scholarly journals The Dioxin Receptor Regulates the Constitutive Expression of the Vav3 Proto-Oncogene and Modulates Cell Shape and Adhesion

2009 ◽  
Vol 20 (6) ◽  
pp. 1715-1727 ◽  
Author(s):  
Jose M. Carvajal-Gonzalez ◽  
Sonia Mulero-Navarro ◽  
Angel Carlos Roman ◽  
Vincent Sauzeau ◽  
Jaime M. Merino ◽  
...  
Keyword(s):  
Cell Shape ◽  
Constitutive Expression ◽  
Focal Adhesions ◽  
Stress Fibers ◽  
Cell Area ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Null Phenotype ◽  
And Migration ◽  
Dioxin Receptor

The dioxin receptor (AhR) modulates cell plasticity and migration, although the signaling involved remains unknown. Here, we report a mechanism that integrates AhR into these cytoskeleton-related functions. Immortalized and mouse embryonic fibroblasts lacking AhR (AhR−/−) had increased cell area due to spread cytoplasms that reverted to wild-type morphology upon AhR re-expression. The AhR-null phenotype included increased F-actin stress fibers, depolarized focal adhesions, and enhanced spreading and adhesion. The cytoskeleton alterations of AhR−/− cells were due to down-regulation of constitutive Vav3 expression, a guanosine diphosphate/guanosine triphosphate exchange factor for Rho/Rac GTPases and a novel transcriptional target of AhR. AhR was recruited to the vav3 promoter and maintained constitutive mRNA expression in a ligand-independent manner. Consistently, AhR−/− fibroblasts had reduced Rac1 activity and increased activation of the RhoA/Rho kinase (Rock) pathway. Pharmacological inhibition of Rac1 shifted AhR+/+ fibroblasts to the null phenotype, whereas Rock inhibition changed AhR-null cells to the AhR+/+ morphology. Knockdown of vav3 transcripts by small interfering RNA induced cytoskeleton defects and changes in adhesion and spreading mimicking those of AhR-null cells. Moreover, vav3−/− MEFs, as AhR−/− mouse embryonic fibroblasts, had increased cell area and enhanced stress fibers. By modulating Vav3-dependent signaling, AhR could regulate cell shape, adhesion, and migration under physiological conditions and, perhaps, in certain pathological states.

scholarly journals Appl1andAppl2are Expendable for Mouse Development But Are Essential for HGF-Induced Akt Activation and Migration in Mouse Embryonic Fibroblasts

2015 ◽  
Vol 231 (5) ◽  
pp. 1142-1150 ◽  
Author(s):  
Yinfei Tan ◽  
Xiaoban Xin ◽  
Francis J. Coffey ◽  
David L. Wiest ◽  
Lily Q. Dong ◽  
...  
Keyword(s):  
Mouse Development ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Akt Activation ◽  
And Migration

Active FHOD1 promotes the formation of functional actin stress fibers

2019 ◽  
Vol 476 (20) ◽  
pp. 2953-2963 ◽  
Author(s):  
Xuemeng Shi ◽  
Shuangshuang Zhao ◽  
Jinping Cai ◽  
Gary Wong ◽  
Yaming Jiu
Keyword(s):  
Myosin Ii ◽  
Critical Role ◽  
Active Form ◽  
Focal Adhesions ◽  
Stress Fibers ◽  
Cell Periphery ◽  
Physiological Relevance ◽  
Human Osteosarcoma Cells ◽  
And Migration ◽  
Actin Stress Fibers

