Biopolym. Cell. 2002; 18(4):312-318.
Subcellular localization of adapter protein Ruk, in HEK293 cells
- Institute of Cell Biology, NAS of Ukraine
14/16, Drahomanov Str., Lviv, Ukraine, 79005
- Institute of Molecular Biology and Genetics, NAS of Ukraine
150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03680
- Ludwig Institute for Cancer Research
91 Riding house str., London, UK, WIP8BT
- The Universuty of Edinburgh
Subcellular localization of adapter protein Ruk1 has been investigated in human embryonic kidney HEK293 cells transfected with pRc/CMV2/Ruk1-Glu-tag. Using immunofluorescence microscopy, it was shown that the recombinant protein was distributed diffusely in both cytoplasm and nucleus, but with punctated structures in the nucleus, which correspond in all probability to the nucleolus. The Ruk1 localization in the nucleus was confirmed by Western blotting of nucleic extracts prepared from transfected HEK293 cells using monoclonal anti-Glu-tag antibodies, as well as from nontransfected HEK293 and U937 cells using polyclonal anti-Ruk antibodies. The ability of affine purified Ruk1 Glu-tagged preparation to bind to DNA from calf thymus, but not to Escherichia coli DNA, was revealed by the immunodot-blot analysis. The nuclear localization of Ruk1 suggests that this adapter protein is involved in some new, yet unrecognized functions, in the eukaryotic cell nucleus.
 Pawson T, Schlessingert J. SH2 and SH3 domains. Curr Biol. 1993;3(7):434-42.
 Cicchetti P, Mayer BJ, Thiel G, Baltimore D. Identification of a protein that binds to the SH3 region of Abl and is similar to Bcr and GAP-rho. Science. 1992;257(5071):803-6.
 Rickles RJ, Botfield MC, Weng Z, Taylor JA, Green OM, Brugge JS, Zoller MJ. Identification of Src, Fyn, Lyn, PI3K and Abl SH3 domain ligands using phage display libraries. EMBO J. 1994;13(23):5598-604.
 Sparks AB, Rider JE, Hoffman NG, Fowlkes DM, Quillam LA, Kay BK. Distinct ligand preferences of Src homology 3 domains from Src, Yes, Abl, Cortactin, p53bp2, PLCgamma, Crk, and Grb2. Proc Natl Acad Sci U S A. 1996;93(4):1540-4.
 Pawson T. Protein modules and signalling networks. Nature. 1995;373(6515):573-80.
 Cohen GB, Ren R, Baltimore D. Modular binding domains in signal transduction proteins. Cell. 1995;80(2):237-48.
 Sahr KE, Laurila P, Kotula L, Scarpa AL, Coupal E, Leto TL, Linnenbach AJ, Winkelmann JC, Speicher DW, Marchesi VT, et al. The complete cDNA and polypeptide sequences of human erythroid alpha-spectrin. J Biol Chem. 1990;265(8):4434-43.
 Wu H, Parsons JT. Cortactin, an 80/85-kilodalton pp60src substrate, is a filamentous actin-binding protein enriched in the cell cortex. J Cell Biol. 1993;120(6):1417-26.
 Pawson T, Scott JD. Signaling through scaffold, anchoring, and adaptor proteins. Science. 1997;278(5346):2075-80.
 Pleiman CM, Hertz WM, Cambier JC. Activation of phosphatidylinositol-3' kinase by Src-family kinase SH3 binding to the p85 subunit. Science. 1994;263(5153):1609-12.
 de Mendez I, Homayounpour N, Leto TL. Specificity of p47phox SH3 domain interactions in NADPH oxidase assembly and activation. Mol Cell Biol. 1997;17(4):2177-85.
 Gout I, Dhand R, Hiles ID, Fry MJ, Panayotou G, Das P, Truong O, Totty NF, Hsuan J, Booker GW, et al. The GTPase dynamin binds to and is activated by a subset of SH3 domains. Cell. 1993;75(1):25-36.
 Bar-Sagi D, Rotin D, Batzer A, Mandiyan V, Schlessinger J. SH3 domains direct cellular localization of signaling molecules. Cell. 1993;74(1):83-91.
 Galisteo ML, Chernoff J, Su YC, Skolnik EY, Schlessinger J. The adaptor protein Nck links receptor tyrosine kinases with the serine-threonine kinase Pak1. J Biol Chem. 1996;271(35):20997-1000.
