DOCSLIB.ORG

Vitronectin–Avb3 Integrin Engagement Directs Hypoxia- Resistant Mtor Activity and Sustained Protein Synthesis Linked to Invasion by Breast Cancer Cells

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Vitronectin–Avb3 Integrin Engagement Directs Hypoxia- Resistant Mtor Activity and Sustained Protein Synthesis Linked to Invasion by Breast Cancer Cells Published OnlineFirst May 30, 2013; DOI: 10.1158/0008-5472.CAN-13-0218 Cancer Tumor and Stem Cell Biology Research Vitronectin–avb3 Integrin Engagement Directs Hypoxia- Resistant mTOR Activity and Sustained Protein Synthesis Linked to Invasion by Breast Cancer Cells Carolina Pola1, Silvia C. Formenti2,3, and Robert J. Schneider1,2,3 Abstract The tumor microenvironment is a crucial player in the ability of cancer cells to acquire the ability to survive under the hypoxic environment and promote migration and invasion. Translational regulation is an essential part of cancer development and progression. Protein synthesis consumes considerable cellular metabolic energy and is therefore highly regulated, in turn controlling tumor cell proliferation and survival in extreme tumor–host conditions. Protein synthesis is typically downregulated by hypoxia, impairing cell proliferation and migration. Here, we show that breast cancer cells expressing integrin avb3, when engaging the extracellular matrix (ECM) protein vitronectin, strongly upregulate both mTOR activity and cap- dependent mRNA translation, which overrides their inhibition by hypoxia and facilitates tumor cell invasion. Interaction of vitronectin with integrin avb3 results in the continued activation of the kinase mTOR despite hypoxia through a mechanism that is dependent on integrin-linked kinase but is independent of focal adhesion kinase. Continuous activation of mTOR despite hypoxia involves release of translation initiation factor eIF4E from its repressor protein 4E-BP1, which is required for vitronectin-mediated tumor cell invasion. As integrin avb3 is associated with breast cancer cell invasion and metastasis to bone, we propose that the interaction with specific ECM proteins can influence cancer cell invasion, in part, by hyperactivation of mTOR, thereby promoting and sustaining protein synthesis under hypoxic conditions. Cancer Res; 73(14); 1–8. Ó2013 AACR. Introduction mRNA translation is a checkpoint that is blocked in response to Tumor cells and their microenvironment maintain a dynam- certain physiologic stresses, including low levels of oxygen ic interaction, exchanging growth factors and cytokines and during hypoxia (6). eIF4F consists of the cap-binding protein transforming a local extracellular matrix (ECM) into an acti- eIF4E, scaffold protein eIF4G, and the ATP-dependent RNA vated stroma (1, 2). The active host microenvironment can helicase, eIF4A (7). Assembly of eIF4F is controlled by the modify gene expression programs in the tumor cell and trigger availability of eIF4E through regulation by the 4E-binding signaling pathways that control cancer cell proliferation, proteins (4E-BP). Hyperphosphorylation of 4E-BP1 by the migration and invasion (3). Components of the tumor stroma mTOR kinase complex 1 (mTORC1) maintains the inactive include a highly structured and tissue-specific ECM, stromal state of 4E-BP1, so that it cannot bind eIF4E (8). Hypoxia cells, infiltrating inflammatory cells, activated fibroblasts, and downregulates mTORC1, activating (dephosphorylating) 4E- endothelial cells (4, 5). BP1, in turn sequestering eIF4E and downregulating cap- Protein synthesis is highly regulated at the initiation phase dependent mRNA translation (6, 9). Both the noncellular – and involves recruitment of ribosomes with the eukaryotic structural components of the ECM (10 12) and hypoxia (6) initiation factor 4F (eIF4F), a complex of proteins, to the m7 can regulate protein synthesis, but their interaction has never GTP capped structure at the 50-end of mRNAs. Initiation of been extensively investigated. We therefore asked whether external signals in the breast ECM play a role in uncoupling breast cancer cell mTOR inhibition during hypoxia to sustain Authors' Affiliations: Department of 1Microbiology and 