Biopolym. Cell. 2016; 32(1):34-40.
http://dx.doi.org/10.7124/bc.00090A
Genomics, Transcriptomics and Proteomics
Focal adhesion kinase (FAK1) regulates SHB phosphorylation and its binding with a range of signaling proteins.
1Dergai O. V., 1Yaruchik A. M., 1Rynditch A. V.
  1. Institute of Molecular Biology and Genetics, NAS of Ukraine
    150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03680

Abstract

Aim. To investigate an effect of the Focal adhesion kinase 1 (FAK1) expression on the level of tyrosine phosphorylation of an adaptor protein SHB and to find functional consequences of this posttranslational modification. Methods. Recombinant DNA construction, protein expression and purification, human cell transfection, western blot. Results. The expression of FAK1 induces the massive tyrosine phosphorylation of SHB adaptor and enhances its interaction in vitro with SH2 domains of a range of the signaling proteins such as PI3K, ABL, CRK and PLCG1. Additionally we have found that Epstein-Barr virus protein LMP2A can partially mimic the FAK1-mediated effect strongly elevating the efficiency and SHB interaction with the mentioned above proteins. While the expression of individual proteins elevated SHB phosphorylation level, the co-expression of LMP2A and FAK1 did not display a synergetic effect. Conclusions. FAK1 as well as LMP2A induce the SHB tyrosine phosphorylation and enhance its interaction with a set of the signaling proteins.
Keywords: FAK1, SHB, LMP2A, phosphorylation
Full text: (PDF, in English)

References

[1] Mon NN, Ito S, Senga T, Hamaguchi M. FAK signaling in neoplastic disorders: a linkage between inflammation and cancer. Ann N Y Acad Sci. 2006;1086:199-212.
[2] Xia H, Nho RS, Kahm J, Kleidon J, Henke CA. Focal adhesion kinase is upstream of phosphatidylinositol 3-kinase/Akt in regulating fibroblast survival in response to contraction of type I collagen matrices via a beta 1 integrin viability signaling pathway. J Biol Chem. 2004;279(31):33024-34.
[3] Schaller MD. Biochemical signals and biological responses elicited by the focal adhesion kinase. Biochim Biophys Acta. 2001;1540(1):1-21.
[4] Chen HC, Guan JL. Association of focal adhesion kinase with its potential substrate phosphatidylinositol 3-kinase. Proc Natl Acad Sci U S A. 1994;91(21):10148-52.
[5] Chen HC, Appeddu PA, Isoda H, Guan JL. Phosphorylation of tyrosine 397 in focal adhesion kinase is required for binding phosphatidylinositol 3-kinase. J Biol Chem. 1996;271(42):26329-34.
[6] Nolan K, Lacoste J, Parsons JT. Regulated expression of focal adhesion kinase-related nonkinase, the autonomously expressed C-terminal domain of focal adhesion kinase. Mol Cell Biol. 1999;19(9):6120-9.
[7] Han DC, Guan JL. Association of focal adhesion kinase with Grb7 and its role in cell migration. J Biol Chem. 1999;274(34):24425-30.
[8] Holmqvist K, Cross M, Riley D, Welsh M. The Shb adaptor protein causes Src-dependent cell spreading and activation of focal adhesion kinase in murine brain endothelial cells. Cell Signal. 2003;15(2):171-9.
[9] Gustafsson K, Jamalpour M, Trinh C, Kharas MG, Welsh M. The Src homology-2 protein Shb modulates focal adhesion kinase signaling in a BCR-ABL myeloproliferative disorder causing accelerated progression of disease. J Hematol Oncol. 2014;7:45.
[10] Matskova LV, Helmstetter C, Ingham RJ, Gish G, Lindholm CK, Ernberg I, Pawson T, Winberg G. The Shb signalling scaffold binds to and regulates constitutive signals from the Epstein-Barr virus LMP2A membrane protein. Oncogene. 2007;26(34):4908-17.
[11] Cross MJ, Lu L, Magnusson P, Nyqvist D, Holmqvist K, Welsh M, Claesson-Welsh L. The Shb adaptor protein binds to tyrosine 766 in the FGFR-1 and regulates the Ras/MEK/MAPK pathway via FRS2 phosphorylation in endothelial cells. Mol Biol Cell. 2002;13(8):2881-93.
[12] Dergai O, Dergai M, Skrypkina I, Matskova L, Tsyba L, Gudkova D, Rynditch A. The LMP2A protein of Epstein-Barr virus regulates phosphorylation of ITSN1 and Shb adaptors by tyrosine kinases. Cell Signal. 2013;25(1):33-40.
[13] Novokhatska O, Dergai M, Tsyba L, Skrypkina I, Filonenko V, Moreau J, Rynditch A. Adaptor proteins intersectin 1 and 2 bind similar proline-rich ligands but are differentially recognized by SH2 domain-containing proteins. PLoS One. 2013;8(7):e70546.
[14] Golubovskaya VM, Zheng M, Zhang L, Li JL, Cance WG. The direct effect of focal adhesion kinase (FAK), dominant-negative FAK, FAK-CD and FAK siRNA on gene expression and human MCF-7 breast cancer cell tumorigenesis. BMC Cancer. 2009;9:280.
[15] Dergai OV, Dergai MV, Gudkova DO, Tsyba LO, Skrypkina IYa, Rynditch AV. Interaction of ubiquitin ligase CBL with LMP2A protein of Epstein-Barr virus occurs via PTB domain of CBL and does not depend on adaptor ITSN1. Biopolym Cell. 2013; 29(2): 131–5.
[16] Xue Y, Ren J, Gao X, Jin C, Wen L, Yao X. GPS 2.0, a tool to predict kinase-specific phosphorylation sites in hierarchy. Mol Cell Proteomics. 2008;7(9):1598-608.
[17] Pawson T, Gish GD, Nash P. SH2 domains, interaction modules and cellular wiring. Trends Cell Biol. 2001;11(12):504-11.
[18] Welsh M, Jamalpour M, Zang G, Åkerblom B. The role of the Src Homology-2 domain containing protein B (SHB) in β cells. J Mol Endocrinol. 2016;56(1):R21-31.
[19] Gesbert F, Sellers WR, Signoretti S, Loda M, Griffin JD. BCR/ABL regulates expression of the cyclin-dependent kinase inhibitor p27Kip1 through the phosphatidylinositol 3-Kinase/AKT pathway. J Biol Chem. 2000;275(50):39223-30.
[20] Hägerkvist R, Mokhtari D, Lindholm C, Farnebo F, Mostoslavsky G, Mulligan RC, Welsh N, Welsh M. Consequences of Shb and c-Abl interactions for cell death in response to various stress stimuli. Exp Cell Res. 2007;313(2):284-91.
[21] Lu J, Lin WH, Chen SY, Longnecker R, Tsai SC, Chen CL, Tsai CH. Syk tyrosine kinase mediates Epstein-Barr virus latent membrane protein 2A-induced cell migration in epithelial cells. J Biol Chem. 2006;281(13):8806-14.
[22] Fotheringham JA, Coalson NE, Raab-Traub N. Epstein-Barr virus latent membrane protein-2A induces ITAM/Syk- and Akt-dependent epithelial migration through αv-integrin membrane translocation. J Virol. 2012;86(19):10308-20.