scholarly journals The Adapter Protein CrkII Regulates Neuronal Wiskott-Aldrich Syndrome Protein, Actin Polymerization, and Tension Development during Contractile Stimulation of Smooth Muscle

2005 ◽  
Vol 280 (24) ◽  
pp. 23380-23389 ◽  
Author(s):  
Dale D. Tang ◽  
Wenwu Zhang ◽  
Susan J. Gunst
Keyword(s):  
Smooth Muscle ◽  
Actin Polymerization ◽  
Adapter Protein ◽  
Tension Development ◽  
Wiskott Aldrich Syndrome ◽  
Syndrome Protein ◽  
Stimulation Of

scholarly journals The Small GTPase Cdc42 Regulates Actin Polymerization and Tension Development during Contractile Stimulation of Smooth Muscle

2004 ◽  
Vol 279 (50) ◽  
pp. 51722-51728 ◽  
Author(s):  
Dale D. Tang ◽  
Susan J. Gunst
Keyword(s):  
Smooth Muscle ◽  
Actin Polymerization ◽  
Myosin Light Chain ◽  
Dominant Negative ◽  
Related Protein ◽  
Wild Type ◽  
Tension Development ◽  
Muscle Tissues ◽  
Syndrome Protein ◽  
Stimulation Of

Contractile stimulation induces actin polymerization in smooth muscle tissues and cells, and the inhibition of actin polymerization depresses smooth muscle force development. In the present study, the role of Cdc42 in the regulation of actin polymerization and tension development in smooth muscle was evaluated. Acetylcholine stimulation of tracheal smooth muscle tissues increased the activation of Cdc42. Plasmids encoding wild type Cdc42 or a dominant negative Cdc42 mutant, Asn-17 Cdc42, were introduced into tracheal smooth muscle strips by reversible permeabilization, and tissues were incubated for 2 days to allow for protein expression. Expression of recombinant proteins was confirmed by immunoblot analysis. The expression of the dominant negative Cdc42 mutant inhibited contractile force and the increase in actin polymerization in response to acetylcholine stimulation but did not inhibit the increase in myosin light chain phosphorylation. The expression of wild type Cdc42 had no significant effect on force, actin polymerization, or myosin light chain phosphorylation. Contractile stimulation increased the association of neuronal Wiskott-Aldrich syndrome protein with Cdc42 and the Arp2/3 (actin-related protein) complex in smooth muscle tissues expressing wild type Cdc42. The agonist-induced increase in these protein interactions was inhibited in tissues expressing the inactive Cdc42 mutant. We conclude that Cdc42 activation regulates active tension development and actin polymerization during contractile stimulation. Cdc42 may regulate the activation of neuronal Wiskott-Aldrich syndrome protein and the actin related protein complex, which in turn regulate actin filament polymerization initiated by the contractile stimulation of smooth muscle.

scholarly journals Fyn and PTP-PEST–mediated Regulation of Wiskott-Aldrich Syndrome Protein (WASp) Tyrosine Phosphorylation Is Required for Coupling T Cell Antigen Receptor Engagement to WASp Effector Function and T Cell Activation

2004 ◽  
Vol 199 (1) ◽  
pp. 99-112 ◽  
Author(s):  
Karen Badour ◽  
Jinyi Zhang ◽  
Fabio Shi ◽  
Yan Leng ◽  
Michael Collins ◽  
...  
Keyword(s):  
T Cell ◽  
Tyrosine Phosphorylation ◽  
T Cell Activation ◽  
Actin Polymerization ◽  
Cell Activation ◽  
Synapse Formation ◽  
Tyrosine Phosphatase ◽  
Structural Constraints ◽  
Wiskott Aldrich Syndrome ◽  
Syndrome Protein

Involvement of the Wiskott-Aldrich syndrome protein (WASp) in promoting cell activation requires its release from autoinhibitory structural constraints and has been attributed to WASp association with activated cdc42. Here, however, we show that T cell development and T cell receptor (TCR)-induced proliferation and actin polymerization proceed normally in WASp−/− mice expressing a WASp transgene lacking the cdc42 binding domain. By contrast, mutation of tyrosine residue Y291, identified here as the major site of TCR-induced WASp tyrosine phosphorylation, abrogated induction of WASp tyrosine phosphorylation and its effector activities, including nuclear factor of activated T cell transcriptional activity, actin polymerization, and immunological synapse formation. TCR-induced WASp tyrosine phosphorylation was also disrupted in T cells lacking Fyn, a kinase shown here to bind, colocalize with, and phosphorylate WASp. By contrast, WASp was tyrosine dephosphorylated by protein tyrosine phosphatase (PTP)-PEST, a tyrosine phosphatase shown here to interact with WASp via proline, serine, threonine phosphatase interacting protein (PSTPIP)1 binding. Although Fyn enhanced WASp-mediated Arp2/3 activation and was required for synapse formation, PTP-PEST combined with PSTPIP1 inhibited WASp-driven actin polymerization and synapse formation. These observations identify key roles for Fyn and PTP-PEST in regulating WASp and imply that inducible WASp tyrosine phosphorylation can occur independently of cdc42 binding, but unlike the cdc42 interaction, is absolutely required for WASp contributions to T cell activation.

