David Cornu1,2, Norbert Bakalara3,4, Emilie Marhuenda5,6,7, Christine Fabre8,9, Cunjie Zhang10,11, Martà Martin-Fernandez12, Thomas Iskratsch13, Ali Saleh8, Luc Bauchet8, Julien Cambedouzou14,9, Jean-Philippe Hugnot8, Hugues Duffau8, James W Dennis10,11. 1. Institut Européen des Membranes, IEM, UMR 5635, University of Montpellier, ENSCM, CNRS, Montpellier, France. david.cornu@enscm.fr. 2. École nationale supérieure de chimie de Montpellier, ENSCM, 240 Avenue du Professeur Emile Jeanbrau, 34090, Montpellier, France. david.cornu@enscm.fr. 3. Institut des Neurosciences de Montpellier (INM) U-1051, University of Montpellier, 80 rue Augustin Fliche, Hôpital Saint-Eloi, 34091, Montpellier, Cedex 5, France. norbert.bakalara@enstbb.fr. 4. École nationale supérieure de chimie de Montpellier, ENSCM, 240 Avenue du Professeur Emile Jeanbrau, 34090, Montpellier, France. norbert.bakalara@enstbb.fr. 5. Institut des Neurosciences de Montpellier (INM) U-1051, University of Montpellier, 80 rue Augustin Fliche, Hôpital Saint-Eloi, 34091, Montpellier, Cedex 5, France. e.marhuenda@qmul.ac.uk. 6. School of Engineering and Materials Science, Queen Mary University of London, London, E1 4NS, UK. e.marhuenda@qmul.ac.uk. 7. Institut Européen des Membranes, IEM, UMR 5635, University of Montpellier, ENSCM, CNRS, Montpellier, France. e.marhuenda@qmul.ac.uk. 8. Institut des Neurosciences de Montpellier (INM) U-1051, University of Montpellier, 80 rue Augustin Fliche, Hôpital Saint-Eloi, 34091, Montpellier, Cedex 5, France. 9. École nationale supérieure de chimie de Montpellier, ENSCM, 240 Avenue du Professeur Emile Jeanbrau, 34090, Montpellier, France. 10. Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, 600 University Ave., Toronto, ON, M5G 1X5, Canada. 11. Department of Molecular Genetics, and Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, M5S 1A8, Canada. 12. Institut Charles Coulomb, UMR 5221, University of Montpellier, CNRS, Montpellier, France. 13. School of Engineering and Materials Science, Queen Mary University of London, London, E1 4NS, UK. 14. Institut Européen des Membranes, IEM, UMR 5635, University of Montpellier, ENSCM, CNRS, Montpellier, France.
Abstract
BACKGROUND: Glioblastomas stem-like cells (GSCs) by invading the brain parenchyma, remains after resection and radiotherapy and the tumoral microenvironment become stiffer. GSC invasion is reported as stiffness sensitive and associated with altered N-glycosylation pattern. Glycocalyx thickness modulates integrins mechanosensing, but details remain elusive and glycosylation enzymes involved are unknown. Here, we studied the association between matrix stiffness modulation, GSC migration and MGAT5 induced N-glycosylation in fibrillar 3D context. METHOD: To mimic the extracellular matrix fibrillar microenvironments, we designed 3D-ex-polyacrylonitrile nanofibers scaffolds (NFS) with adjustable stiffnesses by loading multiwall carbon nanotubes (MWCNT). GSCs neurosphere were plated on NFSs, allowing GSCs migration and MGAT5 was deleted using CRISPR-Cas9. RESULTS: We found that migration of GSCs was maximum at 166 kPa. Migration rate was correlated with cell shape, expression and maturation of focal adhesion (FA), Epithelial to Mesenchymal Transition (EMT) proteins and (β1,6) branched N-glycan binding, galectin-3. Mutation of MGAT5 in GSC inhibited N-glycans (β1-6) branching, suppressed the stiffness dependence of migration on 166 kPa NFS as well as the associated FA and EMT protein expression. CONCLUSION: MGAT5 catalysing multibranched N-glycans is a critical regulators of stiffness induced invasion and GSCs mechanotransduction, underpinning MGAT5 as a serious target to treat cancer.
BACKGROUND:Glioblastomas stem-like cells (GSCs) by invading the brain parenchyma, remains after resection and radiotherapy and the tumoral microenvironment become stiffer. GSC invasion is reported as stiffness sensitive and associated with altered N-glycosylation pattern. Glycocalyx thickness modulates integrins mechanosensing, but details remain elusive and glycosylation enzymes involved are unknown. Here, we studied the association between matrix stiffness modulation, GSC migration and MGAT5 induced N-glycosylation in fibrillar 3D context. METHOD: To mimic the extracellular matrix fibrillar microenvironments, we designed 3D-ex-polyacrylonitrile nanofibers scaffolds (NFS) with adjustable stiffnesses by loading multiwall carbon nanotubes (MWCNT). GSCs neurosphere were plated on NFSs, allowing GSCsmigration and MGAT5 was deleted using CRISPR-Cas9. RESULTS: We found that migration of GSCs was maximum at 166 kPa. Migration rate was correlated with cell shape, expression and maturation of focal adhesion (FA), Epithelial to Mesenchymal Transition (EMT) proteins and (β1,6) branched N-glycan binding, galectin-3. Mutation of MGAT5 in GSC inhibited N-glycans (β1-6) branching, suppressed the stiffness dependence of migration on 166 kPa NFS as well as the associated FA and EMT protein expression. CONCLUSION:MGAT5 catalysing multibranched N-glycans is a critical regulators of stiffness induced invasion and GSCs mechanotransduction, underpinning MGAT5 as a serious target to treat cancer.
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