Interrogation of cross talk between proteins and gene regulatory networks in breast cancer Chambers, Teressa Lee Hiren Karathia Sridhar Hannenhalli

Cross Talk of Protein and Gene Regulation?? HPRD HPRD Ligand INTERACTION Receptor INTERACTION PPI Interactions PTM I A P NETPATH S E E Interact with other Protein molecules Prot ANNOTATION Transport of Pn to Nucleus from Cytoplasm Prot Getting information of pathway events from literatures Activation of TF TF Regulation of Genes

View of EGFR Protein Signaling Pathways

View of EGFR Protein Signaling Pathways

View of EGFR Protein Signaling Pathways

NetPath NetPath is a curated resource of signal transduction pathways in humans. At present there are 20 pathways 10 – Immune signaling pathways 10 – Cancer Signaling pathways

TCGA data set 100 Normal 100 Benign Cancer

Protein Protein Interactions Proteins Proteins Protein Protein Interactions Interfacing TF (ITF) ?? TF Protein DNA Interactions DNA (Genes)

Questions How can we identify context specific ITFs? What are correlation between ITFs centric protein and gene regulatory networks? Which ITF centric paths and target genes are cancer specific?

Collected from Databases and Literatures Pipeline Protein paths (K = 2) 200 Samples Parse XML (SBML) STAT RNA-Seq Collected from Databases and Literatures Target Genes Proteins Target Genes Corr STAT

Comparisons

Comparisons Protein paths (K = 3) STAT STAT Target Genes

All Protein paths (K <= 3) STAT ** Target Genes

** Protein paths (K = 1) STAT ** Target Genes

Protein paths (K = 2) STAT ** Target Genes

** Protein paths (K = 1) ** Target Genes

** Protein paths (K = 2) Target Genes **

Validation of the analyses

Validating a candidate gene

Flow of whole experiment

THANK YOU

Molecular Cross Talk

RELATIONSHIPS OF ALL THE RESOURCES Protein or gene centric functional information i.e., HPRD PUBMED Interaction information Annotation Gene regulation database Gene locus NetPath (Information of signaling pathways) SNPs Pathway Details Biological scientists read full text papers from pubmed database GenProt (Genomic information with SNPs) Genomic Details VALIDATION OF PATHWAY CANDIDATES dbSNP

View of EGFR induced Signaling pathways

RELATIONSHIPS OF ALL THE RESOURCES Protein centric functional annotations i.e., HPRD PUBMED Interaction information Manual annotation Gene regulation database Gene locus NetPath (Information of signaling pathways) SNPs Pathway Details Biological scientists read full text research papers from pubmed database. GenProt (Genomic information with SNPs) Genomic Details VALIDATION OF PATHWAY CANDIDATES dbSNP

Biological databases HPRD (Human Protein Reference Database) PPD (Plasma Protein Database) GenProt NetPATH (Networks and PATHways) Proteinpedia

HPRD (Human Protein Reference Database) The HPRD is a protein information resource that provides extensive information pertaining to human proteins. It includes information as follows: Domain Architecture Protein Function Protein-protein Interaction Post Translation Modification (PTMs) Enzyme-substrate relationships Sub cellular localization Tissue Expression Disease association All this Information collected by Molecular Biologists from full text papers available in Pubmed Database

ARCHITECTURE OF HPRD

ARCHITECTURE OF HPRD

FRONT PAGE OF HPRD @ http://hprd.org

Queries in HPRD

RESULT PAGE OF QUERY

PAGE OF INDIVIDUAL PROTEIN ENTRY

Protein Sequence Information

Protein Interaction Information

Protein Post Translation Modification & Substrate information

GENPROT Genprot is graphically viewer, which depicts human genomic sequence alongside its transcript and SNP (Single Nucleotide Polymorphism) if available on protein coding or non coding DNA sequences.

