Dia1 and IQGAP1 interact in cell migration and phagocytic cup formation

Brandt, Dominique; Marion, Sabrina; Griffiths, Gareth; Watanabe, Takashi; Kaibuchi, Kozo; Grosse, Robert

doi:10.1083/jcb.200612071

Cited by 179 publications

(195 citation statements)

References 26 publications

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“…IQGAP1 is known to associate with E-cadherin, regulate actin assembly, and coordinate the tethering of MT during polarized cell migration through interaction with MT plus-end binding proteins [21,22,24,35,36]. Our data implicate the participation of MT in endothelial IQGAP1-dependent junction remodeling during lymphocyte diapedesis.…”

Section: Discussionmentioning

confidence: 62%

“…In our experiments, the IQGAP1-deficient HUVEC were plated at confluence, then maintained in complete media with 20% FBS for 48 h to promote junction maturation. Hence, in the current experiments, effects of IQGAP1 knockdown on cell migration or repopulation at subconfluent densities were minimized.IQGAP1 is known to associate with E-cadherin, regulate actin assembly, and coordinate the tethering of MT during polarized cell migration through interaction with MT plus-end binding proteins [21,22,24,35,36]. Our data implicate the participation of MT in endothelial IQGAP1-dependent junction remodeling during lymphocyte diapedesis.…”

mentioning

confidence: 62%

“…Next, we sought to characterize the effect of IQGAP1 knockdown on EC cytoskeletal components since IQGAP1 regulates dynamic filamentous-actin (F-actin) polymerization [23,35,36] and MT capture at the cell cortex [21][22][23]. Biochemical analysis of free and polymerized tubulin within EC determined IQGAP1 knockdown decreased the ratio of polymerized tubulin to free tubulin levels in the cytosolic extracts ( Fig.…”

Section: Loss Of Iqgap1 Perturbs Tethering Of Mt At Ajmentioning

confidence: 99%

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Endothelial IQGAP1 regulates efficient lymphocyte transendothelial migration

2009

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Leukocyte movement from the blood to the tissues is a fundamental process in acute and chronic inflammatory diseases. While the role of endothelial cells (EC) to recruit leukocytes to sites of inflammation is well known, the mechanisms that control remodeling of EC shape and adhesive contacts during leukocyte transendothelial migration (TEM) are not completely understood. We studied the role of IQGAP1, an adaptor protein that binds to filamentous-actin and microtubules (MT) at interendothelial junctions, during lymphocyte TEM. EC IQGAP1 knockdown decreases MT tethered to the adherens junction, and decreases lymphocyte TEM to $70% (po0.05) versus control. Similarly, loss of adherens junction-associated MT induced by brief nocodazole (ND) treatment decreases lymphocyte TEM to $65% of control (po0.01). Confocal microscopy imaging indicates that EC IQGAP1 knockdown and MT depolymerization both result in lymphocyte accumulation above the vascular endothelial cadherin (VE-cadherin) junctions and reduces the fraction of adherent lymphocytes that complete diapedesis across interendothelial cell junctions. However, we observe no change in VE-cadherin gap formation underlying adherent lymphocytes among control, IQGAP1 knockdown, or MT depolymerised EC monolayers. These data indicate that IQGAP1 contributes to MT stability at endothelial junctions. Further, IQGAP1 is involved in junction remodeling required for efficient lymphocyte diapedesis, independent of VE-cadherin gap formation.Key words: Adherens junctions . Cytoskeleton . IQGAP1 Supporting Information available online IntroductionLeukocyte extravasation is fundamental to the development of many immune responses including solid-organ allograft rejection. In this process, leukocytes leave the bloodstream and migrate into tissues through the endothelial cells (EC) that line the walls of vessels, i.e. leukocytes undergo transendothelial migration (TEM). Whereas the specific adhesion molecules, chemoattractants, and possibly signaling pathways involved in TEM are unique among different subgroups of leukocytes and vascular beds, the interaction between leukocytes and EC during TEM can be generalized into a multicascade event, described in recent reviews [1][2][3]. EC and leukocyte adhesion molecules mediate tethering and rolling of leukocytes on EC followed by chemokinemediated leukocyte activation, then firm adhesion to the EC. Finally, adherent leukocytes crawl on the surface of endothelium, undergo diapedesis, and enter tissues by mechanisms that are not fully understood.Leukocyte transmigration may occur by either a transcellular, through EC, or paracellular route, between adjacent EC [4][5][6]. 204The latter is associated with structural changes in the interendothelial adhesion structures and EC cytoskeleton [5,7,8]. Cross-linking of adhesion molecules such as CD54 or CD106 is shown to mediate signals that lead to EC actin cytoskeleton remodeling [9][10][11][12]. These signaling cascades promote structural changes in interendothlelial junctions, which might be requ...

