Tyrosine phosphorylation within the SH3 domain regulates CAS subcellular localization, cell migration, and invasiveness

Janoštiak, Radoslav; Tolde, Ondřej; Brůhová, Zuzana; Novotný, Marián; Hanks, Steven K.; Rösel, Daniel; Brábek, Jan

doi:10.1091/mbc.e11-03-0207

Cited by 41 publications

(64 citation statements)

References 47 publications

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“…Accordingly, phosphorylated Cas levels in FAs increased approximately 3-fold when Cul5 expression was inhibited, especially in FAs in the front 6 µm of the cell (Figure 5—figure supplement 3). Since Cas exchanges rapidly between the cytosol and FAs (Janostiak et al, 2011; Machiyama et al, 2014), the increase in pYCas in FAs could be secondary to the increased Cas in the cytosol. Moreover, the chronic increase in pYCas in adhesions could accelerate adhesion turnover regardless of whether SOCS6 targets Cas in the cytosol or in FAs.…”

Section: Resultsmentioning

confidence: 99%

“…Removal of Cas by this mechanism would be expected to open up sites in the FA where other Cas molecules could bind. It is known that most of the Cas in a FA exchanges with Cas from the cytosol with a very rapid (~20 s) rate constant (Janostiak et al, 2011; Machiyama et al, 2014). It seems remarkable that ubiquitination and degradation of pYCas would affect the kinetics of FA disassembly if new Cas molecules could replace degraded molecules with such speed.…”

Section: Discussionmentioning

confidence: 99%

“…Cytoskeletal tension induces Cas tyrosine phosphorylation (Janostiak et al, 2011; Tamada et al, 2004). Mechanical extension of purified Cas promotes its phosphorylation by SFKs in vitro (Sawada et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

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The ubiquitin-proteasome system regulates focal adhesions at the leading edge of migrating cells

Teckchandani

Cooper

2016

eLife

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Cell migration requires the cyclical assembly and disassembly of focal adhesions. Adhesion induces phosphorylation of focal adhesion proteins, including Cas (Crk-associated substrate/p130Cas/BCAR1). However, Cas phosphorylation stimulates adhesion turnover. This raises the question of how adhesion assembly occurs against opposition from phospho-Cas. Here we show that suppressor of cytokine signaling 6 (SOCS6) and Cullin 5, two components of the CRL5SOCS6 ubiquitin ligase, inhibit Cas-dependent focal adhesion turnover at the front but not rear of migrating epithelial cells. The front focal adhesions contain phospho-Cas which recruits SOCS6. If SOCS6 cannot access focal adhesions, or if cullins or the proteasome are inhibited, adhesion disassembly is stimulated. This suggests that the localized targeting of phospho-Cas within adhesions by CRL5SOCS6 and concurrent cullin and proteasome activity provide a negative feedback loop, ensuring that adhesion assembly predominates over disassembly at the leading edge. By this mechanism, ubiquitination provides a new level of spatio-temporal control over cell migration.DOI: http://dx.doi.org/10.7554/eLife.17440.001

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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The ubiquitin-proteasome system regulates focal adhesions at the leading edge of migrating cells

Teckchandani

Cooper

2016

eLife

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“…Alternatively, it was shown that the interaction of the CAS-SH3 domain with its substrates can be regulated by phosphorylation on Tyr1291213. This enables dynamically regulated changes in CAS targeting to focal adhesions, which might be critical for proper subsequent signaling.…”

Section: Discussionmentioning

confidence: 99%

The role of focal adhesion anchoring domains of CAS in mechanotransduction

Braniš

Pataki

Spörrer

et al. 2017

Sci Rep

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CAS is a docking protein, which was shown to act as a mechanosensor in focal adhesions. The unique assembly of structural domains in CAS is important for its function as a mechanosensor. The tension within focal adhesions is transmitted to a stretchable substrate domain of CAS by focal adhesion-targeting of SH3 and CCH domain of CAS, which anchor the CAS protein in focal adhesions. Mechanistic models of the stretching biosensor propose equal roles for both anchoring domains. Using deletion mutants and domain replacements, we have analyzed the relative importance of the focal adhesion anchoring domains on CAS localization and dynamics in focal adhesions as well as on CAS-mediated mechanotransduction. We confirmed the predicted prerequisite of the focal adhesion targeting for CAS-dependent mechanosensing and unraveled the critical importance of CAS SH3 domain in mechanosensing. We further show that CAS localizes to the force transduction layer of focal adhesions and that mechanical stress stabilizes CAS in focal adhesions.