2Radiation Oncol- protein synthesis, a requirement for cancer cell invasion and ogy, and 3NYU Cancer Institute, New York University School of Medicine, New York, New York disease progression (8). To date, ECM regulators of mRNA translation include Note: Supplementary data for this article are available at Cancer Research fi Online (http://cancerres.aacrjournals.org/). bronectin (10, 11) and integrin receptors, several of which are overexpressed on breast cancers and mediate cell–ECM Corresponding Author: Robert J. Schneider, NYU School of Medicine, Alexandria Center for Life Sciences, 450 East 29th Street, New York, NY interactions (13). The regulatory effect of different integrins 10016. Phone: 212-263-6006; Fax: 646-501-4541; E-mail: on protein synthesis has also not been extensively studied. In [email protected] breast tumors, malignant progression and bone metastasis doi: 10.1158/0008-5472.CAN-13-0218 are strongly associated with the expression of integrin avb3 Ó2013 American Association for Cancer Research. (13–15). www.aacrjournals.org OF1 Downloaded from cancerres.aacrjournals.org on September 30, 2021. © 2013 American Association for Cancer Research. Published OnlineFirst May 30, 2013; DOI: 10.1158/0008-5472.CAN-13-0218 Pola et al. In this study, we show that vitronectin engagement of minutes in Dulbecco's Modified Eagle's Media (DMEM). After integrin avb3, typically present on breast cancer cells meta- incubation, cells were added to dishes coated with either 25 static to bone, hyperactivates mTOR, even during hypoxia, mg/mL collagen IV native or denatured, 10 mg/mL laminin 1, or thereby overriding the downregulation of cap-dependent 2.5 mg/mL vitronectin. Cells were allowed to attach for 12 to 16 mRNA translation. Vitronectin is an ECM component localized hours. Plating efficiency was determined as the ratio of plated primarily in the stroma and is generally only abundantly in cells to adhered cells at 12 to 16 hours using the MTT assay for contact with tumor cells after breach of the basement mem- cell number. brane during cancer cell invasion (3, 16). We show that stimulation of breast cancer cell mTOR by integrin avb3- SDS-PAGE, immunoblot analysis vitronectin interaction is linked to strongly increased tumor Cells were washed with ice-cold 1 PBS and lysed in 0.5% cell invasion during hypoxia, through a mechanism that lysis buffer [50 mmol/L HEPES, pH 7.0, 150 mmol/L NaCl, 2 involves integrin-linked kinase (ILK) but not focal adhesion mmol/L EDTA, 0.5% NP-40, 25 mmol/L NaF, 2 mmol/L sodium kinase (FAK), and mTOR activation of eIF4E-dependent mRNA orthovanadate, 25 mmol/L glycerophosphate, protease inhib- translation. Thus, integrin-mediated stimulation of protein itor (Roche)], clarified by centrifugation at 4C for 15 minutes synthesis directed by specific ECM components plays an at 13,000 rpm, protein concentration determined, solubilized important role in promoting hypoxia resistance and tumor and SDS-PAGE carried out, and protein immunoblotting con- cell invasion. ducted. Immunoblotting and protein detection by chemilumi- nescence were carried out (Amersham). Material and Methods Cell lines, cell culture, plasmids, and antibodies Protein synthesis m 35 Mouse monoclonal anti-eIF4A antibody was provided by W. Cells were labeled with 50 Ci of [ S]-methionine per mL Merrick (Case Western Reserve University, Cleveland, OH). (Easytag Express Protein Labeling Mix, Dupont/NEN) in Other antibodies were from commercial sources and included DMEM without cold methionine for 1 hour, washed twice  fi rabbit polyclonal antibodies to eIF4E, eIF4G, 4E-BP1-P (Thr 37/ with ice-chilled 1 PBS, lysed, and clari ed as above and fi 46), p70S6K, p70S6K-P (Thr 389), S6rp, S6rp-P (Ser 235/236), speci c activity determined by trichloroacetic acid (TCA) fi AMPKa-P (Thr 172), FAK, FAK-P (Tyr 576/577), mTOR, mTOR- precipitation onto GF/C lters and liquid scintillation count- P (Ser 2448), Akt and Akt-P (Ser 473), ILK, eIF2a and a-P (Ser ing. In hypoxic cells, labeling was conducted using pre-equil- 51; Cell Signaling Technology); integrin b3 (BD Biosciences ibrated media within the hypoxic chamber. Pharmingen), horseradish peroxidase (HRP)-conjugated