title: Lamellipodial tension, not integrin/ligand binding, is the crucial factor to realise integrin activation and cell migration.
abstract: The molecular clutch (MC) model proposes that actomyosin-driven force transmission permits integrin-dependent cell migration. To investigate the MC, we introduced diverse talin (TLN) and integrin variants into Flp-In™ T-Rex™ HEK293 cells stably expressing uPAR. Vitronectin variants served as substrate providing uPAR-mediated cell adhesion and optionally integrin binding. This particular system allowed us to selectively analyse key MC proteins and interactions, effectively from the extracellular matrix substrate to intracellular f-actin, and to therewith study mechanobiological aspects of MC engagement also uncoupled from integrin/ligand binding. With this experimental approach, we found that for the initial PIP2-dependent membrane/TLN/f-actin linkage and persistent lamellipodia formation the C-terminal TLN actin binding site (ABS) is dispensable. The establishment of an adequate MC-mediated lamellipodial tension instead depends predominantly on the coupling of this C-terminal TLN ABS to the actomyosin-driven retrograde actin flow force. This lamellipodial tension is crucial for full integrin activation eventually determining integrin-dependent cell migration. In the integrin/ligand-independent condition the frictional membrane resistance participates to these processes. Integrin/ligand binding can also contribute but is not necessarily required.
authors.affiliation: Unit of Cell Matrix Signalling, IFOM, The FIRC Institute of Molecular Oncology, Via Adamello 16, 20139 Milan, Italy; CIMaINa (Interdisciplinary Centre for Nanostructured Material and Interfaces) and Department of Physics, Università degli Studi di Milano, Via Celoria 16, 20133 Milano, Italy. Electronic address: carsten.schulte@unimi.it.
authors.forename: Carsten
authors.initials: C
authors.lastname: Schulte
authors.affiliation: Unit of Cell Matrix Signalling, IFOM, The FIRC Institute of Molecular Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Gian Maria Sarra
authors.initials: GM
authors.lastname: Ferraris
authors.affiliation: Imaging Unit, IFOM, The FIRC Institute of Molecular Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Amanda
authors.initials: A
authors.lastname: Oldani
authors.affiliation: CIMaINa (Interdisciplinary Centre for Nanostructured Material and Interfaces) and Department of Physics, Università degli Studi di Milano, Via Celoria 16, 20133 Milano, Italy.
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Galluzzi
authors.affiliation: CIMaINa (Interdisciplinary Centre for Nanostructured Material and Interfaces) and Department of Physics, Università degli Studi di Milano, Via Celoria 16, 20133 Milano, Italy.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Podestà
authors.affiliation: CIMaINa (Interdisciplinary Centre for Nanostructured Material and Interfaces) and Department of Physics, Università degli Studi di Milano, Via Celoria 16, 20133 Milano, Italy.
authors.forename: Luca
authors.initials: L
authors.lastname: Puricelli
authors.affiliation: Unit of Cell Matrix Signalling, IFOM, The FIRC Institute of Molecular Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Valentina
authors.initials: V
authors.lastname: de Lorenzi
authors.affiliation: CIMaINa (Interdisciplinary Centre for Nanostructured Material and Interfaces) and Department of Physics, Università degli Studi di Milano, Via Celoria 16, 20133 Milano, Italy.
authors.forename: Cristina
authors.initials: C
authors.lastname: Lenardi
authors.affiliation: CIMaINa (Interdisciplinary Centre for Nanostructured Material and Interfaces) and Department of Physics, Università degli Studi di Milano, Via Celoria 16, 20133 Milano, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Milani
authors.affiliation: Unit of Cell Matrix Signalling, IFOM, The FIRC Institute of Molecular Oncology, Via Adamello 16, 20139 Milan, Italy.
authors.forename: Nicolai
authors.initials: N
authors.lastname: Sidenius
mesh_terms: D000199:Actins / Q000378:metabolism; D002465:Cell Movement / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D022001:Focal Adhesions / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D016023:Integrins / Q000378:metabolism*; D008024:Ligands; D018613:Microscopy, Confocal; D011485:Protein Binding / Q000502:physiology; D011554:Pseudopodia / Q000378:metabolism*; D016608:Talin / Q000378:metabolism
chemical_list: D000199:Actins; D016023:Integrins; D008024:Ligands; D016608:Talin
keywords: AFM; Focal adhesion; Mechanotransduction; Molecular clutch; PIP(2); Talin; uPAR