3,709 research outputs found
Ezrin phosphorylation on tyrosine 477 regulates invasion and metastasis of breast cancer cells
No full textBackground 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. Methods Using a highly metastatic mouse mammary carcinoma cell line (AC2M2), we tested the effect of over-expressing a non-phosphorylatable form of ezrin (Y477F) on invasive colony growth in 3-dimensional Matrigel cultures, and on local invasion and metastasis in an orthotopic engraftment model. Results AC2M2 cells over-expressing Y477F ezrin exhibited delayed migration in vitro, and cohesive round colonies in 3-dimensional Matrigel cultures, compared to control cells that formed invasive colonies with branching chains of cells and numerous actin-rich protrusions. Moreover, over-expression of Y477F ezrin inhibits local tumor invasion in vivo. Whereas orthotopically injected wild type AC2M2 tumor cells were found to infiltrate into the abdominal wall and visceral organs within three weeks, tumors expressing Y477F ezrin remained circumscribed, with little invasion into the surrounding stroma and abdominal wall. Additionally, Y477F ezrin reduces the number of lung metastatic lesions. Conclusions Our study implicates a role of Y477 ezrin, which is phosphorylated by Src, in regulating local invasion and metastasis of breast carcinoma cells, and provides a clinically relevant model for assessing the Src/ezrin pathway as a potential prognostic/predictive marker or treatment target for invasive human breast cancer.Canadian Breast Cancer Research Alliance (BEE, 017374)Canadian Institutes of Health Research (BEE, 102644)Physicians Society Inc.Association pour le développement de la recherche sur le cancer (France
Towards definition of an ECM parts list: An advance on GO categories
No full textThose of us interested in the extracellular matrix (ECM) are faced with significant challenges of definition. ECM proteins are large, complex and assembled into crosslinked insoluble matrices. This has meant that defining the biochemical composition of ECMs has been difficult. Nonetheless, protein chemistry and molecular biology have defined many familiar ECM proteins — collagens, proteoglycans, laminins, thrombospondins, tenascins, fibronectins, etc. With the completion of many genomes it should now be possible to develop complete “parts lists” for the ECM. Such lists are needed for analyzing data from “omic” approaches such as expression arrays, latest-generation sequencing and proteomics. These approaches generate long lists and it is typically necessary to extract from those lists the genes/proteins of interest. Anyone who attempts to do this using the commonly used gene ontology (GO) categories soon discovers that they are largely useless for defining ECM proteins. Many ECM proteins are unannotated and those which are, are sorted, with little evidence of logic or consistency, into diverse categories such “extracellular matrix,” “basement membrane,” “cell surface” and many others. The human and mouse orthologs are often found in different categories and attempts to use GO categories to extract a complete list of ECM genes or proteins from a data set are unsatisfactory at best
Ezrin regulates microvillus morphogenesis by promoting distinct activities of Eps8 proteins
No full textThe 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.Association pour le développement de la recherche sur le cancer (France) (3267)Association pour le développement de la recherche sur le cancer (France) (4823)France. Agence nationale de la recherche (05BLAN033001)GenHomme Network (Grant 02490-6088
Enrichment of Extracellular Matrix Proteins from Tissues and Digestion into Peptides for Mass Spectrometry Analysis
No full textThe extracellular matrix (ECM) is a complex meshwork of cross-linked proteins that provides biophysical and biochemical cues that are major regulators of cell proliferation, survival, migration, etc. The ECM plays important roles in development and in diverse pathologies including cardio-vascular and musculo-skeletal diseases, fibrosis, and cancer. Thus, characterizing the composition of ECMs of normal and diseased tissues could lead to the identification of novel prognostic and diagnostic biomarkers and potential novel therapeutic targets. However, the very nature of ECM proteins (large in size, cross-linked and covalently bound, heavily glycosylated) has rendered biochemical analyses of ECMs challenging. To overcome this challenge, we developed a method to enrich ECMs from fresh or frozen tissues and tumors that takes advantage of the insolubility of ECM proteins. We describe here in detail the decellularization procedure that consists of sequential incubations in buffers of different pH and salt and detergent concentrations and that results in 1) the extraction of intracellular (cytosolic, nuclear, membrane and cytoskeletal) proteins and 2) the enrichment of ECM proteins. We then describe how to deglycosylate and digest ECM-enriched protein preparations into peptides for subsequent analysis by mass spectrometry.United States. Department of Defense (DOD Innovator Award W81XWH-14-1-0240)National Cancer Institute (U.S.) (Grant U54 CA126515/CA163109)Broad Institute of MIT and HarvardHoward Hughes Medical InstituteNational Cancer Institute (U.S.) (David H. Koch Institute for Integrative Cancer Research at MIT. Grant P30-CA14051
Extracellular matrix signatures of human mammary carcinoma identify novel metastasis promoters
No full textThe 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.National Cancer Institute (U.S.) (Tumor Microenvironment Network, grant no. U54 CA126515/CA163109)National Cancer Institute (U.S.) (David H. Koch Institute Support Grant P30-CA14051)Howard Hughes Medical InstituteBroad Institute of MIT and HarvardNational Institutes of Health (U.S.) (NIH/National Research and Service Award)Virginia and D. K. Ludwig Fund for Cancer ResearchNational Cancer Cente
Author, Philosopher Alexandra Stoddard to Speak March 2 at Williams Library
No full textOXFORD, Miss. – Contemporary philosopher, author, interior designer and speaker Alexandra Stoddard gives an inspirational lecture and reading March 2 at the University of Mississippi
sj-xlsx-3-jhc-10.1369_00221554211061359 – Supplemental material for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas
No full textSupplemental material, sj-xlsx-3-jhc-10.1369_00221554211061359 for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas by Carine Renner, Clarissa Gomez, Mike R. Visetsouk, Isra Taha, Aisha Khan, Stephanie M. McGregor, Paul Weisman, Alexandra Naba, Kristyn S. Masters and Pamela K. Kreeger in Journal of Histochemistry & Cytochemistry</p
sj-pdf-1-jhc-10.1369_00221554211061359 – Supplemental material for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas
No full textSupplemental material, sj-pdf-1-jhc-10.1369_00221554211061359 for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas by Carine Renner, Clarissa Gomez, Mike R. Visetsouk, Isra Taha, Aisha Khan, Stephanie M. McGregor, Paul Weisman, Alexandra Naba, Kristyn S. Masters and Pamela K. Kreeger in Journal of Histochemistry & Cytochemistry</p
sj-xlsx-4-jhc-10.1369_00221554211061359 – Supplemental material for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas
No full textSupplemental material, sj-xlsx-4-jhc-10.1369_00221554211061359 for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas by Carine Renner, Clarissa Gomez, Mike R. Visetsouk, Isra Taha, Aisha Khan, Stephanie M. McGregor, Paul Weisman, Alexandra Naba, Kristyn S. Masters and Pamela K. Kreeger in Journal of Histochemistry & Cytochemistry</p
sj-xlsx-5-jhc-10.1369_00221554211061359 – Supplemental material for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas
No full textSupplemental material, sj-xlsx-5-jhc-10.1369_00221554211061359 for Multi-modal Profiling of the Extracellular Matrix of Human Fallopian Tubes and Serous Tubal Intraepithelial Carcinomas by Carine Renner, Clarissa Gomez, Mike R. Visetsouk, Isra Taha, Aisha Khan, Stephanie M. McGregor, Paul Weisman, Alexandra Naba, Kristyn S. Masters and Pamela K. Kreeger in Journal of Histochemistry & Cytochemistry</p
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