The role of the linker between the SH2 domain and catalytic domain in the regulation and function of Src

Gonfloni, Stefania; Williams, John C.; Hattula, Katarina; Weijland, Albert; Wierenga, Rik K.; Superti‐Furga, Giulio

doi:10.1093/emboj/16.24.7261

Cited by 142 publications

(136 citation statements)

References 48 publications

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“…The cDNA clones pSGT-Src, pSGT-Src(K249E/P250G) [pSGTSrc(KP)], and pSGT-Src(Y416F) were described before (16,17,39). The plasmid pCA-Lyn was a kind gift of Peter Klinken (51).…”

Section: Plasmidsmentioning

confidence: 99%

Deciphering the Cross Talk between hnRNP K and c-Src: the c-Src Activation Domain in hnRNP K Is Distinct from a Second Interaction Site

Adolph

Flach

Mueller

et al. 2007

Molecular and Cellular Biology

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The protein tyrosine kinase c-Src is regulated by two intramolecular interactions. The repressed state is achieved through the interaction of the Src homology 2 (SH2) domain with the phosphorylated C-terminal tail and the association of the SH3 domain with a polyproline type II helix formed by the linker region between SH2 and the kinase domain. hnRNP K, the founding member of the KH domain protein family, is involved in chromatin remodeling, regulation of transcription, and translation of specific mRNAs and is a target in different signal transduction pathways. In particular, it functions as a specific activator and a substrate of the tyrosine kinase c-Src. Here we address the question how hnRNP K interacts with and activates c-Src. We define the proline residues in hnRNP K in the proline-rich motifs P2 (amino acids [aa] 285 to 297) and P3 (aa 303 to 318), which are necessary and sufficient for the specific activation of c-Src, and we dissect the amino acid sequence (aa 216 to 226) of hnRNP K that mediates a second interaction with c-Src. Our findings indicate that the interaction with c-Src and the activation of the kinase are separable functions of hnRNP K. hnRNP K acts as a scaffold protein that integrates signaling cascades by facilitating the cross talk between kinases and factors that mediate nucleic acid-directed processes.

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“…The cDNA clones pSGT-Src, pSGT-Src(K249E/P250G) [pSGTSrc(KP)], and pSGT-Src(Y416F) were described before (16,17,39). The plasmid pCA-Lyn was a kind gift of Peter Klinken (51).…”

Section: Plasmidsmentioning

confidence: 99%

Deciphering the Cross Talk between hnRNP K and c-Src: the c-Src Activation Domain in hnRNP K Is Distinct from a Second Interaction Site

Adolph

Flach

Mueller

et al. 2007

Molecular and Cellular Biology

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“…In addition, p56 lck kinase activity is regulated through intramolecular interaction of the SH3 domain with a proline-rich motif in the SH2-kinase linker region. This interaction maintains a conformation in which the kinase domain is inaccessible (15).…”

mentioning

confidence: 99%

SOCS-6 Negatively Regulates T Cell Activation through Targeting p56 to Proteasomal Degradation

