#xC870;. &#xC774;&#xB7EC;&#xD55C; &#xBA85;&#xBC31;&#xD55C; &#xD615;&#xD0DC;&#xD559; &#xBCC0;&#xD654; &#xB54C; ezrin Eps8/eps8l1a&#xB97C; &#xBC14;&#xC778;&#xB529; &#xAE30;&#xB2A5;&#xC5D0; &#xACB0;&#xD568;&#xC774; &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; Eps8 &#xB610;&#xB294; eps8l1a&#xC640; coexpressed&#xB294; &#xAD00;&#xCC30;&#xB3C4; &#xB9D0;&#xB77C; &#xBC88;&#xB4E4;&#xC5D0; Eps8 &#xB610;&#xB294; Eps8L1a &#xACB0;&#xD568; &#xB3CC;&#xC5F0;&#xBCC0;&#xC774; ezrin &#xD45C;&#xD604; &#xD558;&#xB294; &#xB54C; &#xAD00;&#xCC30; &#xB610;&#xB294;-&#xD65C;&#xB3D9;&#xC744; &#xAC01;&#xAC01; &#xC0C1;&#xD55C;. &#xC6B0;&#xB9AC;&#xC758; &#xB370;&#xC774;&#xD130; &#xC5B4;&#xC148;&#xBE14;&#xB9AC;&#xC640; &#xB9D0;&#xB77C; microvillar &#xD544; &#xB77C; &#xBA58; &#xD2B8;&#xC758; &#xC870;&#xC9C1; ezrin Eps8 &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xC2DC;&#xB108;&#xC9C0; &#xD6A8;&#xACFC; &#xBCF4;&#xC5EC;&#xC900;&#xB2E4;.

Ezrin Eps8 &#xB2E8;&#xBC31;&#xC9C8;&#xC758; &#xACE0;&#xC720; &#xD65C;&#xB3D9;&#xC744; &#xD64D;&#xBCF4; &#xD558; &#xC5EC; microvillus morphogenesis &#xC870;&#xC808;.

Desconocemos los mecanismos que regulan la polimerizaci&oacute;n de filamentos de actina dando por resultado la morfog&eacute;nesis de las microvellosidades de frontera de cepillo en las c&eacute;lulas epiteliales. Eps8, el prototipo de una familia de prote&iacute;nas capaces de tapado y agrupaci&oacute;n de filamentos de actina, se ha demostrado que los filamentos de actina microvillar del paquete. Divulgamos Eps8L1a, un miembro de la familia Eps8 y novela socio interacci&oacute;n ezrin, longitud de microvilli de controles a trav&eacute;s de su actividad que capsula. Agotamiento de Eps8L1a conduce a la formaci&oacute;n de largas microvellosidades, mientras que su sobreexpresi&oacute;n tiene el efecto contrario. Demostramos que Ezrin diferencialmente modula las actividades de actina-capsular y liar de Eps8 y Eps8L1a durante el montaje de microvilli. Coexpresi&oacute;n de ezrin con Eps8 promueve la formaci&oacute;n de membrana volantes y penachos de microvellosidades, mientras que induce la expresi&oacute;n de ezrin y Eps8L1a el agrupamiento de actina que contienen estructuras en la superficie celular. Estos cambios morfol&oacute;gicos distintos se observ&oacute; cuando un mutante de ezrin defectuoso en su uni&oacute;n a Eps8/Eps8L1a es coexpressed con Eps8 o Eps8L1a ni observ&oacute; cuando ezrin es expresado con mutantes de Eps8 o Eps8L1a defectuoso en la actina-liar o - capsular actividades, respectivamente. Nuestros datos muestran un efecto sin&eacute;rgico de ezrin y prote&iacute;nas Eps8 en el montaje y organizaci&oacute;n de microvillar filamentos de actina.

Ezrin regula morfog&eacute;nesis microvilli promoviendo actividades distintas de las prote&iacute;nas Eps8.

The membrane cytoskeletal crosslinker, ezrin, a member of the ERM family of proteins, is frequently over-expressed in human breast cancers, and is required for motility and invasion of epithelial cells. Our group previously showed that ezrin acts co-operatively with the non-receptor tyrosine kinase, Src, in deregulation of cell-cell contacts and scattering of epithelial cells. In particular, ezrin phosphorylation on Y477 by Src is specific to ezrin within the ERM family, and is required for HGF-induced scattering of epithelial cells. We therefore sought to examine the role of Y477 phosphorylation in ezrin on tumor progression.

Ezrin phosphorylation on tyrosine 477 regulates invasion and metastasis of breast cancer cells.

Towards definition of an ECM parts list: an advance on GO categories.

