Myosin IIA/IIB restrict adhesive and protrusive signaling to generate front–back polarity in migrating cells

Vicente‐Manzanares, Miguel; Litwa, Karen; Bachir, Alexia I.; Whitmore, Leanna; Horwitz, Alan Rick

doi:10.1083/jcb.201012159

Cited by 136 publications

(175 citation statements)

References 71 publications

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“…In this context, an interesting observation is that in the triangular cells all internal F-actin bundles (except those along the edges) are crucially dependent on MIIA as the depletion of MIIA results in almost complete removal of all large bundles. This is similar to unconstrained cells where bundling by other myosin isoform, MIIB, is dependent on MIIA and efficient MIIA knockdown results in complete depletion of all F-actin bundles (Vicente-Manzanares et al, 2011). Furthermore, the result that either depletion of Myosin IIA or Myosin IIB by themselves depletes all F-actin bundles are consistent with recent findings that in fibroblast cells MIIA initiates bundle formation whereas MIIB crosslinks and strengthens MIIA initiated bundles (Vicente-Manzanares et al, 2011).…”

Section: Discussionsupporting

confidence: 80%

“…Most importantly, the distribution of MT growth trajectories in cells treated with the ROCK inhibitor was nearly identical to the computationally generated distributions for the random/unguided growth case ( , 2007). In motile cells, myosin IIA initiates Factin bundle formation while myosin IIB stabilizes these F-actin bundles (Wei and Adelstein, 2000;Vicente-Manzanares et al, 2011). To test the role of myosin IIA (MIIA) crosslinked bundles in MT guidance, we depleted MIIA from Rat2 cells by means of RNA interference.…”

Section: Cell-wide Correlation Of F-actin Bundles and Microtubule Tramentioning

confidence: 99%

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Microtubule guidance tested through controlled cell geometry

Huda

Soh

Pilans

et al. 2012

Journal of Cell Science

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SummaryIn moving cells dynamic microtubules (MTs) target and disassemble substrate adhesion sites (focal adhesions; FAs) in a process that enables the cell to detach from the substrate and propel itself forward. The short-range interactions between FAs and MT plus ends have been observed in several experimental systems, but the spatial overlap of these structures within the cell has precluded analysis of the putative long-range mechanisms by which MTs growing through the cell body reach FAs in the periphery of the cell. In the work described here cell geometry was controlled to remove the spatial overlap of cellular structures thus allowing for unambiguous observation of MT guidance. Specifically, micropatterning of living cells was combined with high-resolution in-cell imaging and gene product depletion by means of RNA interference to study the long-range MT guidance in quantitative detail. Cells were confined on adhesive triangular microislands that determined cell shape and ensured that FAs localized exclusively at the vertices of the triangular cells. It is shown that initial MT nucleation at the centrosome is random in direction, while the alignment of MT trajectories with the targets (i.e. FAs at vertices) increases with an increasing distance from the centrosome, indicating that MT growth is a non-random, guided process. The guided MT growth is dependent on the presence of FAs at the vertices. The depletion of either myosin IIA or myosin IIB results in depletion of F-actin bundles and spatially unguided MT growth. Taken together our findings provide quantitative evidence of a role for long-range MT guidance in MT targeting of FAs.

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Section: Discussionsupporting

confidence: 80%

Section: Cell-wide Correlation Of F-actin Bundles and Microtubule Tramentioning

confidence: 99%

Microtubule guidance tested through controlled cell geometry

Huda

Soh

Pilans

et al. 2012

Journal of Cell Science

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“…Importantly, accumulating evidence suggests that myosin II contractility mediates the establishment of polarity in migrating cells through a mechanism whereby the local depletion/dislodging of the Rho guanine nucleotide exchange factor b-Pix (and others) decreases Rac activation, limits protrusiveness, and promotes RhoAdependent adhesion maturation (Kuo et al, 2011;VicenteManzanares et al, 2011). In line with these observations, our results suggest that the Nck deficiency phenocopies -to a significant extent -the inactivation/depletion of myosin II which is characterized by increased protrusiveness, reduced adhesion maturation, and loss of cell polarity (Vicente-Manzanares et al, 2011;Vicente-Manzanares et al, 2007).…”