don- key anti-rabbit or sheep anti-mouse secondary antibodies Lentivirus short hairpin RNA expression vectors (Amersham). Breast cancer cell lines MDA-MB-435, HTB-20, Short hairpin RNAs (shRNA) and scrambled nonsilencing MCF10A, and MDA-MB-231 were obtained from the American (NS) shRNAs were delivered by transduction of cells with lentivirus shRNA expression vectors. Double-stranded shRNAs Type Culture Collection. Cell lines were subjected to IMPACT 0 testing, used within 6 months for this study and grown as for cloning into lentivirus vectors were directed to the 3 - recommended. Human plasma collagen IV, laminin 1 (from untranslated region of mRNAs. Viruses were produced and basement membrane of Engelbreth-Holm-Swarm mouse sar- cells infected/selected with polybrene (4 mg/mL) and puro- m coma), and vitronectin from human plasma were from Sigma. mycin at 0.8 g/mL for 48 hours. MAb 1976/LM609 (anti-human integrin avb3) was from Che- micon International. Transfection of plasmid DNA was carried MTT proliferation assay m Ninety-six–well plates coated with vitronectin (2.5 mg/mL) out using standard conditions (Fugene) with 1 to 2 g of DNA. Transfection of siRNA to eIF4E was carried out on cells at 50% at 37 C for 1 hour were blocked with 1% bovine serum albumin  3 confluency using 5.6 mLof20mmol/L siRNA per 5  10 cells by (BSA) in PBS. MB-435 cells (2 10 per well) were plated in Oligofectamine (Invitrogen), according to manufacturer's triplicate in 1% serum in DMEM and time-lapse analyzed for 7 instructions, in the absence of serum and antibiotics. Normal days. MTT dye (Sigma) was added to wells and incubated for 4 growth media was replaced after 4 to 6 hours. siRNA to hours, cells were solubilized with MTT stripping buffer (10% eIF4E (Ambion): AAGCAAACCUGCGGCUGAUCU. Nonsilen- SDS, 45% isopropanol, 0.04 N HCl) per well, and optical density cing RNA was purchased from Ambion. determined at 570 nm. Hypoxia treatments Cell invasion assay Hypoxia experiments were carried out using a BioSphere Filter undersides of Transwells were coated with vitronectin hypoxia incubator at 0.5% O2, 95% N2,5%CO2 with growing (2.5 mg/mL) at 37 C for 1 hour, plates blocked as above and cells subjected to hypoxia for 24 hours unless otherwise noted.
Load more

Recommended publications
  • Global Analysis Reveals the Complexity of the Human Glomerular Extracellular Matrix
    Global analysis reveals the complexity of the human glomerular extracellular matrix Rachel Lennon,1,2 Adam Byron,1,* Jonathan D. Humphries,1 Michael J. Randles,1,2 Alex Carisey,1 Stephanie Murphy,1,2 David Knight,3 Paul E. Brenchley,2 Roy Zent,4,5 and Martin J. Humphries.1 1Wellcome Trust Centre for Cell-Matrix Research, Faculty of Life Sciences, University of Manchester, Manchester, UK; 2Faculty of Medical and Human Sciences, University of Manchester, Manchester, UK; 3Biological Mass Spectrometry Core Facility, Faculty of Life Sciences, University of Manchester, Manchester, UK; 4Division of Nephrology, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA; and 5Veterans Affairs Hospital, Nashville, TN, USA. *Present address: Edinburgh Cancer Research UK Centre, Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh, UK. Running title: Proteome of the glomerular matrix Word count: Abstract: 208, main text 2765 Corresponding author: Dr Rachel Lennon, Wellcome Trust Centre for Cell-Matrix Research, Michael Smith Building, University of Manchester, Manchester M13 9PT, UK. Phone: 0044 (0) 161 2755498. Fax: 0044 (0) 161 2755082. Email: [email protected] Abstract The glomerulus contains unique cellular and extracellular matrix (ECM) components, which are required for intact barrier function. Studies of the cellular components have helped to build understanding of glomerular disease; however, the full composition and regulation of glomerular ECM remains poorly understood. Here, we employed mass spectrometry–based proteomics of enriched ECM extracts for a global analysis of human glomerular ECM in vivo and identified a tissue-specific proteome of 144 structural and regulatory ECM proteins. This catalogue includes all previously identified glomerular components, plus many new and abundant components.
