GENOME DUPLICATION AND GENE ANNOTATION: AN EXAMPLE FOR A REFERENCE PLANT SPECIES.
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1 GENOME DUPLICATION AND GENE ANNOTATION: AN EXAMPLE FOR A REFERENCE PLANT SPECIES. Alessandra Vigilante, Mara Sangiovanni, Chiara Colantuono, Luigi Frusciante and Maria Luisa Chiusano Dept. of Soil, Plant, Environmental and Animal Production Sciences CAB (Computer Aided Biosciences) group
2 BACKGROUND: Arabidopsis thaliana as a reference genome Arabidopsis thaliana WAS THE FIRST PLANT GENOME TO BE COMPLETELY SEQUENCED
3 BACKGROUND: Arabidopsis thaliana as a reference genome A REFERENCE GENOME SHOULD BE: FULLY RELIABLE SAFELY ANNOTATED WELL UNDERSTOOD IN TERMS OF EVOLUTIONARY HISTORY
4 BACKGROUND: Arabidopsis thaliana as a reference genome A REFERENCE GENOME SHOULD BE: FULLY RELIABLE SAFELY ANNOTATED WELL UNDERSTOOD IN TERMS OF EVOLUTIONARY HISTORY Arabidopsis thaliana GENOME IS: GENE DENSE COMPLEX BECAUSE HIGHLY DUPLICATED AND CLAIMED TO BE ARCHEOPOLYPLOID STILL NOT EXHAUSTIVELY ANNOTATED
5 BACKGROUND: Arabidopsis thaliana as a reference genome Whole genome duplication events Nature (2007) Jaillon O, et al. The grapevine genome sequence suggests ancestral hexaploidization in major angiosperm phyla.
6 BACKGROUND: Arabidopsis thaliana as a reference genome V. vinifera A. thaliana Whole genome duplication events Nature (2007) Jaillon O, et al. The grapevine genome sequence suggests ancestral hexaploidization in major angiosperm phyla.
7 STRATEGY: Unraveling Arabidopsis thaliana genome IT COULD BE USEFUL TO REVIEW THE GENOME IN TERMS OF RELATIONSHIPS BETWEEN DUPLICATED GENES/PARALOGS
8 STRATEGY: Unraveling Arabidopsis thaliana genome IT COULD BE USEFUL TO REVIEW THE GENOME IN TERMS OF RELATIONSHIPS BETWEEN DUPLICATED GENES/PARALOGS NETWORKS OF PARALOG GENES
9 Gene Gene duplication analysis: pipeline Arabidopsis thaliana proteome (TAIR9 release) All-against-all BLASTp versus protein-coding genes E<10-10, Rost s formula Singleton genes Duplicated genes Network extraction Networks of duplicated genes
10 Gene Gene duplication analysis: pipeline Arabidopsis thaliana proteome (TAIR9 release) All-against-all BLASTp versus protein-coding genes E<10-10, Rost s formula Singleton genes Duplicated genes Network extraction Networks of duplicated genes
11 Gene Gene duplication analysis: pipeline Arabidopsis thaliana proteome (TAIR9 release) All-against-all BLASTp versus protein-coding genes E<10-10, Rost s formula Singleton genes Duplicated genes Network extraction Networks of duplicated genes
12 Gene Gene duplication analysis: pipeline Arabidopsis thaliana proteome (TAIR9 release) All-against-all BLASTp versus protein-coding genes E<10-10, Rost s formula Singleton genes Duplicated genes Network extraction Networks of duplicated genes
13 Gene Gene duplication analysis: pipeline A. thaliana GENES AND PARALOGIES ARE REPRESENTED AS AN (UNDIRECTED) GRAPH G(V,E) WHERE: - V ={v 1,..v N } = genes - E ={e 1,..e M } = paralogies
14 Gene Gene duplication analysis: pipeline A. thaliana GENES AND PARALOGIES ARE REPRESENTED AS AN (UNDIRECTED) GRAPH G(V,E) WHERE: - V ={v 1,..v N } = genes - E ={e 1,..e M } = paralogies
15 RESULTS: Gene duplication Gene duplication analysis: analysis pipeline Arabidopsis thaliana proteome (TAIR9 release) All-against-all BLASTp versus protein-coding genes E<10-10, Rost s formula Singleton genes Duplicated genes Networks of duplicated genes
16 RESULTS: Gene duplication Gene duplication analysis: analysis pipeline Arabidopsis thaliana proteome (TAIR9 release) All-against-all BLASTp versus protein-coding genes E<10-10, Rost s formula Singleton genes 5326 Duplicated genes Networks of duplicated genes
17 RESULTS: Gene duplication Gene duplication analysis: analysis pipeline Genes A network contains all and only the genes that share at least one paralogy relationship. Each gene belongs to one and only one network.
