Adaptor protein XB130 is a Rac-controlled component of lamellipodia that regulates cell motility and invasion

ABSTRACT Cooperation between the Neu/ErbB-2 and transforming growth factor β (TGF-β) signaling pathways enhances the invasive and metastatic capabilities of breast cancer cells; however, the underlying mechanisms mediating this synergy have yet to be fully explained. We demonstrate that TGF-β induces the migration and invasion of mammary tumor explants expressing an activated Neu/ErbB-2 receptor, which requires signaling from autophosphorylation sites located in the C terminus. A systematic analysis of mammary tumor explants expressing Neu/ErbB-2 add-back receptors that couple to distinct signaling molecules has mapped the synergistic effect of TGF-β-induced motility and invasion to signals emanating from tyrosine residues 1226/1227 and 1253 of Neu/ErbB-2. Given that the ShcA adaptor protein is known to interact with Neu/ErbB-2 through these residues, we investigated the importance of this signaling molecule in TGF-β-induced cell motility and invasion. The reduction of ShcA expression rendered cells expressing activated Neu/ErbB-2, or add-back receptors signaling specifically through tyrosines 1226/1227 or 1253, unresponsive to TGF-β-induced motility and invasion. In addition, a dominant-negative form of ShcA, lacking its three known tyrosine phosphorylation sites, completely abrogates the TGF-β-induced migration and invasion of breast cancer cells expressing activated Neu/ErbB-2. Our results implicate signaling through the ShcA adaptor as a key component in the synergistic interaction between these pathways.

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Functional Characterization of Obesity-Associated Variants Involving the α and β Isoforms of Human SH2B1

Endocrinology ◽

10.1210/en.2014-1264 ◽

2014 ◽

Vol 155 (9) ◽

pp. 3219-3226 ◽

Cited By ~ 20

Author(s):

Laura R. Pearce ◽

Ray Joe ◽

Michael E. Doche ◽

Hsiao-Wen Su ◽

Julia M. Keogh ◽

...

Keyword(s):

Nerve Growth Factor ◽

Growth Factor ◽

Cell Motility ◽

Neurite Outgrowth ◽

Functional Characterization ◽

Adaptor Protein ◽

Maladaptive Behavior ◽

In Vitro Assays ◽

Nerve Growth ◽

The Individual

Abstract We have previously reported rare variants in sarcoma (Src) homology 2 (SH2) B adaptor protein 1 (SH2B1) in individuals with obesity, insulin resistance, and maladaptive behavior. Here, we identify 4 additional SH2B1 variants by sequencing 500 individuals with severe early-onset obesity. SH2B1 has 4 alternatively spliced isoforms. One variant (T546A) lies within the N-terminal region common to all isoforms. As shown for past variants in this region, T546A impairs SH2B1β enhancement of nerve growth factor-induced neurite outgrowth, and the individual with the T546A variant exhibits mild developmental delay. The other 3 variants (A663V, V695M, and A723V) lie in the C-terminal tail of SH2B1α. SH2B1α variant carriers were hyperinsulinemic but did not exhibit the behavioral phenotype observed in individuals with SH2B1 variants that disrupt all isoforms. In in vitro assays, SH2B1α, like SH2B1β, enhances insulin- and leptin-induced insulin receptor substrate 2 (IRS2) phosphorylation and GH-induced cell motility. None of the variants affect SH2B1α enhancement of insulin- and leptin-induced IRS2 phosphorylation. However, T546A, A663V, and A723V all impair the ability of SH2B1α to enhance GH-induced cell motility. In contrast to SH2B1β, SH2B1α does not enhance nerve growth factor-induced neurite outgrowth. These studies suggest that genetic variants that disrupt isoforms other than SH2B1β may be functionally significant. Further studies are needed to understand the mechanism by which the individual isoforms regulate energy homeostasis and behavior.

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PEAK3/C19orf35 pseudokinase, a new NFK3 kinase family member, inhibits CrkII through dimerization

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1906360116 ◽

2019 ◽

Vol 116 (31) ◽

pp. 15495-15504 ◽

Cited By ~ 3

Author(s):

Mitchell L. Lopez ◽

Megan Lo ◽

Jennifer E. Kung ◽

Małgorzata Dudkiewicz ◽

Gwendolyn M. Jang ◽

...