Abstract The formin FHOD1 acts as a nucleating, capping and bundling protein of actin filaments. In cells, release from the C-terminal diaphanous autoregulatory domain (DAD) of FHOD1 stimulates the protein into the active form. However, the cellular physiological relevance of active form FHOD1 and the phenotypic regulation by FHOD1 depletion are not completely understood. Here, we show that in contrast with the cytosolic diffused expression of auto-inhibited FHOD1, active FHOD1 by C-terminal truncation was recruited into all three types of actin stress fibers in human osteosarcoma cells. Notably, the recruited active FHOD1 was more incorporated with myosin II than α-actinin, and associated with both naïve and mature focal adhesions. Active FHOD1 displayed faster turnover than actin molecules on ventral stress fibers. Moreover, we witnessed the emergence of active FHOD1 from the cell periphery, which subsequently moved centripetally together with transverse arcs. Furthermore, FHOD1 knockdown resulted in defective maturation of actomyosin bundles and subsequently longer non-contractile dorsal stress fibers, whereas the turnover of both actin and myosin II were maintained normally. Importantly, the loss of FHOD1 led to slower actin centripetal flow, resulting in abnormal cell spreading and migration defects. Taken together, these results reveal a critical role of FHOD1 in temporal- and spatial- control of the morphology and dynamics of functional actin stress fibers during variable cell behavior.

scholarly journals Translocation of CrkL to Focal Adhesions Mediates Integrin-Induced Migration Downstream of Src Family Kinases

2003 ◽  
Vol 23 (8) ◽  
pp. 2883-2892 ◽  
Author(s):  
Leiming Li ◽  
Deborah L. Guris ◽  
Masaya Okura ◽  
Akira Imamoto
Keyword(s):  
G Proteins ◽  
Focal Adhesions ◽  
Src Family Kinases ◽  
Adapter Protein ◽  
Related Protein ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Src Signaling ◽  
Downstream Mediator

ABSTRACT The adapter protein Crk-Like (CrkL) can associate with the Src substrate p130 Cas (Cas). The biological role of CrkL downstream of Cas, however, has been largely obscure. Consistent with the ability of CrkL to biochemically associate with Cas, we found that Src triggers translocation of CrkL to focal adhesions (FAs) in a manner dependent on Cas. Forced localization of CRKL to FAs (FA-CRKL) by itself was sufficient to induce activation of Rac1 and Cdc42 and rescued haptotaxis defects of mouse embryonic fibroblasts (MEFs) lacking Src, Yes, and Fyn, three broadly expressed Src family members required for integrin-induced migration. Consistent with Rac1 activation, FA-CRKL induced cotranslocation of a Rac1 activator, Dock1, to focal adhesions. These results therefore indicate a role for CrkL in mediating Src signaling by activating small G proteins at focal adhesions. Furthermore, MEFs lacking CrkL show impaired integrin-induced migration despite expression of a closely related protein, Crk-II, in these cells. These results therefore provide formal evidence that CrkL plays a specific role in integrin-induced migration as a downstream mediator of Src.

scholarly journals Replacing nonmuscle myosin 2A with myosin 2C1 permits gastrulation but not placenta vascular development in mice

2018 ◽  
Vol 29 (19) ◽  
pp. 2326-2335
Author(s):  
Yingfan Zhang ◽  
Chengyu Liu ◽  
Robert S. Adelstein ◽  
Xuefei Ma
Keyword(s):  
Homologous Recombination ◽  
Focal Adhesions ◽  
Nonmuscle Myosin ◽  
Developmentally Delayed ◽  
Mouse Development ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Heavy Chains ◽  
Vascular Formation

Three paralogues of nonmuscle myosin 2 (NM 2A, 2B, and 2C) are expressed in mammals, and the heavy chains are the products of three different genes (Myh9, Myh10, and Myh14, respectively). NM 2A and 2B are essential for mouse development, while 2C is not. Studies on NM 2C are limited and the in vivo function of this paralogue is not clear. Using homologous recombination, cDNA encoding nonmuscle myosin heavy chain 2C1 fused with GFP was introduced into the first coding exon of Myh9, replacing NM 2A expression with NM 2C1 expression in mice. In contrast to A–/A– embryos, which die by embryonic day (E) 6.5, AC1*gfp/AC1*gfp embryos survive through E8.5, demonstrating that NM 2C1 can support mouse development beyond gastrulation. At E9.5 and E10.5, however, AC1*gfp/AC1*gfp embryos are developmentally delayed, with abnormalities in placental vascular formation. The defect in vascular formation is confirmed in allantois explants from AC1*gfp/AC1*gfp embryos. Thus, NM 2C1 cannot support normal placental vascular formation. In addition, AC1*gfp/AC1*gfp mouse embryonic fibroblasts (MEFs) migrate rapidly but with impaired persistence and develop smaller, less mature focal adhesions than A+/A+ MEFs. This is attributed to enhanced NM 2C1 actomyosin stability and different NM 2C1 subcellular localization than in NM 2A.