Section: Discussionsupporting

confidence: 77%

Nck enables directional cell migration through the coordination of polarized membrane protrusion with adhesion dynamics

Chaki¹,

Barhoumi²,

Berginski³

et al. 2013

Journal of Cell Science

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SummaryDirectional migration requires the coordination of cytoskeletal changes essential for cell polarization and adhesion turnover. Extracellular signals that alter tyrosine phosphorylation drive directional migration by inducing reorganization of the actin cytoskeleton. It is recognized that Nck is an important link between tyrosine phosphorylation and actin dynamics; however, the role of Nck in cytoskeletal remodeling during directional migration and the underlying molecular mechanisms remain largely undetermined. In this study, a combination of molecular genetics and quantitative live cell microscopy was used to show that Nck is essential in the establishment of front-back polarity and directional migration of endothelial cells. Time-lapse differential interference contrast and total internal reflection fluorescence microscopy showed that Nck couples the formation of polarized membrane protrusions with their stabilization through the assembly and maturation of cell-substratum adhesions. Measurements by atomic force microscopy showed that Nck also modulates integrin a5b1-fibronectin adhesion force and cell stiffness. Fluorescence resonance energy transfer imaging revealed that Nck depletion results in delocalized and increased activity of Cdc42 and Rac. By contrast, the activity of RhoA and myosin II phosphorylation were reduced by Nck knockdown. Thus, this study identifies Nck as a key coordinator of cytoskeletal changes that enable cell polarization and directional migration, which are crucial processes in development and disease.

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“…In contrast to this variable importance of NMIIA for 2D migration, evidence is consistent (i) in that the NMIIA isoform shows strong recruitment to anterior protrusions during 2D migration and (ii) that NMIIA has a strong and isoform-specific role in stabilizing nascent focal adhesions at the anterior of cells migrating in 2D (14,30,(43)(44)(45). We hypothesized that the limited and variable impact of NMIIA depletion on migration rate in 2D studies may relate to the lack of resistance a cell encounters during 2D migration on rigid surfaces.…”

Section: Discussionmentioning

confidence: 99%

Myosin IIA Heavy Chain Phosphorylation Mediates Adhesion Maturation and Protrusion in Three Dimensions

Rai

Thomas

Beach

et al. 2017

Journal of Biological Chemistry

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Non-muscle myosin II (NMII) is a conserved force-producing cytoskeletal enzyme with important but poorly understood roles in cell migration. To investigate myosin heavy chain (MHC) phosphorylation roles in 3D migration, we expressed GFP-tagged NMIIA wild-type or mutant constructs in cells depleted of endogenous NMIIA protein. We find that individual mutation or double mutation of Ser-1916 or Ser-1943 to alanine potently blocks recruitment of GFP-NM-IIA filaments to leading edge protrusions in 2D, and this in turn blocks maturation of anterior focal adhesions. When placed in 3D collagen gels, cells expressing wild-type GFP MHC-IIA behave like parental cells, displaying robust and active formation and retraction of protrusions. However, cells depleted of NMIIA or cells expressing the mutant GFP MHC-IIA display severe defects in invasion and in stabilizing protrusions in 3D. These studies reveal an NMIIA-specific role in 3D invasion that requires competence for NMIIA phosphorylation at Ser-1916 and Ser-1943. In sum, these results demonstrate a critical and previously unrecognized role for NMIIA phosphorylation in 3D invasion.

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Myosin IIA/IIB restrict adhesive and protrusive signaling to generate front–back polarity in migrating cells

Abstract: Myosin IIA and IIB synergistically generate front–back polarity through their effects on actomyosin bundling formation and stability, and adhesion maturation, which are mediated by localized Rac GEF depletion.

Cited by 136 publications

References 71 publications

Microtubule guidance tested through controlled cell geometry

Microtubule guidance tested through controlled cell geometry

Nck enables directional cell migration through the coordination of polarized membrane protrusion with adhesion dynamics

Myosin IIA Heavy Chain Phosphorylation Mediates Adhesion Maturation and Protrusion in Three Dimensions

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