Lipid binding by the Unique and SH3 domains of c-Src suggests a new regulatory mechanism

Pérez, Yolanda; Maffei, Mariano; Igea, Ana; Amata, Irene; Gairì, Margarida; Nebreda, Ángel R.; Bernadó, Pau; Pons, Miquel

doi:10.1038/srep01295

Cited by 85 publications

(145 citation statements)

References 48 publications

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“…We examined the role of Src's Unique domain, a nonconserved region within SFKs that is thought to mediate SFKlipid interactions (40,41 …”

Section: Resultsmentioning

confidence: 99%

Engineered kinase activation reveals unique morphodynamic phenotypes and associated trafficking for Src family isoforms

Chu¹,

Tsygankov²,

Berginski³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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The Src kinase family comprises nine homologous members whose distinct expression patterns and cellular distributions indicate that they have unique roles. These roles have not been determined because genetic manipulation has not produced clearly distinct phenotypes, and the kinases' homology complicates generation of specific inhibitors. Through insertion of a modified FK506 binding protein (insertable FKBP12, iFKBP) into the protein kinase isoforms Fyn, Src, Lyn, and Yes, we engineered kinase analogs that can be activated within minutes in living cells (RapR analogs). Combining our RapR analogs with computational tools for quantifying and characterizing cellular dynamics, we demonstrate that Src family isoforms produce very different phenotypes, encompassing cell spreading, polarized motility, and production of long, thin cell extensions. Activation of Src and Fyn led to patterns of kinase translocation that correlated with morphological changes in temporally distinct stages. Phenotypes were dependent on N-terminal acylation, not on Src homology 3 (SH3) and Src homology 2 (SH2) domains, and correlated with movement between a perinuclear compartment, adhesions, and the plasma membrane.image analysis | motion classification | live cell imaging | protein engineering | rapamycin

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“…We examined the role of Src's Unique domain, a nonconserved region within SFKs that is thought to mediate SFKlipid interactions (40,41 …”

Section: Resultsmentioning

confidence: 99%

Engineered kinase activation reveals unique morphodynamic phenotypes and associated trafficking for Src family isoforms

Chu¹,

Tsygankov²,

Berginski³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…The N-terminal region of c-Src, including the SH4 and the UD, is intrinsically disordered. Detailed NMR study of this N-terminal region suggested transient secondary structural elements between residues 60–64 and 67–74 230 and revealed that SH4 and UD play a crucial role in the regulation of c-Src by participating in a number of intra- and intermolecular interactions 229 . The UD and SH3 domain bind lipids and interact with each other, but binding of poly-proline peptides to the SH3 domain allosterically inhibits its interaction with the UD.…”

Section: Dynamic Complexes Multivalent Interactions and Dynamic Imentioning

confidence: 99%

Dynamic Protein Interaction Networks and New Structural Paradigms in Signaling

et al. 2016

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Understanding signaling and other complex biological processes requires elucidating the critical roles of intrinsically disordered proteins and regions (IDPs/IDRs), which represent ~30% of the proteome and enable unique regulatory mechanisms. In this review we describe the structural heterogeneity of disordered proteins that underpins these mechanisms and the latest progress in obtaining structural descriptions of ensembles of disordered proteins that are needed for linking structure and dynamics to function. We describe the diverse interactions of IDPs that can have unusual characteristics such as “ultrasensitivity” and “regulated folding and unfolding”. We also summarize the mounting data showing that large-scale assembly and protein phase separation occurs within a variety of signaling complexes and cellular structures. In addition, we discuss efforts to therapeutically target disordered proteins with small molecules. Overall, we interpret the remodeling of disordered state ensembles due to binding and post-translational modifications within an expanded framework for allostery that provides significant insights into how disordered proteins transmit biological information.

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“…c-Src is the leading member of the Src family of non-receptor tyrosine kinases (SFKs) that are expressed in many cells and tissues [5, 6], and are involved in diverse signal transduction pathways [7, 8]. All members of the SFKs possess similar domain arrangements consisting of src homology 3 (SH3), SH2 and kinase (SH1) domains with a common myristoylated and/or palmitoylated membrane–anchoring N-terminal region known as the SH4 domain [9, 10] and a unique domain [11]. Regulation of c-Src activity is crucial for its biological functions.…”

Section: Introductionmentioning

confidence: 99%

Activation of c-Src: A hub for exogenous pro-oxidant-mediated activation of Toll-like receptor 4 signaling

Karki

Zhang

Igwe

2014

Free Radical Biology and Medicine

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Summary To study the role of c-Src kinase in prooxidant-induced stimulation of TLR4, we used LPS-EK and MPLA as TLR4 specific agonists and positive controls, and SIN-1 and PPC as prooxidant sources. We used HEK-Blue mTLR4 cell line that is stably transfected with mouse TLR4 and that expresses optimized SEAP reporter under the control of a promoter inducible by NF-κB transcription factor. The level of SEAP released due to TLR4 stimulation was a measure of NF-κB activation. Treatment with either the prooxidants or LPS-EK increased SEAP release and TNF-α production in these cells. These treatments also increased intracellular ROS accumulation with an enhanced production of nitric oxide and TBARS to confirm oxidant stress in these cells. Pretreatment with c-Src kinase inhibitors, PP2 and CA-pY, which act by different mechanisms, decreased these parameters. Pretreatment with SSG, a c-Src activator, enhanced the effects promoted by LPS-EK and prooxidants, and rescued cells from PP2- and Ca-pY-induced effects. Curiously, prooxidants but not TLR4 agonist increased the ratio of TNFα to IL-10 released suggesting that prooxidants can initiate and maintain an imbalance of TNFα production over IL-10. To different degrees, both prooxidant and TLR4 agonist increased formation of c-Src complexes with TLR4 and IκB-α as coimmunoprecipitates. Both prooxidant and TLR4 agonist increased c-Src phosphorylation of Tyr-42 residue in IκB-α, but prooxidant-induced effect was more robust and much longer lasting. Taken together, these studies provide a mechanism whereby c-Src assumes a central role in prooxidant-induced NF-κB activation in TLR4 signaling. Prooxidant-induced activation of TLR4 through c-Src/NFκB/IκB-α coupling provides a basis for a molecular dissection of the initiation and maintenance of sterile inflammation that may serve as a “pathophysiologic primer” for many diseases.

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Lipid binding by the Unique and SH3 domains of c-Src suggests a new regulatory mechanism

Cited by 85 publications

References 48 publications

Engineered kinase activation reveals unique morphodynamic phenotypes and associated trafficking for Src family isoforms

Engineered kinase activation reveals unique morphodynamic phenotypes and associated trafficking for Src family isoforms

Dynamic Protein Interaction Networks and New Structural Paradigms in Signaling

Activation of c-Src: A hub for exogenous pro-oxidant-mediated activation of Toll-like receptor 4 signaling

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