title: Paxillin mediates sensing of physical cues and regulates directional cell motility by controlling lamellipodia positioning.
abstract: Physical interactions between cells and the extracellular matrix (ECM) guide directional migration by spatially controlling where cells form focal adhesions (FAs), which in turn regulate the extension of motile processes. Here we show that physical control of directional migration requires the FA scaffold protein paxillin. Using single-cell sized ECM islands to constrain cell shape, we found that fibroblasts cultured on square islands preferentially activated Rac and extended lamellipodia from corner, rather than side regions after 30 min stimulation with PDGF, but that cells lacking paxillin failed to restrict Rac activity to corners and formed small lamellipodia along their entire peripheries. This spatial preference was preceded by non-spatially constrained formation of both dorsal and lateral membrane ruffles from 5-10 min. Expression of paxillin N-terminal (paxN) or C-terminal (paxC) truncation mutants produced opposite, but complementary, effects on lamellipodia formation. Surprisingly, pax-/- and paxN cells also formed more circular dorsal ruffles (CDRs) than pax+ cells, while paxC cells formed fewer CDRs and extended larger lamellipodia even in the absence of PDGF. In a two-dimensional (2D) wound assay, pax-/- cells migrated at similar speeds to controls but lost directional persistence. Directional motility was rescued by expressing full-length paxillin or the N-terminus alone, but paxN cells migrated more slowly. In contrast, pax-/- and paxN cells exhibited increased migration in a three-dimensional (3D) invasion assay, with paxN cells invading Matrigel even in the absence of PDGF. These studies indicate that paxillin integrates physical and chemical motility signals by spatially constraining where cells will form motile processes, and thereby regulates directional migration both in 2D and 3D. These findings also suggest that CDRs may correspond to invasive protrusions that drive cell migration through 3D extracellular matrices.
authors.affiliation: Vascular Biology Program, Department of Pathology, Children's Hospital Boston and Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Julia E
authors.initials: JE
authors.lastname: Sero
authors.forename: Charles K
authors.initials: CK
authors.lastname: Thodeti
authors.forename: Akiko
authors.initials: A
authors.lastname: Mammoto
authors.forename: Chris
authors.initials: C
authors.lastname: Bakal
authors.forename: Sheila
authors.initials: S
authors.lastname: Thomas
authors.forename: Donald E
authors.initials: DE
authors.lastname: Ingber
mesh_terms: D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects; D002452:Cell Count; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects; D002645:Chickens; D003094:Collagen / Q000378:metabolism; D004338:Drug Combinations; D004622:Embryo, Mammalian / Q000166:cytology; D005109:Extracellular Matrix / Q000187:drug effects / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D022001:Focal Adhesions / Q000187:drug effects / Q000378:metabolism; D055786:Gene Knockout Techniques; D006801:Humans; D007797:Laminin / Q000378:metabolism; D051379:Mice; D009154:Mutation / Q000235:genetics; D051419:Paxillin / Q000737:chemistry / Q000378:metabolism*; D010641:Phenotype; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism; D011554:Pseudopodia / Q000187:drug effects / Q000378:metabolism*; D013997:Time Factors; D016596:Vinculin / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000378:metabolism
chemical_list: D004338:Drug Combinations; D007797:Laminin; D051419:Paxillin; D010982:Platelet-Derived Growth Factor; D011509:Proteoglycans; C059485:matrigel; D016596:Vinculin; D003094:Collagen; D020744:rac GTP-Binding Proteins