BRCT Repeats As Phosphopeptide-Binding Modules Involved in Protein Targeting

We used a proteomic approach to identify phosphopeptide-binding modules mediating signal transduction events in the DNA damage response pathway. Using a library of partially degenerate phosphopeptides, we identified tandem BRCT (BRCA1 carboxyl-terminal) domains in PTIP (Pax transactivation domain-interacting protein) and in BRCA1 as phosphoserine- or phosphothreonine-specific binding modules that recognize substrates phosphorylated by the kinases ATM (ataxia telangiectasia–mutated) and ATR(ataxia telangiectasia– and RAD3-related) in response to γ-irradiation. PTIP tandem BRCT domains are responsible for phosphorylation-dependent protein localization into 53BP1- and phospho-H2AX (γ-H2AX)–containing nuclear foci, a marker of DNA damage. These findings provide a molecular basis for BRCT domain function in the DNA damage response and may help to explain why the BRCA1 BRCT domain mutation Met1775 → Arg, which fails to bind phosphopeptides, predisposes women to breast and ovarian cancer.

[1]  T. Halazonetis,et al.  P53 Binding Protein 1 (53bp1) Is an Early Participant in the Cellular Response to DNA Double-Strand Breaks , 2000, The Journal of cell biology.

[2]  D. Chan,et al.  Utilization of Oriented Peptide Libraries to Identify Substrate Motifs Selected by ATM* , 2000, The Journal of Biological Chemistry.

[3]  M. Yaffe,et al.  Mapping specificity determinants for protein-protein association using protein fusions and random peptide libraries. , 2000, Methods in enzymology.

[4]  M. Sternberg,et al.  The BRCA1 C-terminal domain: structure and function. , 2000, Mutation research.

[5]  R. Abraham.  Cell cycle checkpoint signaling through the ATM and ATR kinases. , 2001, Genes & development.

[6]  J. Glover,et al.  Crystal structure of the BRCT repeat region from the breast cancer-associated protein BRCA1 , 2001, Nature Structural Biology.

[7]  J. Sarkaria,et al.  Inhibition of ATM and ATR kinase activities by the radiosensitizing agent, caffeine. , 1999, Cancer research.

[8]  Michael B Yaffe,et al.  Proteomic Screen Finds pSer/pThr-Binding Domain Localizing Plk1 to Mitotic Substrates , 2003, Science.

[9]  D. Livingston,et al.  In search of the tumour-suppressor functions of BRCA1 and BRCA2 , 2000, Nature.

[10]  Junjie Chen,et al.  Tumor Suppressor P53 Binding Protein 1 (53bp1) Is Involved in DNA Damage–Signaling Pathways , 2001, The Journal of cell biology.

[11]  B. Price,et al.  Caffeine inhibits the checkpoint kinase ATM , 1999, Current Biology.

[12]  M. Yaffe,et al.  Phosphoserine/threonine-binding domains. , 2001, Current opinion in cell biology.

[13]  J. Glover,et al.  Structural Consequences of a Cancer-causing BRCA1-BRCT Missense Mutation* , 2003, The Journal of Biological Chemistry.

[14]  T. Pawson,et al.  Assembly of Cell Regulatory Systems Through Protein Interaction Domains , 2003, Science.

[15]  S. Elledge,et al.  53BP1, a Mediator of the DNA Damage Checkpoint , 2002, Science.

[16]  J. Gurdon,et al.  Swift Is a Novel BRCT Domain Coactivator of Smad2 in Transforming Growth Factor β Signaling , 2001, Molecular and Cellular Biology.

[17]  M. Yaffe.  Phosphotyrosine-binding domains in signal transduction , 2002, Nature Reviews Molecular Cell Biology.

[18]  Robert D. Finn,et al.  Pfam 3.1: 1313 multiple alignments and profile HMMs match the majority of proteins , 1999, Nucleic Acids Res..

[19]  G. Dressler,et al.  PTIP, a novel BRCT domain-containing protein interacts with Pax2 and is associated with active chromatin. , 2000, Nucleic acids research.

[20]  X. Le,et al.  Measuring DNA damage using capillary electrophoresis with laser-induced fluorescence detection. , 2000, Methods.

[21]  S. Elledge,et al.  The DNA damage response: putting checkpoints in perspective , 2000, Nature.

[22]  Marc J. Prindle,et al.  BRCT Domain-Containing Protein PTIP Is Essential for Progression through Mitosis , 2003, Molecular and Cellular Biology.

[23]  S. T. Kim,et al.  Substrate Specificities and Identification of Putative Substrates of ATM Kinase Family Members* , 1999, The Journal of Biological Chemistry.