Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Paper
  • Published:

Role of the Brk SH3 domain in substrate recognition

Oncogene volume 23, pages 2216–2223 (2004)Cite this article

Abstract

Breast tumor kinase (Brk) is a nonreceptor tyrosine kinase that is overexpressed in a high percentage of breast carcinomas. Brk contains SH3, SH2, and tyrosine kinase catalytic domains in a similar arrangement as Src family kinases. In this study, we explored the roles of the SH3 and SH2 domains in Brk regulation and substrate binding. We introduced a series of mutations into Brk that were predicted to disrupt the intramolecular interactions involving the SH3 and SH2 domains. These mutant forms of Brk displayed higher activity than wild-type Brk when expressed in human embryonic kidney HEK293 cells. These studies also allowed us to pinpoint the intramolecular binding site for the SH3 domain. To examine substrate binding, we compared binding and phosphorylation of Sam68, a physiological substrate of Brk. These experiments showed that the SH3 domain plays a particularly important role in substrate recognition. We confirmed this conclusion using a series of synthetic peptides in which a substrate sequence was coupled to an SH3 or SH2 ligand. The SH3-binding substrate had a significantly lower Km than a control, while no difference was observed between an SH2-binding substrate and a control. Taken together, our data suggest that SH3 interactions will govern phosphorylation of many substrates by Brk.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  • Alexandropoulos K and Baltimore D . (1996). Genes Dev., 10, 1341–1355.

  • Barker KT, Jackson LE and Crompton MR . (1997). Oncogene, 15, 799–805.

  • Barker SC, Kassel DB, Weigl D, Huang X, Luther MA and Knight WB . (1995). Biochemistry, 34, 14843–14851.

  • Briggs SD, Sharkey M, Stevenson M and Smithgall TE . (1997). J. Biol. Chem., 272, 17899–17902.

  • Brown MT and Cooper JA . (1996). Biochim. Biophys. Acta, 1287, 121–149.

  • Coyle JH, Guzik BW, Bor YC, Jin L, Eisner-Smerage L, Taylor SJ, Rekosh D and Hammarskjold ML . (2003). Mol. Cell. Biol., 23, 92–103.

  • Derry JJ, Prins GS, Ray V and Tyner AL . (2003). Oncogene, 22, 4212–4220.

  • Derry JJ, Richard S, Valderrama Carvajal H, Ye X, Vasioukhin V, Cochrane AW, Chen T and Tyner AL . (2000). Mol. Cell. Biol., 20, 6114–6126.

  • Easty DJ, Mitchell PJ, Patel K, Florenes VA, Spritz RA and Bennett DC . (1997). Int. J. Cancer, 71, 1061–1065.

  • Erpel T, Superti-Furga G and Courtneidge SA . (1995). EMBO J., 14, 963–975.

  • Fumagalli S, Totty NF, Hsuan JJ and Courtneidge SA . (1994). Nature, 368, 871–874.

  • Hantschel O, Nagar B, Guettler S, Kretzschmar J, Dorey K, Kuriyan J and Superti-Furga G . (2003). Cell, 112, 845–857.

  • Harvey AJ and Crompton MR . (2003). Oncogene, 22, 5006–5010.

  • Jove R and Hanafusa H . (1987). Annu. Rev. Cell Biol., 3, 31–56.

  • Kamalati T, Jolin HE, Fry MJ and Crompton MR . (2000). Oncogene, 19, 5471–5476.

  • Kamalati T, Jolin HE, Mitchell PJ, Barker KT, Jackson LE, Dean CJ, Page MJ, Gusterson BA and Crompton MR . (1996). J. Biol. Chem., 271, 30956–30963.

  • LaFevre-Bernt M, Sicheri F, Pico A, Porter M, Kuriyan J and Miller WT . (1998). J. Biol. Chem., 273, 32129–32134.

  • Liu X, Brodeur SR, Gish G, Songyang Z, Cantley LC, Laudano AP and Pawson T . (1993). Oncogene, 8, 1119–1126.