scholarly journals Protein 4.1G Regulates Cell Adhesion, Spreading, and Migration of Mouse Embryonic Fibroblasts through the β1 Integrin Pathway

2015 ◽  
Vol 291 (5) ◽  
pp. 2170-2180 ◽  
Author(s):  
Lixiang Chen ◽  
Ting Wang ◽  
Yaomei Wang ◽  
Jingxin Zhang ◽  
Yuanming Qi ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Β1 Integrin ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
And Migration

scholarly journals β-Actin specifically controls cell growth, migration, and the G-actin pool

2011 ◽  
Vol 22 (21) ◽  
pp. 4047-4058 ◽  
Author(s):  
Tina M. Bunnell ◽  
Brandon J. Burbach ◽  
Yoji Shimizu ◽  
James M. Ervasti
Keyword(s):  
Focal Adhesions ◽  
Whole Body ◽  
Filamentous Actin ◽  
Embryonic Fibroblasts ◽  
Actin Isoforms ◽  
Cellular Processes ◽  
Expression Of Genes ◽  
Primary Mouse ◽  
And Migration ◽  
Severe Growth

Ubiquitously expressed β-actin and γ-actin isoforms play critical roles in most cellular processes; however, their unique contributions are not well understood. We generated whole-body β-actin–knockout (Actb−/−) mice and demonstrated that β-actin is required for early embryonic development. Lethality of Actb−/−embryos correlated with severe growth impairment and migration defects in β-actin–knockout primary mouse embryonic fibroblasts (MEFs) that were not observed in γ-actin–null MEFs. Migration defects were associated with reduced membrane protrusion dynamics and increased focal adhesions. We also identified migration defects upon conditional ablation of β-actin in highly motile T cells. Of great interest, ablation of β-actin altered the ratio of globular actin (G-actin) to filamentous actin in MEFs, with corresponding changes in expression of genes that regulate the cell cycle and motility. These data support an essential role for β-actin in regulating cell migration and gene expression through control of the cellular G-actin pool.

scholarly journals Gab2 regulates cytoskeletal organization and migration of mammary epithelial cells by modulating RhoA activation

2011 ◽  
Vol 22 (1) ◽  
pp. 105-116 ◽  
Author(s):  
Maria Teresa Herrera Abreu ◽  
William E. Hughes ◽  
Katarina Mele ◽  
Ruth J. Lyons ◽  
Danny Rickwood ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cancer Progression ◽  
Mammary Epithelial Cells ◽  
Focal Adhesions ◽  
Metastatic Potential ◽  
Mammary Epithelial ◽  
Stress Fibers ◽  
Membrane Recruitment ◽  
Rhoa Activation ◽  
And Migration

The docking protein Gab2 is overexpressed in several human malignancies, including breast cancer, and is associated with increased metastatic potential. Here we report that Gab2 overexpression in MCF-10A mammary epithelial cells led to delayed cell spreading, a decrease in stress fibers and mature focal adhesions, and enhanced cell migration. Expression of a Gab2 mutant uncoupled from 14-3-3-mediated negative feedback (Gab22×A) led to a more mesenchymal morphology and acquisition of invasive potential. Expression of either Gab2 or Gab22×A led to decreased activation of RhoA, but only the latter increased levels of Rac-GTP. Expression of constitutively active RhoA in MCF-10A/Gab2 cells restored stress fibers and focal adhesions, indicating that Gab2 signals upstream of RhoA to suppress these structures. Mutation of the two Shp2-binding sites to phenylalanine (Gab2ΔShp2) markedly reduced the effects of Gab2 on cellular phenotype and RhoA activation. Expression of Gab2 or Gab22×A, but not Gab2ΔShp2, promoted Vav2 phosphorylation and plasma membrane recruitment of p190A RhoGAP. Knockdown of p190A RhoGAP reversed Gab2-mediated effects on stress fibers and focal adhesions. The identification of a novel pathway downstream of Gab2 involving negative regulation of RhoA by p190A RhoGAP sheds new light on the role of Gab2 in cancer progression.