Genprot main interface

Integration of HPRD in GENPROT

PATH FOR KNOWLEDGE DISCOVERY Data (Biological Data) Information Knowledge OLAP OLTP Obtained Raw DATA From DATABASE (Biological Database) Storing Information in DATABASE (Biological Database) Storing Knowledge in Knowledge Base (Biological Data Warehouse) Data Manipulation Data Mining

DATA MINING APPROACH ON HPRD Biological Data (Interpreted by Biologist) Biologial meaningful Information Knowledge discovered (Data mining process) OLAP OLTP Obtained Raw DATA From DATABASE (Pubmed) Storing Information in DATABASE (HPRD) Storing Knowledge in Knowledge Base (Biological Data Warehouse) Data Interpretation Data Mining

HPRD report on Chromosome dimension

HPRD Report on Disease dimension

HPRD report based on Localization dimension

PTM event distribution

HPRD report on Protein and cDNA length

HPRD report based on Protein expression in Tissues

HPRD report based on Protein expression in Tissues

RELATIONSHIPS OF ALL THE RESOURCES HPRD or PPD PUBMED Interaction information Annotation Gene regulation database Gene locus NetPath (Information of signaling pathways) SNPs Pathway Details Biological scientists read full text papers from pubmed database GenProt (Genomic information with SNPs) Genomic Details VALIDATION OF PATHWAY CANDIDATES dbSNP

POSSIBLE APPLICATIONS OF HPRD IN MOLECULAR BIOLOGY 1. Comparative interactome analysis between human and other species by using data mining and wet laboratory approach 2. Design the strategy for validating the protein-protein interactions.

Example of comparative interactome analysis 25,464 24,587 105 Human Worm ORTHOLOGS ANALYSIS 42 21 PPI out of 36 found to be with high confidence 36 63 Fly 5,625 Human: 25,464 PPI (HPRD) 9 out of these 21 PPI was taken for validation done by Coimmunoprecipitation experiment C.elegans: 5,625 PPI (Li et al., 2004) CO-IMMUNOPRECIPITATION (CO-IP) EXPERIMENT S.cerevisiae: 15,675 PPI (MIPS & DIP) D.melanogaster: 20,439 PPI (Glot et al., 2003)

Objectives for validating biomolecules Specific objective - 1: Analysis of protein complexes of adapter proteins that are not well characterized in EGFR1 pathway. Experiment techniques: Protein digestion & Mass Spectroscopy Specific objective - 2: Validating the interactions/interactors of EGFR1 pathway molecules identified through large-scale yeast 2-hybrid (Y2H) screens through biochemical and functional assays. Experiment techniques: Co-IP and Western Blotting

Human ProteinPedia Human Proteinpedia is a community portal for sharing and integration of human protein data. Human Proteinpedia is information source of Protein-Protein interactions Tissue expression Expression in Cell line Subcellular Localization Enzyme Substrate relationships All together are obtained from number of experiment evidences. For example 1. Co-immunoprecipitation. 6. Western Blotting. 2. Mass spectroscopy. 7. Yeast 2 Hybrid experiments. 3. Fluorescence based experiments 4. Immunohistochemistry 5. Protein peptide Microarray

Human Proteinpedia

QUERING IN HUMAN PROTEINPEDIA

Results of Query

Results

Mass spectroscopy results

Mass spectroscopy peaks

CONCLUSION HPRD is a knowledgebase of Human Proteins that can assist the biomedical community in discovering genomic, transcriptomic and proteomic information together in an integrated view in terms of sequence, function and protein interactions, which provides facilities to a scientist to differentiate between ordered and disordered states of human cells.

CAREER PATH EDUCATION B.Sc (Biochemistry) M.Sc (Biomedical Technology) M.Sc (Bioinformatics) Joined as Research Scientist at Institute of Bioinformatics Worked for developing and designing HPRD, NetPath & Genprot.

Dept. of Biomathematics and Bioinformatics Under the supervision of HPRD development team Joined as Ph.D student funded by AGAUR in Dept. of Biomathematics and Bioinformatics at UDL Under the supervision of Dr. Rui Alves

“MOLTES GRACIES”