scholarly journals TheSaccharomyces cerevisiaeHomologue of Human Wiskott–Aldrich Syndrome Protein Las17p Interacts with the Arp2/3 Complex

1999 ◽  
Vol 10 (10) ◽  
pp. 3521-3538 ◽  
Author(s):  
Ammar Madania ◽  
Pascal Dumoulin ◽  
Sandrine Grava ◽  
Hiroko Kitamoto ◽  
Claudia Schärer-Brodbeck ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Temperature Sensitive ◽  
Multicopy Suppressor ◽  
Wiskott Aldrich Syndrome ◽  
Regulatory Cascades ◽  
Src Homology ◽  
Synthetically Lethal ◽  
Actin Patch ◽  
Syndrome Protein

Yeast Las17 protein is homologous to the Wiskott–Aldrich Syndrome protein, which is implicated in severe immunodeficiency. Las17p/Bee1p has been shown to be important for actin patch assembly and actin polymerization. Here we show that Las17p interacts with the Arp2/3 complex. LAS17 is an allele-specific multicopy suppressor of ARP2 and ARP3 mutations; overexpression restores both actin patch organization and endocytosis defects in ARP2 temperature-sensitive (ts) cells. Six of seven ARP2 ts mutants and at least oneARP3 ts mutant are synthetically lethal withlas17Δ ts confirming functional interaction with the Arp2/3 complex. Further characterization of las17Δcells showed that receptor-mediated internalization of α factor by the Ste2 receptor is severely defective. The polarity of normal bipolar bud site selection is lost. Las17-gfp remains localized in cortical patches in vivo independently of polymerized actin and is required for the polarized localization of Arp2/3 as well as actin. Coimmunoprecipitation of Arp2p with Las17p indicates that Las17p interacts directly with the complex. Two hybrid results also suggest that Las17p interacts with actin, verprolin, Rvs167p and several other proteins including Src homology 3 (SH3) domain proteins, suggesting that Las17p may integrate signals from different regulatory cascades destined for the Arp2/3p complex and the actin cytoskeleton.

scholarly journals PHD2 deletion in endothelial or arterial smooth muscle cells reveals vascular cell type-specific responses in pulmonary hypertension and fibrosis

Angiogenesis ◽  
2022 ◽  
Author(s):  
Harri Elamaa ◽  
Mika Kaakinen ◽  
Marjut Nätynki ◽  
Zoltan Szabo ◽  
Veli-Pekka Ronkainen ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Actin Polymerization ◽  
Vascular Tone ◽  
Muscle Cells ◽  
Low Oxygen ◽  
Arterial Smooth Muscle ◽  
Tension Development ◽  
Hemodynamic Stress ◽  
Arterial Smooth Muscle Cells

AbstractHypoxia plays an important regulatory role in the vasculature to adjust blood flow to meet metabolic requirements. At the level of gene transcription, the responses are mediated by hypoxia-inducible factor (HIF) the stability of which is controlled by the HIF prolyl 4-hydroxylase-2 (PHD2). In the lungs hypoxia results in vasoconstriction, however, the pathophysiological relevance of PHD2 in the major arterial cell types; endothelial cells (ECs) and arterial smooth muscle cells (aSMCs) in the adult vasculature is incompletely characterized. Here, we investigated PHD2-dependent vascular homeostasis utilizing inducible deletions of PHD2 either in ECs (Phd2∆ECi) or in aSMCs (Phd2∆aSMC). Cardiovascular function and lung pathologies were studied using echocardiography, Doppler ultrasonography, intraventricular pressure measurement, histological, ultrastructural, and transcriptional methods. Cell intrinsic responses were investigated in hypoxia and in conditions mimicking hypertension-induced hemodynamic stress. Phd2∆ECi resulted in progressive pulmonary disease characterized by a thickened respiratory basement membrane (BM), alveolar fibrosis, increased pulmonary artery pressure, and adaptive hypertrophy of the right ventricle (RV). A low oxygen environment resulted in alterations in cultured ECs similar to those in Phd2∆ECi mice, involving BM components and vascular tone regulators favoring the contraction of SMCs. In contrast, Phd2∆aSMC resulted in elevated RV pressure without alterations in vascular tone regulators. Mechanistically, PHD2 inhibition in aSMCs involved  actin polymerization -related tension development via activated cofilin. The results also indicated that hemodynamic stress, rather than PHD2-dependent hypoxia response alone, potentiates structural remodeling of the extracellular matrix in the pulmonary microvasculature and respiratory failure.