    [Show full text]
  • Bruch's Membrane Abnormalities in PRDM5-Related Brittle Cornea
    Porter et al. Orphanet Journal of Rare Diseases (2015) 10:145 DOI 10.1186/s13023-015-0360-4 RESEARCH Open Access Bruch’s membrane abnormalities in PRDM5-related brittle cornea syndrome Louise F. Porter1,2,3, Roberto Gallego-Pinazo4, Catherine L. Keeling5, Martyna Kamieniorz5, Nicoletta Zoppi6, Marina Colombi6, Cecilia Giunta7, Richard Bonshek2,8, Forbes D. Manson1 and Graeme C. Black1,9* Abstract Background: Brittle cornea syndrome (BCS) is a rare, generalized connective tissue disorder associated with extreme corneal thinning and a high risk of corneal rupture. Recessive mutations in transcription factors ZNF469 and PRDM5 cause BCS. Both transcription factors are suggested to act on a common pathway regulating extracellular matrix genes, particularly fibrillar collagens. We identified bilateral myopic choroidal neovascularization as the presenting feature of BCS in a 26-year-old-woman carrying a novel PRDM5 mutation (p.Glu134*). We performed immunohistochemistry of anterior and posterior segment ocular tissues, as expression of PRDM5 in the eye has not been described, or the effects of PRDM5-associated disease on the retina, particularly the extracellular matrix composition of Bruch’smembrane. Methods: Immunohistochemistry using antibodies against PRDM5, collagens type I, III, and IV was performed on the eyes of two unaffected controls and two patients (both with Δ9-14 PRDM5). Expression of collagens, integrins, tenascin and fibronectin in skin fibroblasts of a BCS patient with a novel p.Glu134* PRDM5 mutation was assessed using immunofluorescence. Results: PRDM5 is expressed in the corneal epithelium and retina. We observe reduced expression of major components of Bruch’s membrane in the eyes of two BCS patients with a PRDM5 Δ9-14 mutation.
    [Show full text]
  • Integrin Adhesion Molecules in the Human Endometrium. Correlation with the Normal and Abnormal Menstrual Cycle
    Integrin adhesion molecules in the human endometrium. Correlation with the normal and abnormal menstrual cycle. B A Lessey, … , S M Albelda, C A Buck J Clin Invest. 1992;90(1):188-195. https://doi.org/10.1172/JCI115835. Research Article Integrins are a class of cell adhesion molecules that participate in cell-cell and cell-substratum interactions and are present on essentially all human cells. The distribution of nine different alpha and beta integrin subunits in human endometrial tissue at different stages of the menstrual cycle was determined using immunoperoxidase staining. Glandular epithelial cells expressed primarily alpha 2, alpha 3, and alpha 6 (collagen/laminin receptors), while stromal cells expressed predominantly alpha 5 (fibronectin receptor). The presence of alpha 1 on glandular epithelial cells was cycle specific, found only during the secretory phase. Expression of both subunits of the vitronectin receptor, alpha v beta 3, also underwent cycle specific changes on endometrial epithelial cells. Immunostaining for alpha v increased throughout the menstrual cycle, while the beta 3 subunit appeared abruptly on cycle day 20 on luminal as well as glandular epithelial cells. Discordant luteal phase biopsies (greater than or equal to 3 d "out of phase") from infertility patients exhibited delayed epithelial beta 3 immunostaining. These results demonstrate similarities, as well as specific differences, between endometrium and other epithelial tissues. Certain integrin moieties appear to be regulated within the cycling endometrium and disruption of integrin expression may be associated with decreased uterine receptivity and infertility. Find the latest version: https://jci.me/115835/pdf Integrin Adhesion Molecules in the Human Endometrium Correlation with the Normal and Abnormal Menstrual Cycle Bruce A.