Keyword(s):

Cell Motility ◽

Family Member ◽

Cancer Progression ◽

Family Members ◽

Adaptor Protein ◽

Dimerization Domain ◽

Unique Role ◽

Membrane Ruffling ◽

Kinase Family ◽

Evolutionarily Conserved

Members of the New Kinase Family 3 (NKF3), PEAK1/SgK269 and Pragmin/SgK223 pseudokinases, have emerged as important regulators of cell motility and cancer progression. Here, we demonstrate that C19orf35 (PEAK3), a newly identified member of the NKF3 family, is a kinase-like protein evolutionarily conserved across mammals and birds and a regulator of cell motility. In contrast to its family members, which promote cell elongation when overexpressed in cells, PEAK3 overexpression does not have an elongating effect on cell shape but instead is associated with loss of actin filaments. Through an unbiased search for PEAK3 binding partners, we identified several regulators of cell motility, including the adaptor protein CrkII. We show that by binding to CrkII, PEAK3 prevents the formation of CrkII-dependent membrane ruffling. This function of PEAK3 is reliant upon its dimerization, which is mediated through a split helical dimerization domain conserved among all NKF3 family members. Disruption of the conserved DFG motif in the PEAK3 pseudokinase domain also interferes with its ability to dimerize and subsequently bind CrkII, suggesting that the conformation of the pseudokinase domain might play an important role in PEAK3 signaling. Hence, our data identify PEAK3 as an NKF3 family member with a unique role in cell motility driven by dimerization of its pseudokinase domain.

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Role for a Cindr–Arf6 axis in patterning emerging epithelia

Molecular Biology of the Cell ◽

10.1091/mbc.e11-04-0305 ◽

2011 ◽

Vol 22 (23) ◽

pp. 4513-4526 ◽

Cited By ~ 20

Author(s):

Ruth I. Johnson ◽

Alanna Sedgwick ◽

Crislyn D'Souza-Schorey ◽

Ross L. Cagan

Keyword(s):

Cell Motility ◽

Adaptor Protein ◽

Time Lapse ◽

Similar Relationship ◽

Surface Adhesion ◽

Cell Movements ◽

Surface Receptors ◽

Time Lapse Microscopy ◽

Gtpase Activating Proteins ◽

Regulatory Cascade

Patterning of the Drosophila pupal eye is characterized by precise cell movements. In this paper, we demonstrate that these movements require an Arf regulatory cycle that connects surface receptors to actin-based movement. dArf6 activity—regulated by the Arf GTPase–activating proteins (ArfGAPs) dAsap and dArfGAP3 and the Arf GTP exchange factors Schizo and dPsd—promoted large cellular extensions; time-lapse microscopy indicated that these extensions presage cell rearrangements into correct epithelial niches. During this process, the Drosophila eye also requires interactions between surface Neph1/nephrin adhesion receptors Roughest and Hibris, which bind the adaptor protein Cindr (CD2AP). We provide evidence that Cindr forms a physical complex with dArfGAP3 and dAsap. Our data suggest this interaction sequesters ArfGAP function to liberate active dArf6 elsewhere in the cell. We propose that a Neph1/nephrin–Cindr/ArfGAP complex accumulates to limit local Arf6 activity and stabilize adherens junctions. Our model therefore links surface adhesion via an Arf6 regulatory cascade to dynamic modeling of the cytoskeleton, accounting for precise cell movements that organize the functional retinal field. Further, we demonstrate a similar relationship between the mammalian Cindr orthologue CD2AP and Arf6 activity in cell motility assays. We propose that this Cindr/CD2AP-mediated regulation of Arf6 is a widely used mechanism in emerging epithelia.

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Phosphorylation of Tyrosine Residues 31 and 118 on Paxillin Regulates Cell Migration through an Association with Crk in Nbt-II Cells

The Journal of Cell Biology ◽

10.1083/jcb.148.5.957 ◽

2000 ◽

Vol 148 (5) ◽

pp. 957-970 ◽

Cited By ~ 170

Author(s):

Valérie Petit ◽

Brigitte Boyer ◽

Delphine Lentz ◽

Christopher E. Turner ◽

Jean Paul Thiery ◽

...

Keyword(s):