Choi

Son

Park

et al. 2010

Journal of Biological Chemistry

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The T cell-specific tyrosine kinase, p56 lck , plays crucial roles in T cell receptor (TCR)-mediated T cell activation. Here, we report that SOCS-6 (suppressor of cytokine signaling-6) is a negative regulator of p56 lck . SOCS-6 was identified as a protein binding to the kinase domain of p56 lck through yeast two-hybrid screening. SOCS-6 bound specifically to p56 lck (F505), which mimics the active form of p56 lck , but not to wild type p56 lck . In Jurkat T cells, SOCS-6 binding to p56 lck was detected 1-2 h after TCR stimulation. Confocal microscopy showed that upon APC-T cell conjugation, SOCS-6 was recruited to the immunological synapse and colocalized with the active form of p56 lck . SOCS-6 promoted p56lck ubiquitination and its subsequent targeting to the proteasome. Moreover, SOCS-6 overexpression led to repression of TCR-dependent interleukin-2 promoter activity. These results establish that SOCS-6 acts as a negative regulator of T cell activation by promoting ubiquitin-dependent proteolysis.The T lymphocyte-specific member of the Src-type tyrosine kinase family, p56 lck , plays essential roles in development, antigen-induced T cell activation, and proliferation (1-3). During TCR 4 -mediated T cell activation, p56 lck is activated and transmits a positive signal by interacting with CD4/CD8 (4, 5). Upon engagement of the antigen-major histocompatibility complex with TCR-CD4-CD8 complexes, p56 lck phosphorylates the immunoreceptor tyrosine-based activation motifs of the CD3 -chain and provides the binding site for ZAP-70 tyrosine kinase (6). Subsequent ZAP-70 activation leads to phosphorylation of signaling proteins, including LAT, and amplification of TCR-mediated signaling (7,8). In this process, p56 lck migrates to T cell-APC contact regions known as immunological synapses (IS) and thereby enhances TCR signaling (9, 10). Given the importance of p56 lck function in T cell activation, establishing how p56 lck activity is regulated is essential to understand that activation.The structure of p56 lck is similar to that of other Src family kinases and exhibits the following common domains: N-terminal attachment sites for saturated fatty acid addition, a unique region; an Src homology 3 (SH3) domain; an SH2 domain; a tyrosine kinase domain (SH1); and a C-terminal negative regulatory site, tyrosine 505 (Tyr-505) (11). p56 lck kinase activity is regulated by intramolecular interactions through its regulatory domains and/or phosphorylation (11). For example, phosphorylation of Tyr-394 in the activation loop of the kinase domain increases enzymatic activity, whereas phosphorylation of Tyr-505 renders the enzyme less active. Phosphorylated Tyr-505 interacts with its own SH2 domain, promoting a closed conformation and maintaining the kinase in an inactive state (12). Phenylalanine substitution of the regulatory tyrosine residue (Y505F) results in a constitutively active form of the protein.The status of the regulatory tyrosine is maintained both by a kinase, Csk, which phosphorylates it (13), and by a transmembrane...

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“…When phosphorylated, Tyr 505 interacts with the SH2 domain of Lck, which effectively suppresses tyrosine kinase activity (10 -13). Interactions between the SH3 domain and the linker connecting the SH2 domain to the catalytic domain build up the clamp and play an additional essential role in stabilizing the autoinhibited state (14). Suppressive phosphorylation of Lck at Tyr 505 is accomplished by the tyrosine kinase C-terminal Src kinase (Csk) (15).…”

mentioning

confidence: 99%

Genetically Encoded Förster Resonance Energy Transfer Sensors for the Conformation of the Src Family Kinase Lck

Paster

Paar

Eckerstorfer

et al. 2009

The Journal of Immunology

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The current model for regulation of the Src family kinase member Lck postulates a strict correlation between structural condensation of the kinase backbone and catalytic activity. The key regulatory tyrosine 505, when phosphorylated, interacts with the Src homology 2 domain on the same molecule, effectively suppressing tyrosine kinase activity. Dephosphorylation of Tyr 505 upon TCR engagement is supposed to lead to unfolding of the kinase structure and enhanced kinase activity. Studies on the conformation-activity relationship of Lck in living cells have not been possible to date because of the lack of tools providing spatiotemporal resolution of conformational changes. We designed a biochemically active, conformation-sensitive Förster resonance energy transfer biosensor of human Lck using the complete kinase backbone. Live cell imaging in Jurkat cells demonstrated that our biosensor performed according to Src family kinase literature. A Tyr 505 to Phe mutation opened the structure of the Lck sensor, while changing the autophosphorylation site Tyr 394 to Phe condensed the molecule. The tightly packed structure of a high-affinity YEEI tail mutant showed that under steady-state conditions the bulk of Lck molecules exist in a mean conformational configuration. Although T cell activation commenced normally, we could not detect a change in the conformational status of our Lck biosensor during T cell activation. Together with biochemical data we conclude that during T cell activation, Lck is accessible to very subtle regulatory mechanisms without the need for acute changes in Tyr 505 and Tyr 394 phosphorylation and conformational alterations.

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The role of the linker between the SH2 domain and catalytic domain in the regulation and function of Src

Cited by 142 publications

References 48 publications

Deciphering the Cross Talk between hnRNP K and c-Src: the c-Src Activation Domain in hnRNP K Is Distinct from a Second Interaction Site

Deciphering the Cross Talk between hnRNP K and c-Src: the c-Src Activation Domain in hnRNP K Is Distinct from a Second Interaction Site

SOCS-6 Negatively Regulates T Cell Activation through Targeting p56 to Proteasomal Degradation

Genetically Encoded Förster Resonance Energy Transfer Sensors for the Conformation of the Src Family Kinase Lck

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