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Section: Discussionmentioning

confidence: 62%

mentioning

confidence: 62%

Section: Loss Of Iqgap1 Perturbs Tethering Of Mt At Ajmentioning

confidence: 99%

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Endothelial IQGAP1 regulates efficient lymphocyte transendothelial migration

2009

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“…This regulation of the myosin heavy chain differs from the usual regulation through the light chain ( Conti and Adelstein, 2008 ). Talin, ␣ -actinin, and the actin nucleator mDia1 also appeared regulated (to a lesser extent), and all have been implicated previously in phagocytosis (respectively: [( Turner et al, 1989 ;Greenberg et al, 1990 ); ( Crowley and Horwitz, 1995 ;Izaguirre et al, 1999 ); ( Meng et al, 2004 ;Brandt et al, 2007 )]). Although phospho-paxillin was downregulated in our imaging studies, it was not prominent in our immunoblots and was unaffected by blebbistatin, which otherwise mimicked CD47 ' s inhibitory effects ( Figs.…”

Section: Phosphotyrosine Activation Of Nmm Iia In Phagocytosismentioning

confidence: 97%

Inhibition of “self” engulfment through deactivation of myosin-II at the phagocytic synapse between human cells

Tsai¹,

Discher²

2008

J Exp Med

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(2008). Inhibition of "self " engulfment through deactivation of myosin-II at the phagocytic synapse between human cells. Retrieved from http://repository.upenn.edu/cbe_papers/108Inhibition of "self " engulfment through deactivation of myosin-II at the phagocytic synapse between human cells AbstractPhagocytosis of foreign cells or particles by macrophages is a rapid process that is inefficient when faced with "self " cells that display CD47-although signaling mechanisms in self-recognition have remained largely unknown. With human macrophages, we show the phagocytic synapse at cell contacts involves a basal level of actin-driven phagocytosis that, in the absence of species-specific CD47 signaling, is made more efficient by phospho-activated myosin. We use "foreign" sheep red blood cells (RBCs) together with CD47-blocked, antibody-opsonized human RBCs in order to visualize synaptic accumulation of phosphotyrosine, paxillin, Factin, and the major motor isoform, nonmuscle myosin-IIA. When CD47 is functional, the macrophage counter-receptor and phosphatase-activator SIRPα localizes to the synapse, suppressing accumulation of phosphotyrosine and myosin without affecting F-actin. On both RBCs and microbeads, human CD47 potently inhibits phagocytosis as does direct inhibition of myosin. CD47-SIRPα interaction initiates a dephosphorylation cascade directed in part at phosphotyrosine in myosin. A point mutation turns off this motor's contribution to phagocytosis, suggesting that self-recognition inhibits contractile engulfment. Comments

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“…Proteins were dialyzed against 2 mM NaH 2 PO 4 pH7, 150 mM NaCl, 0.1 mM EGTA, 0.1 mM MgCl 2 , 0.5 mM DTT. Pyrenyl actin assembly assays were performed with minor modifications, as described [54] using a reaction mix containing 10 µl energy-regenerating mix (150 mM creatine phosphate, 20 mM ATP, 2 mM EGTA, 20 mM MgCl 2 ), 10 µl pyrene-labeled actin (2 µM final concentration, 10% labeled; Hypermol) and proteins at required amounts. Pyrene fluorescence was measured at 407 nm with excitation at 365 nm, and the kinetics of actin filament assembly were monitored using a SHIMADZU RF-5301PC spectrofluorphotometer.…”

Section: Northern Blottingmentioning

confidence: 99%

Differing and isoform-specific roles for the formin DIAPH3 in plasma membrane blebbing and filopodia formation

et al. 2011

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Plasma membrane (PM) blebs are dynamic actin-rich cell protrusions that occur, e.g., during cytokinesis, amoeboid cell motility and cell attachment. Using a targeted siRNA screen against 21 actin nucleation factors, we identify a novel and essential role of the human diaphanous formin DIAPH3 in PM blebbing during cell adhesion. Suppression of DIAPH3 inhibited blebbing to promote rapid cell spreading involving β1-integrin. Multiple isoforms of DIAPH3 were detected on the mRNA and protein level of which isoforms 3 and 7 were the largest and most abundant isoforms that however did not induce formation of actin-rich protrusions. Rather, PM blebbing specifically involved the low abundance isoform 1 of DIAPH3 and activation of isoform 7 by deletion of the diaphanous-autoregulatory domain caused the formation of filopodia. Dimerization and actin assembly activity were essential for induction of specific cell protrusions by DIAPH3 isoforms 1 and 7. Our data suggest that the N-terminal region comprising the GTPasebinding domain determined the subcellular localization of the formin as well as its protrusion activity between blebs and filopodia. We propose that isoform-selective actin assembly by DIAPH3 exerts specific and differentially regulated functions during cell adhesion and motility.

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Dia1 and IQGAP1 interact in cell migration and phagocytic cup formation

Cited by 179 publications

References 26 publications

Endothelial IQGAP1 regulates efficient lymphocyte transendothelial migration

Endothelial IQGAP1 regulates efficient lymphocyte transendothelial migration

Inhibition of “self” engulfment through deactivation of myosin-II at the phagocytic synapse between human cells

Differing and isoform-specific roles for the formin DIAPH3 in plasma membrane blebbing and filopodia formation

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