Cell Migration ◽

Cell Motility ◽

Critical Role ◽

Adaptor Protein ◽

Signaling Molecules ◽

Sh2 Domain ◽

Wild Type ◽

Tyrosine Residues ◽

Pathological Conditions

Identification of signaling molecules that regulate cell migration is important for understanding fundamental processes in development and the origin of various pathological conditions. The migration of Nara Bladder Tumor II (NBT-II) cells was used to determine which signaling molecules are specifically involved in the collagen-mediated locomotion. We show here that paxillin is tyrosine phosphorylated after induction of motility on collagen. Overexpression of paxillin mutants in which tyrosine 31 and/or tyrosine 118 were replaced by phenylalanine effectively impaired cell motility. Moreover, stimulation of motility by collagen preferentially enhanced the association of paxillin with the SH2 domain of the adaptor protein CrkII. Mutations in both tyrosine 31 and 118 diminished the phosphotyrosine content of paxillin and prevented the formation of the paxillin–Crk complex, suggesting that this association is necessary for collagen-mediated NBT-II cell migration. Other responses to collagen, such as cell adhesion and spreading, were not affected by these mutations. Overexpression of wild-type paxillin or Crk could bypass the migration-deficient phenotype. Both the SH2 and the SH3 domains of CrkII are shown to play a critical role in this collagen-mediated migration. These results demonstrate the important role of the paxillin–Crk complex in the collagen-induced cell motility.

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Calpain-mediated Proteolysis of Paxillin Negatively Regulates Focal Adhesion Dynamics and Cell Migration

Journal of Biological Chemistry ◽

10.1074/jbc.m110.187294 ◽

2011 ◽

Vol 286 (12) ◽

pp. 9998-10006 ◽

Cited By ~ 53

Author(s):

Christa L. Cortesio ◽

Lindsy R. Boateng ◽

Timothy M. Piazza ◽

David A. Bennin ◽

Anna Huttenlocher

Keyword(s):

Cell Migration ◽

Cell Motility ◽

Focal Adhesion ◽

Focal Adhesions ◽

Adaptor Protein ◽

Time Lapse ◽

Negative Regulator ◽

Proteolytic Fragment ◽

Proteolytic Site ◽

And Migration

The dynamic turnover of integrin-mediated adhesions is important for cell migration. Paxillin is an adaptor protein that localizes to focal adhesions and has been implicated in cell motility. We previously reported that calpain-mediated proteolysis of talin1 and focal adhesion kinase mediates adhesion disassembly in motile cells. To determine whether calpain-mediated paxillin proteolysis regulates focal adhesion dynamics and cell motility, we mapped the preferred calpain proteolytic site in paxillin. The cleavage site is between the paxillin LD1 and LD2 motifs and generates a C-terminal fragment that is similar in size to the alternative product paxillin delta. The calpain-generated proteolytic fragment, like paxillin delta, functions as a paxillin antagonist and impairs focal adhesion disassembly and migration. We generated mutant paxillin with a point mutation (S95G) that renders it partially resistant to calpain proteolysis. Paxillin-deficient cells that express paxillin S95G display increased turnover of zyxin-containing adhesions using time-lapse microscopy and also show increased migration. Moreover, cancer-associated somatic mutations in paxillin are common in the N-terminal region between the LD1 and LD2 motifs and confer partial calpain resistance. Taken together, these findings suggest a novel role for calpain-mediated proteolysis of paxillin as a negative regulator of focal adhesion dynamics and migration that may function to limit cancer cell invasion.

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HGAL inhibits lymphoma dissemination by interacting with multiple Cytoskeletal proteins

Blood Advances ◽

10.1182/bloodadvances.2021004304 ◽

2021 ◽

Author(s):

Xiaoyu Jiang ◽

Xiaoqing Lu ◽

Andrew J Gentles ◽

Dekuang Zhao ◽

Seth A. Wander ◽

...

Keyword(s):

Cell Motility ◽

B Cell ◽

Germinal Center ◽

Molecular Mechanisms ◽

Early Stage ◽

Adaptor Protein ◽

Cytoskeletal Proteins ◽

Proteomic Approach

Human Germinal Center Associated Lymphoma (HGAL) is an adaptor protein specifically expressed in germinal center lymphocytes. High expression of HGAL is a predictor of prolonged survival of Diffuse Large B-Cell (DLBCL) and classical Hodgkin lymphomas. Furthermore, HGAL expression is associated with early stage DLBCL, thus potentially limiting lymphoma dissemination. In our previous studies, we demonstrated that HGAL regulates B-cell receptor signaling and cell motility in vitro and deciphered some molecular mechanisms underlying these effects. Herein, by using novel animal models for in vivo DLBCL dispersion, we demonstrate that HGAL decreases lymphoma dissemination and prolongs survival. Further, by using an unbiased proteomic approach we demonstrate that HGAL may interact with multiple cytoskeletal proteins whereby implicating a multiplicity of effects in regulating lymphoma motility and spread. Specifically, we show that HGAL interacts with tubulin and this interaction may also contribute to HGAL effects on cell motility. These findings recapitulate previous observations in humans, establish the role of HGAL in lymphoma in vivo dissemination, and explain improved survival of patients with HGAL expressing lymphomas.

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