scholarly journals Insights into In Vitro Wound Closure on Two Biopolyesters—Polylactide and Polyhydroxyoctanoate

Materials ◽  
2020 ◽  
Vol 13 (12) ◽  
pp. 2793 ◽  
Author(s):  
Tomasz Witko ◽  
Daria Solarz ◽  
Karolina Feliksiak ◽  
Katarzyna Haraźna ◽  
Zenon Rajfur ◽  
...  
Keyword(s):  
Cell Shape ◽  
Wound Closure ◽  
Cellular Migration ◽  
Wound Dressings ◽  
Embryonic Fibroblasts ◽  
Cell Processes ◽  
Migration Strategies ◽  
And Migration ◽  
Microscopic Techniques

Two bio-based polymers have been compared in this study, namely: polylactide (PLA) and polyhydroxyoctanoate (PHO). Due to their properties such as biocompatibility, and biointegrity they are considered to be valuable materials for medical purposes, i.e., creating scaffolds or wound dressings. Presented biopolymers were investigated for their impact on cellular migration strategies of mouse embryonic fibroblasts (MEF) 3T3 cell line. Advanced microscopic techniques, including confocal microscopy and immunofluorescent protocols, enabled the thorough analysis of the cell shape and migration. Application of wound healing assay combined with dedicated software allowed us to perform quantitative analysis of wound closure dynamics. The outcome of the experiments demonstrated that the wound closure dynamics for PLA differs from PHO. Single fibroblasts grown on PLA moved 1.5-fold faster, than those migrating on the PHO surface. However, when a layer of cells was considered, the wound closure was by 4.1 h faster for PHO material. The accomplished work confirms the potential of PLA and PHO as excellent candidates for medical applications, due to their properties that propagate cell migration, vitality, and proliferation—essential cell processes in the healing of damaged tissues.

scholarly journals Lipid binding promotes oligomerization and focal adhesion activity of vinculin

2014 ◽  
Vol 207 (5) ◽  
pp. 643-656 ◽  
Author(s):  
Krishna Chinthalapudi ◽  
Erumbi S. Rangarajan ◽  
Dipak N. Patil ◽  
Eric M. George ◽  
David T. Brown ◽  
...  
Keyword(s):  
Cell Migration ◽  
Focal Adhesion ◽  
Lipid Binding ◽  
Stress Fibers ◽  
Actin Binding ◽  
Null Mouse ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Growth And Survival ◽  
Adhesion Activity

Adherens junctions (AJs) and focal adhesion (FA) complexes are necessary for cell migration and morphogenesis, and for the development, growth, and survival of all metazoans. Vinculin is an essential regulator of both AJs and FAs, where it provides links to the actin cytoskeleton. Phosphatidylinositol 4,5-bisphosphate (PIP2) affects the functions of many targets, including vinculin. Here we report the crystal structure of vinculin in complex with PIP2, which revealed that PIP2 binding alters vinculin structure to direct higher-order oligomerization and suggests that PIP2 and F-actin binding to vinculin are mutually permissive. Forced expression of PIP2-binding–deficient mutants of vinculin in vinculin-null mouse embryonic fibroblasts revealed that PIP2 binding is necessary for maintaining optimal FAs, for organization of actin stress fibers, and for cell migration and spreading. Finally, photobleaching experiments indicated that PIP2 binding is required for the control of vinculin dynamics and turnover in FAs. Thus, through oligomerization, PIP2 directs a transient vinculin sequestration at FAs that is necessary for proper FA function.

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