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Protein Intrinsic Disorder, Cell Signaling and Alternative Splicing

Protein Intrinsic Disorder, Cell Signaling and Alternative Splicing. Outline of Talk. Examples of intrinsically disordered proteins Prediction of natural disordered regions Disorder and cell signaling Disorder and molecular recognition Disorder and alternative splicing

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Protein Intrinsic Disorder, Cell Signaling and Alternative Splicing

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  1. Protein Intrinsic Disorder, Cell Signaling and Alternative Splicing

  2. Outline of Talk • Examples of intrinsically disordered proteins • Prediction of natural disordered regions • Disorder and cell signaling • Disorder and molecular recognition • Disorder and alternative splicing • Protein isoforms and functional diversity via the linkage of alternative splicing and intrinsic disorder

  3. Molecular Recognition Element (MoRE) CDK Cyclin A p27kip1 3D structure from: Russo A et al., Nature 382:325-331 (1996)

  4. Disorder and Function Dunker AK et al., Adv Protein Chem 62: 25-49 (2002)

  5. Disordered Sequence Data Attribute Selection or Extraction Separate Training and Testing Sets Predictor Training Predictor Validation on Out-of-Sample Data Prediction Prediction of Disorder

  6. www.disprot.org

  7. p53 MoREs PONDR® VL-XT Score Oldfield et al., Biochemistry 44: 12454-12470 (2005)

  8. Protein Interaction Domains http://www.mshri.on.ca/pawson/domains.html

  9. GYF Domain and CD2 Chain B Freund et al., (2002) Embo J. 21:5985-5995

  10. GYF Domain of CD2 Binding Protein Freund et al., (1999) Nat. Struct. Biol. 6:656-660

  11. CD2: Binding Partner of GYF Domain Consensus sequence (GYF binding sites) has the sequence: ppppghr. The peptide in the crystal structure has the aa sequence: shrppppghrv. Freund et al., (1999) Nat. Struct. Biol. 6:656-660

  12. Analysis of Signaling Interactions • Examined each interaction on Pawson’s website. • Almost all of the interactions involved ordered regions binding to disordered partners. • Conclusion: if Pawson’s examples are typical, then a very significant proportion of protein-protein signaling interactions use disordered regions.

  13. Parallel Paradigms Catalysis AA seq →3-D Structure→ Function Signaling AA seq →Disordered→Function Ensemble

  14. Alternative Splicing and Intrinsic Disorder • Find proteins with both ordered and disordered regions. • Find mRNA alternative splicing information for these proteins and map to the ordered and disordered regions. • For alternatively spliced regions of mRNA, do they code for ordered protein more often or do they code for disordered protein more often?

  15. Alternative Splicing 5’ UTR 3’ UTR Coding Sequence

  16. Alternative Splicing 5’ UTR 3’ UTR Coding Sequence mRNA Transcription Translation Protein sequence

  17. Alternative Splicing 5’ UTR 3’ UTR Coding Sequence mRNA 2 mRNA 1 Transcription Translation Isoform 1 Isoform 2

  18. Alternative Splicing 5’ UTR 3’ UTR Coding Sequence mRNA 2 mRNA 1 Transcription Translation Isoform 1 Isoform 2 AS region Folding

  19. Disordered AS regions Structural Studies of AS Structured AS regions Pyrophosphorylase RAC1 Tumor necrosis factor Sulphotransferase Glutathione S-transferase

  20. ASED dataset: 46 proteins 74 characterized AS regions >19,000 charaterized residues, 35% ID Studying the Relationship IDAS ASG (AS Gallery) DisProt  SwissProt (VarSplic) Database of proteins with experimentally determinedstructure and disorder www.disprot.org

  21. Results on ASED Distribution of structurally characterized AS regions

  22. Enlarging the Dataset ASED dataset PONDR® VSL1 ID predictor (> 80% accuracy) Validation ASSP dataset 558AS human proteins fromSwissProt 1,266AS regions Analysis

  23. Global Results AS regions disorder distributions in ASED and ASSP

  24. Alternative Splicing and Disorder • Ordered Proteins: active site residues non-local in sequence, become associated by protein folding • Disordered Proteins and regions: functional residues localized in squence • Functional regions for signaling and regulation are located one after another • Alternative splicing edits functional sets and thereby leads to regulatory and signaling diversity

  25. Breast Cancer Protein 1 (BRCA1)

  26. Summary • Protein signaling interactions involve intrinsic disorder (ID) a high percentage of the time. • Alternative splicing (AS) often occurs in regions of pre-mRNA that code for intrinsic disorder. • AS + ID facilitate regulatory and signaling diversity. • Is AS + ID the critical combination for the evolution of multi-cellular organisms?

  27. Acknowledgements Temple University Zoran Obradovic Slobodan Vucetic Vladimir Vacic Kang Peng Rockefeller University Lilia Iakoucheva Sebat University of Wisconsin John Markley Chris Oldfield UCSF Ethan Garner PNNL Richard Smith Eric Ackerman Indiana University Predrag Radivojac Pedro Romero Marc Cortese Gerard Go Amrita Mohan Jie Sun Siama Zaida Jack Yang University of Idaho Celeste J. Brown Chris Williams Molecular Kinetics Vladimir Uversky Yugong Cheng

  28. Support • NSF CSE II 9711532 • NIH R01 LM007688 • USDA 2000 1740 • INGEN®, Lilly Endowment • Molecular Kinetics

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