Distinct roles for Crk adaptor isoforms in actin reorganization induced by extracellular signals

Antoku, Susumu; Mayer, Bruce J.

doi:10.1242/jcs.054627

Cited by 54 publications

(73 citation statements)

References 61 publications

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“…The SH2 and SH3 domains of CRKII and CRKL are highly conserved, their interaction specificities are almost identical, and redundancy has been reported in other pathways (14,43,44). However, there is also evidence for unique functions, since mice singly deficient for either CRK or CRKL exhibit severe developmental defects (10,11).…”

Section: Figure 7 Crk/crkl-deficient T Cells Have Defects In Migratimentioning

confidence: 98%

CRK proteins selectively regulate T cell migration into inflamed tissues

Huang¹,

Clarke²,

Karimi³

et al. 2015

J. Clin. Invest.

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R e s e a R c h a R t i c l e1 0 2 0 jci.org Volume 125 Number 3 March 2015 therapeutic implications, since they can carry out graftversus-leukemia (GVL) responses with minimal GVHD. Results Generation and characterization of T cell-specific CRK T cells (data not shown).Western blotting of purified CD4 + T cells from Dko and WT mice revealed that levels of CRKI, CRKII, and CRKL in the mutant T cells were reduced by at least 95% ( Figure 1A and data not shown), and flow cytometric analysis confirmed loss of CRK protein expression (Figure 1, B and C).CRK/CRKL Dko mice were born at Mendelian ratios, and spleen, thymus, and lymph nodes exhibited normal cellularity (data not shown). Thymocyte populations were similar in WT and Dko mice, indicating that T cell development proceeded normally ( Figure 1D). Peripheral lymphoid organs showed no differences in proportions of CD4 + and CD8 + T cells, naive (CD62L hi CD44 lo ), memory (CD62L lo CD44 hi ), or activated (CD69 hi ) T cell subsets (Figure 1, D and E, and data not shown). Thus, these mice represented a suitable source of mature CRK/CRKL Dko T cells for functional studies. CRK/CRKL-deficient T cells show defects in adhesion and polarization.Since CRK proteins control adhesion in non-hematopoietic cells (4), we first asked whether integrin-dependent adhesion is defective in CRK/CRKL Dko T cells. On plates coated with ICAM-1, the ligand for the β 2 integrin LFA-1, WT preactivated CD4 + T cells showed about 10% basal binding, which was increased 2-to 5-fold after stimulation with the chemokines CXCL12 or CCL21, or anti-CD3 (Figure 2A). CRK/CRKL Dko T cells showed a significant reduction in adhesion to ICAM-1 under both basal and stimulated conditions. Indeed, chemokine stimulation induced almost no increased adhesion in these cells. Treatment with PMA bypassed the defect. This is most likely because the defect in CRK/CRKL Dko cells lies upstream of PKC signaling in the pathway leading to integrin activation, though PKC activation could also drive a parallel

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Section: Figure 7 Crk/crkl-deficient T Cells Have Defects In Migratimentioning

confidence: 98%

CRK proteins selectively regulate T cell migration into inflamed tissues

Huang¹,

Clarke²,

Karimi³

et al. 2015

J. Clin. Invest.

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“…These results suggest that although JNK phosphorylation dramatically declines in the absence of Crk and CrkL, activation of JNKs alone is not sufficient to drive cell proliferation in the absence of Crk and CrkL. required for PDGF-stimulated remodeling of the actin cytoskeleton and cell migration (44). At first we cultured fibroblasts in low serum (0.5% FBS) for 4 days.…”

Section: Independent Roles Of Crk or Crkl In Cell Growth-because Tranmentioning

confidence: 99%

Fibroblast Growth Requires CT10 Regulator of Kinase (Crk) and Crk-like (CrkL)