The extracellular matrix (ECM) is a major component of tumors and a significant contributor to cancer progression. In this study, we use proteomics to investigate the ECM of human mammary carcinoma xenografts and show that primary tumors of differing metastatic potential differ in ECM composition. Both tumor cells and stromal cells contribute to the tumor matrix and tumors of differing metastatic ability differ in both tumor- and stroma-derived ECM components. We define ECM signatures of poorly and highly metastatic mammary carcinomas and these signatures reveal up-regulation of signaling pathways including TGFβ and VEGF. We further demonstrate that several proteins characteristic of highly metastatic tumors (LTBP3, SNED1, EGLN1, and S100A2) play causal roles in metastasis, albeit at different steps. Finally we show that high expression of LTBP3 and SNED1 correlates with poor outcome for ER(-)/PR(-)breast cancer patients. This study thus identifies novel biomarkers that may serve as prognostic and diagnostic tools. DOI: http://dx.doi.org/10.7554/eLife.01308.001.

Extracellular matrix signatures of human mammary carcinoma identify novel metastasis promoters.

Colorectal cancer is the third most frequently diagnosed cancer and the third cause of cancer deaths in the United States. Despite the fact that tumor cell-intrinsic mechanisms controlling colorectal carcinogenesis have been identified, novel prognostic and diagnostic tools as well as novel therapeutic strategies are still needed to monitor and target colon cancer progression. We and others have previously shown, using mouse models, that the extracellular matrix (ECM), a major component of the tumor microenvironment, is an important contributor to tumor progression. In order to identify candidate biomarkers, we sought to define ECM signatures of metastatic colorectal cancers and their metastases to the liver.

Extracellular matrix signatures of human primary metastatic colon cancers and their metastases to liver.

The extracellular matrix (ECM) is a fundamental component of multicellular organisms that provides mechanical and chemical cues that orchestrate cellular and tissue organization and functions. Degradation, hyperproduction or alteration of the composition of the ECM cause or accompany numerous pathologies. Thus, a better characterization of ECM composition, metabolism, and biology can lead to the identification of novel prognostic and diagnostic markers and therapeutic opportunities. The development over the last few years of high-throughput ("omics") approaches has considerably accelerated the pace of discovery in life sciences. In this review, we describe new bioinformatic tools and experimental strategies for ECM research, and illustrate how these tools and approaches can be exploited to provide novel insights in our understanding of ECM biology. We also introduce a web platform "the matrisome project" and the database MatrisomeDB that compiles in silico and in vivo data on the matrisome, defined as the ensemble of genes encoding ECM and ECM-associated proteins. Finally, we present a first draft of an ECM atlas built by compiling proteomics data on the ECM composition of 14 different tissues and tumor types.

Article	Year
Spatial recruitment and activation of the Fes kinase by ezrin promotes HGF-induced cell scattering. The EMBO journal\| PubMed ID: 18046454	2008
Ezrin is a target for oncogenic Kit mutants in murine erythroleukemia. Blood\| PubMed ID: 18182570	2008
Inhibition of endothelial cell chemotaxis toward FGF-2 by gefitinib associates with downregulation of Fes activity. International journal of oncology\| PubMed ID: 19885553	2009
Emerging role for ERM proteins in cell adhesion and migration. Cell adhesion & migration\| PubMed ID: 21343695	2011
Overview of the matrisome--an inventory of extracellular matrix constituents and functions. Cold Spring Harbor perspectives in biology\| PubMed ID: 21937732	2012
The matrisome: in silico definition and in vivo characterization by proteomics of normal and tumor extracellular matrices. Molecular & cellular proteomics : MCP\| PubMed ID: 22159717	2011
Ezrin regulates microvillus morphogenesis by promoting distinct activities of Eps8 proteins. Molecular biology of the cell\| PubMed ID: 22262457	2012
Ezrin phosphorylation on tyrosine 477 regulates invasion and metastasis of breast cancer cells. BMC cancer\| PubMed ID: 22397367	2012
Towards definition of an ECM parts list: an advance on GO categories. Matrix biology : journal of the International Society for Matrix Biology\| PubMed ID: 23199376	2012
Extracellular matrix signatures of human mammary carcinoma identify novel metastasis promoters. eLife\| PubMed ID: 24618895	2014
Extracellular matrix signatures of human primary metastatic colon cancers and their metastases to liver. BMC cancer\| PubMed ID: 25037231	2014
The Extracellular Matrix: Tools and Insights for the "Omics" Era. Matrix biology : journal of the International Society for Matrix Biology\| PubMed ID: 26163349	2015

Alexandra Naba

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