Protein Intrinsic Disorder Cell Signaling and Alternative Splicing

Protein Intrinsic Disorder, Cell Signaling and Alternative Splicing Center For Computational Biology and Bioinformatics

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

Molecular Recognition Element (Mo. RE) CDK Cyclin A 3 D structure from: Russo A et al. , Nature 382: 325 -331 (1996) p 27 kip 1

Disorder and Function Category Change Examples Descriptions Molecular Recognition D O 113 Inter- and Intra-protein, ss. DNA, ds. DNA, t. RNA, r. RNA, m. RNA, n. RNA, bilayers, ligands, co-factors, metals Protein Modification Variable 36 Acetylation, fatty acylation, glycosylation, methylation, phosphorylation, ADPribosylation, ubiquitination, proteolytic digestion Entropic Chains Variable 17 Linkers, spacers, bristles, clocks, springs, detergents, self-transport Dunker AK et al. , Adv Protein Chem 62: 25 -49 (2002)

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

www. disprot. org

PONDR® VL-XT Score p 53 Mo. REs Oldfield et al. , Biochemistry 44: 12454 -12470 (2005)

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

GYF Domain and CD 2 Chain B Freund et al. , (2002) Embo J. 21: 5985 -5995

GYF Domain of CD 2 Binding Protein Freund et al. , (1999) Nat. Struct. Biol. 6: 656 -660

CD 2: 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

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.

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

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

Alternative Splicing 5’ UTR Coding Sequence 3’ UTR

Alternative Splicing 5’ UTR m. RNA Protein sequence Coding Sequence Transcription Translation 3’ UTR

Alternative Splicing 5’ UTR m. RNA 1 Isoform 1 Coding Sequence Transcription Translation 3’ UTR m. RNA 2 Isoform 2

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

Structural Studies of AS Disordered AS regions Pyrophosphorylase Structured AS regions RAC 1 Tumor necrosis factor Glutathione S-transferase Sulphotransferase

Studying the Relationship ID AS Dis. Prot Database of proteins with experimentally determined structure and disorder www. disprot. org ASG (AS Gallery) Swiss. Prot (Var. Splic) ASED dataset: 46 proteins 74 characterized AS regions >19, 000 charaterized residues, 35% ID

Results on ASED Distribution of structurally characterized AS regions

Enlarging the Dataset PONDR® VSL 1 ID predictor (> 80% accuracy) Validation Analysis ASED dataset ASSP dataset 558 AS human proteins from Swiss. Prot 1, 266 AS regions

Global Results AS regions disorder distributions in ASED and ASSP

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

Breast Cancer Protein 1 (BRCA 1)

Summary • Protein signaling interactions involve intrinsic disorder (ID) a high percentage of the time. • Alternative splicing (AS) often occurs in regions of pre-m. RNA 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?

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

Support • • • NSF CSE II 9711532 NIH R 01 LM 007688 USDA 2000 1740 INGEN®, Lilly Endowment Molecular Kinetics