    [Show full text]
  • Supplementary Table 1: Adhesion Genes Data Set
    Supplementary Table 1: Adhesion genes data set PROBE Entrez Gene ID Celera Gene ID Gene_Symbol Gene_Name 160832 1 hCG201364.3 A1BG alpha-1-B glycoprotein 223658 1 hCG201364.3 A1BG alpha-1-B glycoprotein 212988 102 hCG40040.3 ADAM10 ADAM metallopeptidase domain 10 133411 4185 hCG28232.2 ADAM11 ADAM metallopeptidase domain 11 110695 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 195222 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 165344 8751 hCG20021.3 ADAM15 ADAM metallopeptidase domain 15 (metargidin) 189065 6868 null ADAM17 ADAM metallopeptidase domain 17 (tumor necrosis factor, alpha, converting enzyme) 108119 8728 hCG15398.4 ADAM19 ADAM metallopeptidase domain 19 (meltrin beta) 117763 8748 hCG20675.3 ADAM20 ADAM metallopeptidase domain 20 126448 8747 hCG1785634.2 ADAM21 ADAM metallopeptidase domain 21 208981 8747 hCG1785634.2|hCG2042897 ADAM21 ADAM metallopeptidase domain 21 180903 53616 hCG17212.4 ADAM22 ADAM metallopeptidase domain 22 177272 8745 hCG1811623.1 ADAM23 ADAM metallopeptidase domain 23 102384 10863 hCG1818505.1 ADAM28 ADAM metallopeptidase domain 28 119968 11086 hCG1786734.2 ADAM29 ADAM metallopeptidase domain 29 205542 11085 hCG1997196.1 ADAM30 ADAM metallopeptidase domain 30 148417 80332 hCG39255.4 ADAM33 ADAM metallopeptidase domain 33 140492 8756 hCG1789002.2 ADAM7 ADAM metallopeptidase domain 7 122603 101 hCG1816947.1 ADAM8 ADAM metallopeptidase domain 8 183965 8754 hCG1996391 ADAM9 ADAM metallopeptidase domain 9 (meltrin gamma) 129974 27299 hCG15447.3 ADAMDEC1 ADAM-like,
    [Show full text]
  • Integrins: Roles in Cancer Development and As Treatment Targets
    British Journal of Cancer (2004) 90, 561 – 565 & 2004 Cancer Research UK All rights reserved 0007 – 0920/04 $25.00 www.bjcancer.com Minireview Integrins: roles in cancer development and as treatment targets 1 ,1,2 H Jin and J Varner* 1John and Rebecca Moores Comprehensive Cancer Center, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0912, USA; 2Department of Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0912, USA The integrin family of cell adhesion proteins promotes the attachment and migration of cells on the surrounding extracellular matrix (ECM). Through signals transduced upon integrin ligation by ECM proteins or immunoglobulin superfamily molecules, this family of proteins plays key roles in regulating tumour growth and metastasis as well as tumour angiogenesis. Several integrins play key roles in promoting tumour angiogenesis and tumour metastasis. Antagonists of several integrins (a5b1, avb3 and avb5) are now under evaluation in clinical trials to determine their potential as therapeutics for cancer and other diseases. British Journal of Cancer (2004) 90, 561 – 565. doi:10.1038/sj.bjc.6601576 www.bjcancer.com & 2004 Cancer Research UK Keywords: angiogenesis; metastasis; apoptosis; integrin a5b1; integrin avb3 During the last 10 years, novel insights into the mechanisms sequences (e.g., integrin a4b1 recognises EILDV and REDV in that regulate cell survival as well as cell migration and invasion alternatively spliced CS-1 fibronectin). Inhibitors of integrin have led to the development of novel integrin-based therapeutics function include function-blocking monoclonal antibodies, pep- for the treatment of cancer. Several integrins play important tide antagonists and small molecule peptide mimetics matrix roles in promoting cell proliferation, migration and survival (reviewed in Hynes, 1992; Cheresh, 1993).
