The SH4-Unique-SH3-SH2 domains dictate specificity in signaling that differentiate c-Yes from c-Src

Department of Microbiology, Immunology, and Cell Biology, West Virginia University School of Medicine, PO Box 9300, Morgantown 26506, USA.
Journal of Cell Science (Impact Factor: 5.43). 07/2003; 116(Pt 12):2585-98. DOI: 10.1242/jcs.00466
Source: PubMed


c-Src and c-Yes are highly homologous members of the Src family of non-receptor tyrosine kinases. The overall sequence similarity between c-Src and c-Yes allows them to perform many overlapping functions. However, the phenotypes of the c-src and c-yes knockout mice, and cells derived from them, are quite different, indicating functional specificity between the two proteins. Specifically, c-src-/- cells are deficient in several processes that require dynamic regulation of the actin cytoskeleton. In order to begin to understand why c-Yes is unable to compensate for c-Src signaling, we used a series of Src/Yes chimeras in which the non-catalytic functional domains of Src527F were replaced by those of c-Yes. Using chicken embryo fibroblasts as a model system, our results indicate that the c-Yes N-terminal SH4-Unique domains are sufficient to inhibit the ability of Src527F to alter cell morphology, induce actin filament rearrangements or stimulate motility or invasive potential. The data also indicate that the SH4-Unique-SH3-SH2 domains of c-Yes work cooperatively and prevent activation of signaling proteins associated with Src527F transformation, including activation of phosphatidylinositol 3-kinase, phosphorylation of c-Raf and Akt and downregulation of RhoA-GTP. These data indicate that c-Yes may not modulate signals associated with c-Src-induced changes in actin filament integrity and may explain why c-Yes fails to compensate for c-Src signaling in src-/- cells.