 McPherson PS. Regulatory role of SH3 domain-mediated protein-protein interactions in synaptic vesicle endocytosis. Cell Signal. 1999;11(4):229-38.
 Ribon V, Herrera R, Kay BK, Saltiel AR. A role for CAP, a novel, multifunctional Src homology 3 domain-containing protein in formation of actin stress fibers and focal adhesions. J Biol Chem. 1998;273(7):4073-80.
 Dustin ML, Olszowy MW, Holdorf AD, Li J, Bromley S, Desai N, Widder P, Rosenberger F, van der Merwe PA, Allen PM, Shaw AS. A novel adaptor protein orchestrates receptor patterning and cytoskeletal polarity in T-cell contacts. Cell. 1998;94(5):667-77.
 Kirsch KH, Georgescu MM, Ishimaru S, Hanafusa H. CMS: an adapter molecule involved in cytoskeletal rearrangements. Proc Natl Acad Sci U S A. 1999;96(11):6211-6.
 Perez-Villar JJ, O'Day K, Hewgill DH, Nadler SG, Kanner SB. Nuclear localization of the tyrosine kinase Itk and interaction of its SH3 domain with karyopherin alpha (Rch1alpha). Int Immunol. 2001;13(10):1265-74.
 Zhou D, Chen S. PNRC2 is a 16 kDa coactivator that interacts with nuclear receptors through an SH3-binding motif. Nucleic Acids Res. 2001;29(19):3939-48.
 Jiang M, Axe T, Holgate R, Rubbi CP, Okorokov AL, Mee T, Milner J. p53 binds the nuclear matrix in normal cells: binding involves the proline-rich domain of p53 and increases following genotoxic stress. Oncogene. 2001;20(39):5449-58.
 Grishin AV, Azhipa O, Semenov I, Corey SJ. Interaction between growth arrest-DNA damage protein 34 and Src kinase Lyn negatively regulates genotoxic apoptosis. Proc Natl Acad Sci U S A. 2001;98(18):10172-7.
 Dong Chen, Waters SB, Holt KH, Pessin JE. SOS phosphorylation and disassociation of the Grb2-SOS complex by the ERK and JNK signaling pathways. J Biol Chem. 1996;271(11):6328-32.
 Gout I, Middleton G, Adu J, Ninkina NN, Drobot LB, Filonenko V, Matsuka G, Davies AM, Waterfield M, Buchman VL. Negative regulation of PI 3-kinase by Ruk, a novel adaptor protein. EMBO J. 2000;19(15):4015-25.
 Take H, Watanabe S, Takeda K, Yu ZX, Iwata N, Kajigaya S. Cloning and characterization of a novel adaptor protein, CIN85, that interacts with c-Cbl. Biochem Biophys Res Commun. 2000;268(2):321-8.
 Borinstein SC, Hyatt MA, Sykes VW, Straub RE, Lipkowitz S, Boulter J, Bogler O. SETA is a multifunctional adapter protein with three SH3 domains that binds Grb2, Cbl, and the novel SB1 proteins. Cell Signal. 2000;12(11-12):769-79.
 Peterson GL. A simplification of the protein assay method of Lowry et al. which is more generally applicable. Anal Biochem. 1977;83(2):346-56.
 Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970;227(5259):680-5.
 Watanabe S, Take H, Takeda K, Yu ZX, Iwata N, Kajigaya S. Characterization of the CIN85 adaptor protein and identification of components involved in CIN85 complexes. Biochem Biophys Res Commun. 2000;278(1):167-74.
 Kzhyshkowska J, Sch?tt H, Liss M, Kremmer E, Stauber R, Wolf H, Dobner T. Heterogeneous nuclear ribonucleoprotein E1B-AP5 is methylated in its Arg-Gly-Gly (RGG) box and interacts with human arginine methyltransferase HRMT1L1. Biochem J. 2001;358(Pt 2):305-14.
 Craggs G, Finan PM, Lawson D, Wingfield J, Perera T, Gadher S, Totty NF, Kellie S. A nuclear SH3 domain-binding protein that colocalizes with mRNA splicing factors and intermediate filament-containing perinuclear networks. J Biol Chem. 2001;276(32):30552-60.
 Jahn T, Seipel P, Coutinho S, Miething C, Peschel C, Duyster J. Grb4/Nckbeta acts as a nuclear repressor of v-Abl-induced transcription from c-jun/c-fos promoter elements. J Biol Chem. 2001;276(46):43419-27.