    [Show full text]
  • The Dynamic Interaction Between Extracellular Matrix Remodeling and Breast Tumor Progression
    cells Review The Dynamic Interaction between Extracellular Matrix Remodeling and Breast Tumor Progression Jorge Martinez 1,* and Patricio C. Smith 2,* 1 Cell Biology Laboratory, INTA, University of Chile, Santiago 7810000, Chile 2 School of Dentistry, Faculty of Medicine, Pontificia Universidad Católica de Chile, Santiago 8330024, Chile * Correspondence: [email protected] (J.M.); [email protected] (P.C.S.); Tel.: + 56-2-2987-1419 (J.M.); +56-2-2354-8400 (P.C.S.) Abstract: Desmoplastic tumors correspond to a unique tissue structure characterized by the abnormal deposition of extracellular matrix. Breast tumors are a typical example of this type of lesion, a property that allows its palpation and early detection. Fibrillar type I collagen is a major component of tumor desmoplasia and its accumulation is causally linked to tumor cell survival and metastasis. For many years, the desmoplastic phenomenon was considered to be a reaction and response of the host tissue against tumor cells and, accordingly, designated as “desmoplastic reaction”. This notion has been challenged in the last decades when desmoplastic tissue was detected in breast tissue in the absence of tumor. This finding suggests that desmoplasia is a preexisting condition that stimulates the development of a malignant phenotype. With this perspective, in the present review, we analyze the role of extracellular matrix remodeling in the development of the desmoplastic response. Importantly, during the discussion, we also analyze the impact of obesity and cell metabolism as critical drivers of tissue remodeling during the development of desmoplasia. New knowledge derived from the dynamic remodeling of the extracellular matrix may lead to novel targets of interest for early diagnosis or therapy in the context of breast tumors.
    [Show full text]
  • Blood Vitronectin Is a Major Activator of LIF and IL-6 in the Brain Through Integrin–FAK and Upar Signaling Matthew P
    © 2018. Published by The Company of Biologists Ltd | Journal of Cell Science (2018) 131, jcs202580. doi:10.1242/jcs.202580 RESEARCH ARTICLE Blood vitronectin is a major activator of LIF and IL-6 in the brain through integrin–FAK and uPAR signaling Matthew P. Keasey1, Cuihong Jia1, Lylyan F. Pimentel1,2, Richard R. Sante1, Chiharu Lovins1 and Theo Hagg1,* ABSTRACT Microglia and astrocytes express the VTN receptors αvβ3 and α β We defined how blood-derived vitronectin (VTN) rapidly and potently v 5 integrin (Herrera-Molina et al., 2012; Kang et al., 2008; activates leukemia inhibitory factor (LIF) and pro-inflammatory Milner, 2009; Welser-Alves et al., 2011). Microglia and astrocytes, interleukin 6 (IL-6) in vitro and after vascular injury in the brain. as well as endothelial cells, are major producers of pro- α in vitro Treatment with VTN (but not fibrinogen, fibronectin, laminin-111 or inflammatory cytokines, such as IL-6 and TNF , and collagen-I) substantially increased LIF and IL-6 within 4 h in after traumatic or ischemic injury to the brain (Banner et al., 1997; C6-astroglioma cells, while VTN−/− mouse plasma was less effective Erta et al., 2012; Lau and Yu, 2001) or upon self-induction by IL-6 than that from wild-type mice. LIF and IL-6 were induced by (Van Wagoner and Benveniste, 1999). IL-6 is a major regulator of a intracerebral injection of recombinant human (rh)VTN in mice, but variety of inflammatory disorders and a target for therapies (Hunter induction seen upon intracerebral hemorrhage was less in VTN−/− and Jones, 2015).
    [Show full text]
  • Cell Adhesion Molecules in Normal Skin and Melanoma
    biomolecules Review Cell Adhesion Molecules in Normal Skin and Melanoma Cian D’Arcy and Christina Kiel * Systems Biology Ireland & UCD Charles Institute of Dermatology, School of Medicine, University College Dublin, D04 V1W8 Dublin, Ireland; [email protected] * Correspondence: [email protected]; Tel.: +353-1-716-6344 Abstract: Cell adhesion molecules (CAMs) of the cadherin, integrin, immunoglobulin, and selectin protein families are indispensable for the formation and maintenance of multicellular tissues, espe- cially epithelia. In the epidermis, they are involved in cell–cell contacts and in cellular interactions with the extracellular matrix (ECM), thereby contributing to the structural integrity and barrier for- mation of the skin. Bulk and single cell RNA sequencing data show that >170 CAMs are expressed in the healthy human skin, with high expression levels in melanocytes, keratinocytes, endothelial, and smooth muscle cells. Alterations in expression levels of CAMs are involved in melanoma propagation, interaction with the microenvironment, and metastasis. Recent mechanistic analyses together with protein and gene expression data provide a better picture of the role of CAMs in the context of skin physiology and melanoma. Here, we review progress in the field and discuss molecular mechanisms in light of gene expression profiles, including recent single cell RNA expression information. We highlight key adhesion molecules in melanoma, which can guide the identification of pathways and Citation: D’Arcy, C.; Kiel, C. Cell strategies for novel anti-melanoma therapies. Adhesion Molecules in Normal Skin and Melanoma. Biomolecules 2021, 11, Keywords: cadherins; GTEx consortium; Human Protein Atlas; integrins; melanocytes; single cell 1213. https://doi.org/10.3390/ RNA sequencing; selectins; tumour microenvironment biom11081213 Academic Editor: Sang-Han Lee 1.