  • Llor X, Serfas MS, Bie W, Vasioukhin V, Polonskaia M, Derry J, Abbott CM and Tyner AL . (1999). Clin. Cancer Res., 5, 1767–1777.

  • Mayer BJ and Baltimore D . (1994). Mol. Cell. Biol., 14, 2883–2894.

  • Mayer BJ, Hirai H and Sakai R . (1995). Curr. Biol., 5, 296–305.

  • McPherson RA, Taylor MM, Hershey ED and Sturgill TW . (2000). Oncogene, 19, 3616–3622.

  • Miller WT . (2003). Acc. Chem. Res., 36, 393–400.

  • Mitchell PJ, Barker KT, Martindale JE, Kamalati T, Lowe PN, Page MJ, Gusterson BA and Crompton MR . (1994). Oncogene, 9, 2383–2390.

  • Mitchell PJ, Sara EA and Crompton MR . (2000). Oncogene, 19, 4273–4282.

  • Moarefi I, LaFevre-Bernt M, Sicheri F, Huse M, Lee CH, Kuriyan J and Miller WT . (1997). Nature, 385, 650–653.

  • Nagar B, Hantschel O, Young MA, Scheffzek K, Veach D, Bornmann W, Clarkson B, Superti-Furga G and Kuriyan J . (2003). Cell, 112, 859–871.

  • Nguyen JT and Lim WA . (1997). Nat. Struct. Biol., 4, 256–260.

  • Pawson T, Gish GD and Nash P . (2001). Trends Cell Biol., 11, 504–511.

  • Pellicena P, Stowell KR and Miller WT . (1998). J. Biol. Chem., 273, 15325–15328.

  • Qiu H and Miller WT . (2002). J. Biol. Chem., 277, 34634–34641.

  • Scott MP and Miller WT . (2000). Biochemistry, 39, 14531–14537.

  • Serfas MS and Tyner AL . (2003). Oncol. Res., 13, 409–419.

  • Sicheri F and Kuriyan J . (1997). Curr. Opin. Struct. Biol., 7, 777–785.

  • Sicheri F, Moarefi I and Kuriyan J . (1997). Nature, 385, 602–609.

  • Songyang Z, Carraway KL, III Eck MJ, Harrison SC, Feldman RA, Mohammadi M, Schlessinger J, Hubbard SR, Smith DP, Eng C, Lorenzo MJ, Poner BAJ, Mayer BJ and Cantley LC . (1995). Nature, 373, 536–539.

  • Taylor SJ and Shalloway D . (1994). Nature, 368, 867–871.

  • Tian M and Martin GS . (1997). Mol. Biol. Cell, 8, 1183–1193.

  • Vasioukhin V, Serfas MS, Siyanova EY, Polonskaia M, Costigan VJ, Liu B, Thomason A and Tyner AL . (1995). Oncogene, 10, 349–357.

  • Wang D, Huang XY and Cole PA . (2001). Biochemistry, 40, 2004–2010.

  • Xu W, Harrison SC and Eck MJ . (1997). Nature, 385, 595–602.

  • Young MA, Gonfloni S, Superti-Furga G, Roux B and Kuriyan J . (2001). Cell, 105, 115–126.

Download references

Acknowledgements

This work was supported by a grant from the NIH to WTM (CA28146).

Author information

Authors and Affiliations

  1. Department of Physiology and Biophysics, Basic Science Tower, T-6, School of Medicine, State University of New York at Stony Brook, Stony Brook, 11794-8661, NY, USA

    Haoqun Qiu & W Todd Miller

Authors
  1. Haoqun Qiu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. W Todd Miller
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to W Todd Miller.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Qiu, H., Miller, W. Role of the Brk SH3 domain in substrate recognition. Oncogene 23, 2216–2223 (2004). https://doi.org/10.1038/sj.onc.1207339

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.onc.1207339

Keywords

This article is cited by

Search

Advanced search

Quick links