Park

Koptyra

Curran

2016

Journal of Biological Chemistry

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Edited by Xiao-Fan WangThe chicken tumor virus oncoprotein, v-Crk, induces a substantial increase in protein tyrosine phosphorylation through protein-protein interactions mediated by its SH2 2 and SH3 domains, leading to cell transformation (1). CrkII and Crk-like (CrkL) represent closely related cellular homologues of v-Crk and have similar structures and functions (2). CrkI is a splice variant of CrkII that lacks the C-terminal SH3 domain. Crk family proteins (CrkI, CrkII, and CrkL) interact with many proteins through their SH2 and SH3 domains. Overexpression of Crk family proteins induces transformation of cultured cells (3,4). CrkI is a more potent transforming gene than CrkII and CrkL in part because, like v-Crk, it lacks a C-terminal regulatory phosphorylation site (3). Overexpression of Crk and CrkL has been reported in several human cancers, including oral squamous cell carcinoma (5), ovarian carcinoma (6), colon cancer (7), lung cancer (8, 9), breast cancer (10, 11), gastric cancer (12), and glioblastoma (13,14). Reduced expression of either Crk or CrkL by RNA interference-mediated knockdown lowered the in vivo tumor formation of human ovarian (15), synovial sarcoma (16), glioblastoma (14), breast cancer (10), head and neck squamous cell carcinoma (17), and rhabdomyosarcoma (18) cell lines. Taken together, these reports imply that elevated levels of Crk family proteins promote cell transformation and enhance tumor cell growth (for review see Refs. 19 and 20).To investigate functions of endogenous Crk and CrkL in biological processes at the cellular level, we developed mouse strains and cell lines harboring individual and combined floxed alleles of Crk and CrkL. Application of the Cre-loxP recombination-based conditional knock-out approach to cultured fibroblasts enabled us to demonstrate that endogenous Crk and CrkL are important for maintaining cell structural integrity and proper cell motility (21). We also discovered that Crk and CrkL are required for cell transformation induced by viral oncogenes (22). Here we utilized the conditional knock-out system to examine whether endogenous Crk and CrkL are required for cell growth. Our findings clearly demonstrate that Crk and CrkL play essential, overlapping roles in fibroblast proliferation by enabling cells to progress from the G 1 phase to the S phase. Our study also demonstrates that expression of any one protein among CrkI, CrkII, and CrkL at physiological levels is sufficient to secure cell proliferation. ResultsCrk and CrkL Are Essential for Fibroblast Proliferation-We used the Neon system (Life Technologies) to achieve rapid and efficient gene expression in growing fibroblasts by transduction of synthetic mRNA (synRNA). To reduce innate immune responses and increase ectopic protein expression, we synthesized mRNA using the modified ribonucleotides, pseudo-UTP, and methyl-CTP (23,24). When fibroblasts immortalized by T antigen or the 3T3 protocol were transfected with synthetic green fluorescent protein mRNA (synGFP), both cell types in monolayer cul...

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“…The conserved sequence P 248 NAY in the RT loop combined with some mutations might play a role in the activation of Abl kinase (Reichman et al 2005). Since CrkII also carries the SH2 domain which recruits downstream targets and the Abl family of kinases, a delayed feedback loop is formed (Antoku and Mayer 2009). Abl interactor proteins Abi1 and Abi2 regulate c-Abl kinase activity by interacting with the C-terminal PxxP sequences and c-Abl SH3/SH2 domains.…”

Section: Regulationmentioning

confidence: 99%

“…Such vital processes as migration and invasiveness (Yamada et al 2011), actin reorganization induced by extracellular signals (Antoku and Mayer 2009), shaping spines (Sheng and Kim 2000;Ehlers 2002), among others take a central role when new treatments rely on an understanding of cellular control mechanisms.…”

Section: Regulationmentioning

confidence: 99%

SH3 domains: modules of protein–protein interactions

2012

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Src homology 3 (SH3) domains are involved in the regulation of important cellular pathways, such as cell proliferation, migration and cytoskeletal modifications. Recognition of polyproline and a number of noncanonical sequences by SH3 domains has been extensively studied by crystallography, nuclear magnetic resonance and other methods. High-affinity peptides that bind SH3 domains are used in drug development as candidates for anticancer treatment. This review summarizes the latest achievements in deciphering structural determinants of SH3 function.

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Distinct roles for Crk adaptor isoforms in actin reorganization induced by extracellular signals

Cited by 54 publications

References 61 publications

CRK proteins selectively regulate T cell migration into inflamed tissues

CRK proteins selectively regulate T cell migration into inflamed tissues

Fibroblast Growth Requires CT10 Regulator of Kinase (Crk) and Crk-like (CrkL)

SH3 domains: modules of protein–protein interactions

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