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“…Although post-translational modification of conventional SH3 domains by tyrosine phosphorylation has been documented, for example for Btk, Abl and recently for the CAS/BCAR1 SH3 domain, in contrast to the Crk SH3C, this occurs on the conserved tyrosine in the hydrophobic ligand binding pocket and thereby reduces the affinity of the SH3 domain for its ligands [14][15][16][17][18]. The SH3 domain of CAS/BCAR1, a focal adhesion protein shown to be associated with resistance to anti-estrogens in breast cancer, is phosphorylated on this tyrosine in Src-transformed cells [19].…”

Section: Introductionmentioning

confidence: 99%

Commentary: The carboxyl‐terminal Crk SH3 domain: Regulatory strategies and new perspectives

Sriram¹,

Birge²

2012

FEBS Letters

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a b s t r a c tSince their discovery as cellular counterparts of viral oncogenes more than two decades ago, enormous progress has been made in unraveling the complex regulatory pathways of signal transduction initiated by the Crk family of proteins. New structural and biochemical studies have uncovered novel insights into both negative and positive regulation of Crk mediated by its atypical carboxylterminal SH3 domain (SH3C). Moreover, SH3C is tyrosine phosphorylated by receptor tyrosine kinases and non-receptor tyrosine kinases, thereby permitting assemblages of other SH2/PTB domain containing proteins. Such non-canonical signaling by the Crk SH3C reveals new regulatory strategies for adaptor proteins.Ó 2012 Federation of European Biochemical Societies. Published by Elsevier B.V.Named for their ability to elevate cellular tyrosine phosphorylation and transform primary chicken embryo fibroblasts (CEFs) [1], CT10 regulator of kinase (Crk) is the prototypical member of a class of Src Homology-2 (SH2) and Src Homology-3 (SH3) domain-containing proteins that lack intrinsic enzymatic activity but instead function to promote protein-protein interactions [2,3]. Over the past two decades, enormous progress in our understanding of signal transduction has emerged from studies on Crk. The canonical signaling pathway, defined by recognition of specific phosphotyrosine motifs by the SH2 domain (in the context of p-Tyr.X.X.Pro) [4] and Polyproline Type II motifs by the N-terminal SH3 domain {in the context of Pro. X. X. Pro. X. (Lys/Arg)} [5], identified Crk as a connector between Receptor Tyrosine Kinases (RTKs) and/or their substrates and GTPase effector pathways (Fig. 1). Along with efforts on Grb2 [6], these insights were instrumental in defining missing links between integrins and/or tyrosine phosphorylated growth factor receptors and downstream effector pathways that regulate growth and differentiation, and also more generally for defining elements of modular domains in signaling. In recent years, new layers of regulation for Crk have emerged, pointing to not only unusual levels of auto-inhibition mediated by its atypical carboxylterminal SH3 domain, but also unexpected features of SH3 domain signaling that activate non-canonical pathways.CrkII and CrkL each contain an atypical C-terminal SH3 domain (SH3C), defined by its inability to bind to Polyproline Type II (PPII) motifs [7,22]. In both proteins, the atypical SH3 domain retains core structural characteristics of SH3 domains, consisting of a five-stranded b-barrel which forms the standard SH3 fold. However, the aromatic amino acids -F141, W169, Y186 which line the canonical PPII binding pocket on the Crk SH3N and are conserved in conventional SH3 domains, are replaced by polar residues -Q244, Q274 and H290 on the SH3C surface (Fig. 2), thereby rationalizing the drastically reduced affinity of this domain for proline rich sequences.The carboxyl-terminal region of Crk is considered inhibitory, since it contains elements that constrain binding to the N-terminal SH2 and...

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Tyrosine phosphorylation within the SH3 domain regulates CAS subcellular localization, cell migration, and invasiveness

Cited by 41 publications

References 47 publications

The ubiquitin-proteasome system regulates focal adhesions at the leading edge of migrating cells

The ubiquitin-proteasome system regulates focal adhesions at the leading edge of migrating cells

The role of focal adhesion anchoring domains of CAS in mechanotransduction

Commentary: The carboxyl‐terminal Crk SH3 domain: Regulatory strategies and new perspectives

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