    [Show full text]
  • Ivtigation of the Effects of Mechanical Strain in Human Tenocytes
    Investigation of the effects of Mechanical Strain in Human Tenocytes Eleanor Rachel Jones In partial fulfilment of the requirements for the Degree of Doctor of Philosophy University of East Anglia, Biological Sciences September 2012 This copy of the thesis has been supplied on condition that anyone who consults it is understood to recognise that its copyright rests with the author and that use of any information derived there from must be in accordance with current UK Copyright Law. In addition, any quotation or extract must be included full attribution. Abstract Tendinopathies are a range of diseases characterised by pain and insidious degeneration. Although poorly understood, onset is often associated with physical activity. Metalloproteinases are regulated differentially in tendinopathy causing disruptions in extracellular matrix (ECM) homeostasis. An increase in the anti-inflammatory cytokine TGFβ has also been documented. This project aims to investigate the effect of cyclic tensile strain loading and TGFβ stimulation on protease and ECM protein expression by human tenocytes and begin to characterise the pathway of mechanotransduction. Human tenocytes were seeded at 1.5x106 cells/ml into collagen gels (rat tail type I, 1mg/ml) and stretched using a sinusoidal waveform of 0-5% at 1Hz using the Flexcell FX4000T™ system. Cultures were treated with or without 1ng/ml TGFβ1 or inhibitors of TGFβRI, metalloproteinases, RGD, Mannose-6-phosphate, integrin β1 and a thrombospondin as appropriate. qRT-PCR and a cell based luciferase assay were used to assess RNA and TGFβ activity respectively. The prolonged application of 5% cyclic mechanical strain in a 3D culture system induced an anabolic response in protease and matrix genes.
    [Show full text]
  • Downregulation of Lysyl Oxidase and Lysyl Oxidase-Like Protein 2
    Xu et al. Experimental & Molecular Medicine (2019) 51:20 https://doi.org/10.1038/s12276-019-0211-9 Experimental & Molecular Medicine ARTICLE Open Access Downregulation of lysyl oxidase and lysyl oxidase-like protein 2 suppressed the migration and invasion of trophoblasts by activating the TGF-β/collagen pathway in preeclampsia Xiang-Hong Xu 1,YuanhuiJia1,XinyaoZhou1,DandanXie1, Xiaojie Huang1, Linyan Jia1, Qian Zhou1, Qingliang Zheng1, Xiangyu Zhou1,KaiWang1 and Li-Ping Jin1 Abstract Preeclampsia is a pregnancy-specific disorder that is a major cause of maternal and fetal morbidity and mortality with a prevalence of 6–8% of pregnancies. Although impaired trophoblast invasion in early pregnancy is known to be closely associated with preeclampsia, the underlying mechanisms remain elusive. Here we revealed that lysyl oxidase (LOX) and LOX-like protein 2 (LOXL2) play a critical role in preeclampsia. Our results demonstrated that LOX and LOXL2 expression decreased in preeclamptic placentas. Moreover, knockdown of LOX or LOXL2 suppressed trophoblast cell migration and invasion. Mechanistically, collagen production was induced in LOX-orLOXL2-downregulated trophoblast cells through activation of the TGF-β1/Smad3 pathway. Notably, inhibition of the TGF-β1/Smad3 pathway 1234567890():,; 1234567890():,; 1234567890():,; 1234567890():,; could rescue the defects caused by LOX or LOXL2 knockdown, thereby underlining the significance of the TGF-β1/ Smad3 pathway downstream of LOX and LOXL2 in trophoblast cells. Additionally, induced collagen production and activated TGF-β1/Smad3 were observed in clinical samples from preeclamptic placentas. Collectively, our study suggests that the downregulation of LOX and LOXL2 leading to reduced trophoblast cell migration and invasion through activation of the TGF-β1/Smad3/collagen pathway is relevant to preeclampsia.
