Genomics of Microbial Eukaryotes

Igor GrigorievFungal Genomics Program Head

US DOE Joint Genome Institute, Walnut Creek, CA

<ivgrigoriev@lbl.gov>

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Outline

Eukaryotic Genome Annotation

Fungal Genomics Program

MycoCosm

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Are you in the right room?

genome.jgi.doe.gov

IMG

MycoCosm

100+ annotated eukaryotic genomes

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Started with Human Genome Project

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Protein-based methods build CDS exons around known protein alignments.(Fgenesh, GeneWise)

GenBank protein

Transcript-based methods map or assemble transcripts on the genome, including UTRs (EST_map, Combest)

EST contig

Predict model

Predict model

Ab initio methods use knowledge of known genes’ structures to predict start, stop, and splice sites in CDS only. (Fgenesh+, GeneMark)

Train on known genes

ATG TGA

GT AG

exons introns5’UTR5’UTR3’UTR3’UTR

Promoter PolyA

Gene model

Eukaryotic Gene Prediction

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Predicted protein

Protein Annotation

Higher order assignments:

Gene Ontology terms

EC numbers --> KEGG pathways

Gene families, with and without other species

Possible orthologs

(in nr, SwissProt, KEGG, KOG)

Possible paralog

(Blastp+MCL)

Domain

(InterPro, tmhmm)

Signal peptide

(signalP)

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EST Support is Critical for Eukaryotes

0%10%20%30%40%50%60%70%80%90%

100%

N.heam

atoco

cca

L.bic

olor

P.blake

sleea

nus

S.rose

us

M.g

ram

inico

la

A.niger

*

N.disc

reta

*

W.co

cos

G.trab

eum

A.acu

leatu

s

Other Genes

Supported by ESTs

Sanger 454 Illumina

5531

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EST profile

CombEST gene models

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Best Models

FGENESH

Representative set

GENEWISE

EXTERNAL MODELS

Multiple gene predictors offer several different gene models at each gene locus; A single best model from each locus is automatically selected based on homology and EST support; These compose a non-redundant (or Filtered) gene set for further analysis This set is further improved during community-driven manual curation

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Genomic assembly and EST contigs

An

no

tati

on

Pip

elin

e

Gene predictions Gene predictions

Protein annotationsProtein annotations

Transcript + protein mapsTranscript + protein maps

Repeat maskRepeat mask

Manual curationManual curation

Bring it all together

Analysis

Gene familiesGene expressionPhylogenomicsProteomicsProtein targetingetc

Annotation

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Many Genes of Eco-responsive Daphnia pulex

First crustacean, aquatic animal sequenced, new model organism30,940 predicted D.pulex genes in ~200Mb genome85% supported by 1+ lines of evidence Colbourne et al, Science, 2011

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Half of Daphnia Genes have no Homologs

With Evgeny Zdobnov’s group (Univ. Genève)

* Of 716 highly conserved single copy orthologs, Daphnia is missing only two

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Outline

Eukaryotic Genome Annotation

Fungal Genomics Program

MycoCosm

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Fungal Genomics for Energy and Environment

Grow Grow DegradeDegrade

Lignocellulose degradation

Plant symbiontsand pathogens

SugarFermentation

FermentFerment

Bio-refinery

GOAL: Scale up sequencing and analysis of fungal diversity for DOE science and applications

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• Plant feedstock health• Symbiosis

• Plant Pathogenicity

• Biocontrol

• Biorefinery fungi• Lignocellulose degradation

• Sugar fermentation

• Industrial organisms

• Fungal diversity• Phylogentic

• Ecologic

Genomic Encyclopedia of Fungi Launched

www.jgi.doe.gov/fungi 100+ fungal genomes600+ registered users5000+ visitors/month

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Distinct Mechanisms of Cellulose Degradation

White rotP.chrysosporium

Cellobiohydrolase IIGH6(CBH50)

Cellobiohydrolase IGH7 (CBH58,62)Endoglucanases

GH5-CBM1,GH12

GH3 -glucosidase

Cellulose

No cellulose binding domain CBM1 in brown rot!

Fe2+ + H2O2 Fe3+ + HO- + HO.

Fe3+

Glucose

Copper radical oxidasesGlucose oxidases

Iron reductase

Brown rotPostia placenta

Martinez et al, PNAS 2009

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Diverse Basidiomycota

• FGP09 pilots• Basidio jam (Mar 2010)• 3 CSP11 proposals• Basidio jam (Mar 2011)

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Future Grand Challenges

Fungal isolates & groups

Systems of interacting organisms

Systems in wild

MODELING

FUNCTION

SEQUENCE

1. 1000 fungal genomessampling fungal diversity

2. Model fungisampling 100s of conditions

3. Fungal ecosystems: Bioenergy crops symbionts & pathogens Biorefinery Fungal metagenomes

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Leadership in Sequencing Fungi

68%

23%

Ascomycota

Basidiomycota

Blastocladiomycota

Chytridiomycota

Glomeromycota

Microsporidia

Neocallimastigomycota

Unknown

Zygomycota

13%10%

31%

5%

41%

DOE Joint GenomeInstitute

Broad Institute

Sanger Institute

Washington Univ

other

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Annotation and Analysis Tools

• Automated Annotation• Pipeline

• Genomics Analysis Platform• Genome Centric• Comparative Genomics

• Community Resource• Integrated data• User tools• Training

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Genome-Centric View

Comparative View

www.jgi.doe.gov/fungi

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Genome-Centric View

Focus: functional genomics, user data deposition and curation

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New Comparative View

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Community Building Tools

•

• Jamborees: • Genome analysis for publications

• MycoCosm Tutorials: • On-line video, MGM, workshops w/ large

meetings (Asilomar, JGI Users, MSA)

• Preparation for CSP: • Large meetings and focused groups

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Summary

Eukaryotic Annotation Recipe:• Combined gene predictors,

experimental data, and community annotation

Fungal Genomics Program: • Scaled-up sequencing &

comparative analysis of fungi relevant for energy & environment (jgi.doe.gov/fungi)

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Outline

Eukaryotic Genome Annotation

Fungal Genomics Program

MycoCosm