Tyrosine Phosphorylation of WASP Promotes Calpain-Mediated Podosome Disassembly In Myeloid Cells.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1498-1498
Author(s):  
Lee Macpherson ◽  
James Monypenny ◽  
Michael P Blundell ◽  
Giles O Cory ◽  
Jessica Tomé-García ◽  
...  
Keyword(s):  
Tyrosine Phosphorylation ◽  
Actin Polymerization ◽  
Conflicts Of Interest ◽  
Myeloid Cells ◽  
Reference List ◽  
Wiskott Aldrich Syndrome ◽  
Open Conformation ◽  
New Findings ◽  
Syndrome Protein

Abstract Abstract 1498 We have previously shown that a point mutation (Ile294Thr) in the Wiskott Aldrich Syndrome Protein (WASP) detected in a Wiskott Aldrich Syndrome (WAS) patient led to enhanced actin polymerising activity of WASP 1 and well as increased instability2. We also showed that macrophages from this patient displayed an increased number of actin based adhesion structures called podosomes. Additionally, podosomes in macrophages from this patient were extremely dynamic with a high rate of turnover. Based on these results we proposed that the active open conformation of WASP promotes actin polymerisation but it also induces podosome disassembly and adhesion turnover. Although the idea that the same open conformation of WASP leads to both assembly and disassembly of podosomes may be counterintuitive at first, it is possible that for termination of podosomes actin polymerisation and integrin recruitment have to be discontinued and the same constituents of growing podosomes such as active WASP may contribute to the disassembly process. We have also shown that the rapid turn-over of podosomes involves cleavage of WASP by the protease calpain, further supporting a role of WASP in both podosome formation and disassembly. However, the specific signalling mechanisms that make active WASP susceptible to cleavage by calpain leading to podosome disassembly remain unknown and need further clarification. Phosphorylation of WASP Y291 (human) or Y293 (mouse) promotes the open conformation of WASP and results in enhanced actin polymerisation. We now report that in myeloid cells, tyrosine phosphorylation of WASP negatively regulates the stability of podosomes, leading to their calpain-dependent disassembly. Additionally, we found that constitutive phosphorylation of WASP results in extensive degradation in a process that involves calpain. Our data also indicate that phosphorylation of WASP sustains the open/active conformation that promotes cleavage of WASP by calpain. Interestingly, phosphorylated WASP can bind to the WASP interacting protein (WIP) and other proteins that form a complex with WASP in podosomes such as Nck, cortactin. Taken together, our data indicate that in myeloid cells, tyrosine phosphorylation sustains the open conformation of WASP and it enhances its susceptibility to calpain-mediated cleavage preventing accumulation of actin filaments and integrin associated proteins in podosomes. This process facilitates podosome disassembly and cell translocation. These new findings support the key role of WASP as a protein that integrates actin polymerisation and cell adhesion required for mobilisation of myeloid cells during the immune response. Reference List 1. Ancliff PJ, Blundell MP, Cory GO et al. Two novel activating mutations in the Wiskott-Aldrich syndrome protein result in congenital neutropenia. Blood 2006;108:2182-2189. 2. Moulding DA, Blundell MP, Spiller DG et al. Unregulated actin polymerization by WASp causes defects of mitosis and cytokinesis in X-linked neutropenia. J Exp. Med 2007;204:2213–2224. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Cryptosporidium parvum Infection Requires Host Cell Actin Polymerization

2001 ◽  
Vol 69 (9) ◽  
pp. 5940-5942 ◽  
Author(s):  
David A. Elliott ◽  
Daniel J. Coleman ◽  
Michael A. Lane ◽  
Robin C. May ◽  
Laura M. Machesky ◽  
...  
Keyword(s):  
Host Cell ◽  
Cryptosporidium Parvum ◽  
Actin Polymerization ◽  
Protozoan Parasite ◽  
Parasite Infection ◽  
Cell Cytoplasm ◽  
Wiskott Aldrich Syndrome ◽  
Host Cell Cytoplasm ◽  
Syndrome Protein ◽  
Intracellular Protozoan Parasite

ABSTRACT The intracellular protozoan parasite Cryptosporidium parvum accumulates host cell actin at the interface between the parasite and the host cell cytoplasm. Here we show that the actin polymerizing proteins Arp2/3, vasodilator-stimulated phosphoprotein (VASP), and neural Wiskott Aldrich syndrome protein (N-WASP) are present at this interface and that host cell actin polymerization is necessary for parasite infection.

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