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Available from: Xianglin Shi, Aug 19, 2014
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    • "Several SFKs contain residues in the N-terminal region that are phosphorylated and dephosphorylated in cellular processes (Joung et al., 1995; Hansen et al., 1997; Johnson et al., 2000). Moreover, swapping the Unique domains of Src and Yes interchanges the functional specificity of the two SFKs (Hoey et al., 2000; Summy et al., 2003; Werdich and Penn, 2005). The versatility and relevance of the active role of the Unique domain in Src function was confirmed by the discovery of binding by the Unique domain to different targets, such as acidic lipids, the SH3 domain, and calmodulin (Pérez et al., 2013). "
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    ABSTRACT: Members of the Src family of kinases (SFKs) are non-receptor tyrosine kinases involved in numerous signal transduction pathways. The catalytic, SH3 and SH2 domains are attached to the membrane-anchoring SH4 domain through the intrinsically disordered "Unique" domains, which exhibit strong sequence divergence among SFK members. In the last decade, structural and biochemical studies have begun to uncover the crucial role of the Unique domain in the regulation of SFK activity. This mini-review discusses what is known about the phosphorylation events taking place on the SFK Unique domains, and their biological relevance. The modulation by phosphorylation of biologically relevant inter- and intra- molecular interactions of Src, as well as the existence of complex phosphorylation/dephosphorylation patterns observed for the Unique domain of Src, reinforces the important functional role of the Unique domain in the regulation mechanisms of the Src kinases and, in a wider context, of intrinsically disordered regions in cellular processes.
    Frontiers in Genetics 06/2014; 5:181. DOI:10.3389/fgene.2014.00181
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    • "The SFKs are highly homologous non-receptor cytoplasmic tyrosine kinases that modulate diverse cellular processes including adhesion, migration, division, death and survival [1]–[5]. Dysregulation of individual SFKs, including Lyn, occurs in several different types of tumor [4], [6]–[9]. Although the functions of SFKs appear to be influenced by the microenvironment as well as cell type and post-translational modifications [4], [6]–[9], little attention has been paid to the role of SFKs in promoting cell survival through regulation of autophagy. "
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    ABSTRACT: Members of the Src family kinases (SFK) can modulate diverse cellular processes, including division, death and survival, but their role in autophagy has been minimally explored. Here, we investigated the roles of Lyn, a SFK, in promoting the survival of human glioblastoma tumor (GBM) cells in vitro and in vivo using lentiviral vector-mediated expression of constitutively-active Lyn (CA-Lyn) or dominant-negative Lyn (DN-Lyn). Expression of either CA-Lyn or DN-Lyn had no effect on the survival of U87 GBM cells grown under nutrient-rich conditions. In contrast, under nutrient-deprived conditions (absence of supplementation with L-glutamine, which is essential for growth of GBM cells, and FBS) CA-Lyn expression enhanced survival and promoted autophagy as well as inhibiting cell death and promoting proliferation. Expression of DN-Lyn promoted cell death. In the nutrient-deprived GBM cells, CA-Lyn expression enhanced AMPK activity and reduced the levels of pS6 kinase whereas DN-Lyn enhanced the levels of pS6 kinase. Similar results were obtained in vitro using another cultured GBM cell line and primary glioma stem cells. On propagation of the transduced GBM cells in the brains of nude mice, the CA-Lyn xenografts formed larger tumors than control cells and autophagosomes were detectable in the tumor cells. The DN-Lyn xenografts formed smaller tumors and contained more apoptotic cells. Our findings suggest that on nutrient deprivation in vitro Lyn acts to enhance the survival of GBM cells by promoting autophagy and proliferation as well as inhibiting cell death, and Lyn promotes the same effects in vivo in xenograft tumors. As the levels of Lyn protein or its activity are elevated in several cancers these findings may be of broad relevance to cancer biology.
    PLoS ONE 08/2013; 8(8):e70804. DOI:10.1371/journal.pone.0070804 · 3.23 Impact Factor
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    • "RecombinantSTYkinasewasincubatedwithrecombinantsubstrateinthe presenceof3mCiof[ 32 P]ATPand2.5mMATPinatotalvolumeof50mLof kinasebuffer(20mMTris-HCl,pH7.5,5mMMgCl2,and0.5mMMnCl2).Exact amountsofkinaseandsubstrateproteinsusedfortheindividualexperiments aregiveninthefigurelegends.Thereactionwascarriedoutfor10minat23°C andstoppedbyadding12mLofSDSsamplebuffer.Kinasereactionswith CPK4wereaccomplishedinkinasebuffercontaining7mMCaCl2and1mM DTTinsteadofMnCl2,andYeskinasereactionswereperformedin20mM MOPS,pH7.9,and5mMMgCl2.PurifiedYeskinase(Summyetal.,2003)was polyacrylamidegelfollowedbyautoradiography.Fortheinhibitionstudies, theSTYkinasewaspreincubatedfor10minwiththeinhibitorsJNJ-10198409, tyrphostin,andJanex1(Cayman)attheindicatedconcentrations.Thereaction wasthencarriedoutasdescribedbefore.Thereactionproductwasseparated ona12%SDS-polyacrylamidegel,theCoomassieblue-stainedproteinbands werecutoutfromthegel,andtheradioactivitywasdeterminedbyliquid scintillationcounting. "
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    ABSTRACT: In Arabidopsis (Arabidopsis thaliana), transit peptides for chloroplast-destined preproteins can be phosphorylated by the protein kinases STY8, STY17, and STY46. In this study, we have investigated the in vitro properties of these plant-specific kinases. Characterization of the mechanistic functioning of STY8 led to the identification of an essential threonine in the activation segment, which is phosphorylated by an intramolecular mechanism. STY8 is inhibited by specific tyrosine kinase inhibitors, although it lacked the ability to phosphorylate tyrosine residues in vitro. In vivo analysis of sty8, sty17, and sty46 Arabidopsis knockout/knockdown mutants revealed a distinct function of the three kinases in the greening process and in the efficient differentiation of chloroplasts. Mutant plants displayed not only a delayed accumulation of chlorophyll but also a reduction of nucleus-encoded chloroplast proteins and a retarded establishment of photosynthetic capacity during the first 6 h of deetiolation, supporting a role of cytosolic STY kinases in chloroplast differentiation.
    Plant physiology 07/2011; 157(1):70-85. DOI:10.1104/pp.111.182774 · 6.84 Impact Factor
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