    [Show full text]
  • Nephronectin As a Matrix Effector in Cancer
    cancers Review Nephronectin as a Matrix Effector in Cancer Synnøve Norvoll Magnussen 1,* , Jimita Toraskar 2,3 , Elin Hadler-Olsen 1,4, Tonje S. Steigedal 2,3 and Gunbjørg Svineng 1 1 Department of Medical Biology, Faculty of Health Sciences, UiT—The Arctic University of Norway, 9037 Tromsø, Norway; [email protected] (E.H.-O.); [email protected] (G.S.) 2 Department of Clinical and Molecular Medicine, Faculty of Medicine and Health Sciences, Norwegian University of Science and Technology (NTNU), 7491 Trondheim, Norway; [email protected] (J.T.); [email protected] (T.S.S.) 3 Cancer Clinic, St. Olavs Hospital HF, 7006 Trondheim, Norway 4 The Public Dental Health Service Competence Center of Northern Norway, 9271 Tromsø, Norway * Correspondence: [email protected] Simple Summary: The extracellular matrix provides an important scaffold for cells and tissues of multicellular organisms. The scaffold not only provides a secure anchorage point, but also functions as a reservoir for signalling molecules, sequestered and released when necessary. A dysregulated extracellular matrix may therefore modulate cellular behaviour, as seen during cancer progression. The extracellular matrix protein nephronectin was discovered two decades ago and found to regulate important embryonic developmental processes. Loss of either nephronectin or its receptor, integrin α8β1, leads to underdeveloped kidneys. Recent findings show that nephronectin is also dysregulated in breast cancer and plays a role in promoting metastasis. To enable therapeutic intervention, it is important to fully understand the role of nephronectin and its receptors in cancer progression. In Citation: Magnussen, S.N.; this review, we summarise the literature on nephronectin, analyse the structure and domain-related Toraskar, J.; Hadler-Olsen, E.; functions of nephronectin and link these functions to potential roles in cancer progression.
    [Show full text]
  • Structural and Functional Modulation of Perineuronal Nets: in Search of Important Players with Highlight on Tenascins
    cells Review Structural and Functional Modulation of Perineuronal Nets: In Search of Important Players with Highlight on Tenascins Ana Jakovljevi´c 1,*,†, Milena Tuci´c 1,†, Michaela Blažiková 2 , Andrej Koreni´c 1, Yannis Missirlis 3, Vera Stamenkovi´c 1,4 and Pavle Andjus 1 1 Center for Laser Microscopy, Institute for Physiology and Biochemistry “Jean Giaja”, Faculty of Biology, University of Belgrade, 11000 Belgrade, Serbia; [email protected] (M.T.); [email protected] (A.K.); [email protected] or [email protected] (V.S.); [email protected] (P.A.) 2 Light Microscopy Core Facility, Institute of Molecular Genetics CAS, Prague 142 20, Czech Republic; [email protected] 3 Laboratory of Biomechanics and Biomedical Engineering, Department of Mechanical Engineering and Aeronautics, University of Patras, 26504 Patras, Greece; [email protected] 4 Center for Integrative Brain Research, Seattle Children’s Research Institute, 1900 9th Ave, Seattle, WA 98125, USA * Correspondence: [email protected] † These authors contributed equally to this work. Abstract: The extracellular matrix (ECM) of the brain plays a crucial role in providing optimal conditions for neuronal function. Interactions between neurons and a specialized form of ECM, perineuronal nets (PNN), are considered a key mechanism for the regulation of brain plasticity. Such an assembly of interconnected structural and regulatory molecules has a prominent role in Citation: Jakovljevi´c,A.; Tuci´c,M.; the control of synaptic plasticity. In this review, we discuss novel ways of studying the interplay Blažiková, M.; Koreni´c,A.; Missirlis, Y.; Stamenkovi´c,V.; Andjus, P.
    [Show full text]