18 RESULTS: Gene duplication Networks analysis: applications pipeline NETWORKS ARE A USEFUL TOOL TO DEEPLY INVESTIGATE RELATIONSHIPS BETWEEN SUBSETS OF GENES SHARING DUPLICATION RELATIONSHIPS NETWORKS CAN BE A USEFUL TOOL TO REFINE GENE ANNOTATION
19 RESULTS: Networks Gene duplication Networks applications applications analysis: for the pipeline annotation of unknown information The 19% of the proteome is still annotated as unknown protein NETWORKS CAN HELP IN REFINING GENE ANNOTATION
20 RESULTS: Networks Gene duplication applications Networks applications analysis: for study pipeline of relationships between gene familes
21 RESULTS: Networks Gene duplication applications Networks applications analysis: for study pipeline of relationships between gene familes Networks can be a useful tool for highlighting evolutionary relationships between different gene families
22 RESULTS: Gene Networks duplication of applications analysis: duplicated pipeline genes
23 RESULTS: Gene Networks duplication of applications analysis: duplicated pipeline genes
24 RESULTS: Gene Networks duplication of applications analysis: duplicated pipeline genes
25 Gene RESULTS: duplication Networks 2-gene applications analysis: networks pipeline
26 Gene RESULTS: duplication Networks 2-gene applications analysis: networks pipeline Arabidopsis thaliana chromosomes
27 Gene RESULTS: duplication Networks 2-gene applications analysis: networks pipeline All protein-coding genes
28 Gene RESULTS: duplication Networks 2-gene applications analysis: networks pipeline All genes involved in two-gene networks
29 RESULTS: Gene 2-gene duplication Networks networks 2-gene networks applications analysis: and singleton pipeline genes AT LEAST 5% OF THE ENTIRE PROTEOME HAS A SINGLE PARALOGY RELATIONSHIP AT LEAST 20% OF THE ENTIRE PROTEOME IS A SINGLETON ABOUT ONE QUARTER OF THE ENTIRE PROTEOME HAS ZERO OR AT MOST ONE PARALOGY RELATIONSHIP
30 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline WHAT ABOUT SINGLETON GENES?
31 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline Singleton genes not having protein-coding paralogs Blastn mrna sequences against ESTs Singleton genes validated or not validated by ESTs 3588
32 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline Singleton genes not having protein-coding paralogs 3588 Comparative analysis (Biomart Ensembl) Singleton s orthologs in different species (O.sativa, V.vinifera, S.bicolor, P.trichocarpa)
33 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline SINGLETON GENES ANALYSIS RESULTS 3588 singleton genes
34 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline SINGLETON GENES ANALYSIS RESULTS 3588 singleton genes 2516 confirmed by ESTs 1072 Not confirmed by ESTs
35 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline SINGLETON GENES ANALYSIS RESULTS 3588 singleton genes 2516 confirmed by ESTs 1072 Not confirmed by ESTs 1072 Present only in A. thaliana
36 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline SINGLETON GENES ANALYSIS RESULTS 3588 singleton genes 2516 confirmed by ESTs 1072 Not confirmed by ESTs 481 Present only in A. thaliana 1072 Present only in A. thaliana
37 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline SINGLETON GENES ANALYSIS RESULTS 2516 confirmed by ESTs
38 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline SINGLETON GENES ANALYSIS RESULTS 481 out of 2516 present only in A.thaliana
39 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline SINGLETON GENES ANALYSIS RESULTS 1110 Not confirmed by ESTs and present only in A.thaliana
40 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline Singleton genes not having protein-coding paralogs 3588 BLASTn vs: Pseudogenes Transposons Other RNA Intergenic regions Singleton genes not having protein-coding/ non protein-coding paralogs and any significant hit in the genome 250
41 RESULTS: Gene duplication Networks Singleton 2-gene networks applications analysis: genes analysis pipeline Singleton genes not having protein-coding paralogs 3588 Blastx protein sequences against transcript Searching for ORF annotation errors MT1A (METALLOTHIONEIN 1A) MT1B (METALLOTHIONEIN 1B) Gene family: metallothionein binds to and detoxifies excess copper and other metals, limiting oxidative damage. copper ion binding / metal ion binding
42 CONCLUSIONS This work was helpful to highlight several crucial issues related to gene annotation Networks of paralogs -Are useful for the investigation of the gene annotation of reference and novel genomes -Provide a useful tool for deeper investigations of gene families -Reveal intriguing issues on the genome organization of Arabidopsis thaliana
43 FUNDING & ACKNOWLEDGEMENTS Prof. Luigi Frusciante Maria Luisa Chiusano PRINCIPAL INVESTIGATOR Nunzio D Agostino BIOLOGIST Alessandra Traini PHYSICIST Miriam Di Filippo BIOLOGIST Alessandra Vigilante BIOLOGIST Mara Sangiovanni Chiara Colantuono COMPUTER SCIENTIST STUDENT Mario Aversano TECHNICAL ASSISTANT
44 THANK YOU FOR THE ATTENTION! Reggia di Portici, XVIII century
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