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Home , Fibrobacteres, Methanopyrus, Thermomicrobia, Thermotogae

More Questions Than Answers:

Comparative genomics, evolutionary analysis, and knowing what we do not know about DNA repair.

Jonathan A. Eisen September 20, 2005

TIGRTIGR More Questions Outline • Background • More questions – I. Case study - H. pylori – II. New subfamilies and patterns – III. Recent duplications – IV. Missing pathways – V. Unusual patterns – VI. Even more questions • Answers – Better informatics – Experiments • The end

TIGRTIGR Background

Genome sequencing and evolutionary analysis

TIGRTIGR Fleischmann et al. 1995 TIGRTIGR Prokaryotic

TIGRTIGR From C. M. Fraser TIGRTIGR Why Completeness is Important

• Improves characterization of features • Better comparative genomics • Presence/absence is less subjective • Missing sequence might be important (e.g., centromere) • Allows researchers to focus on biology not sequencing • Facilitates large scale correlation studies • Controls for contamination

TIGRTIGR see Fraser et al. (2002). J. Bacteriol. 184: 6403-5 Analysis of Complete Genomes

• Identification/prediction of • Characterization of features • Characterization of genome features • Prediction of gene function • Prediction of pathways • Integration with known biological data • Comparative genomics

TIGRTIGR “Nothing in biology makes sense except in the light of evolution.”

T. H. Dobzhansky (1973)

TIGRTIGR Evolutionary Perspective and Comparative Biology

• Comparative biology is the analysis of differences and similarities between .

• An evolutionary perspective is useful in such studies because this allows one to focus not just on the levels and degrees of similarity or difference but on how and why similarities and differences came to be.

TIGRTIGR More Questions I: A Case Study The Unbearable Lightness of Helicobacter pylori’s DNA Repair Gene List

TIGRTIGR TIGRTIGR TIGRTIGR Tomb et al. 1997 H. pylori and MutS

• H. pylori has a MutS-like gene but not MutL

• Prior to this genome, all species that encoded a MutS homolog also encoded a MutL homolog

• Experimental studies have shown MutS and MutL always work together in mismatch repair

• Problem: what do we conclude about H. pylori mismatch repair

TIGRTIGR See Eisen et al, 1997 Two Prokaryotic MutS Subfamilies

A. B. B.burgdorferi S pyogenes

5 T.pallidum B. subtilis D. radiodurans Syn. sp MutS2 A. aeolicus A. aeolicus M. genitalium

D. radiodurans 4 3 M. pneumoniae B.burgdorferi * S. pyogenes S. pyogenes B. subtilis

B. subtilis Syn. sp Syn. sp H. pylori MutS1 MutS-I lineage 2 N. gonorrhoaea A. aeolicus MutS-II lineage Gene loss 1 H. influenzae D. radiodurans Species Tree * Gene Duplications 1-5 Gene Loss E. coli B. burgdorferi

TIGRTIGR Eisen, 1998, NAR 26: 4291-4300. H. pylori No MMR

TIGRTIGR PHYLOGENENETIC PREDICTION OF GENE FUNCTION

EXAMPLE A METHOD EXAMPLE B

2A CHOOSE GENE(S) OF INTEREST 5

1 3 4 3A 2 2B 5 1A 2A 1B IDENTIFY HOMOLOGS 6 3B

ALIGN SEQUENCES

1A 2A 3A 1B 2B 3B 1 2 3 4 5 6

CALCULATE GENE TREE

Duplication?

1A 2A 3A 1B 2B 3B 1 2 3 4 5 6

OVERLAY KNOWN FUNCTIONS ONTO TREE

Duplication?

1A 2A 3A 1B 2B 3B 1 2 3 4 5 6

INFER LIKELY FUNCTION OF GENE(S) OF INTEREST Ambiguous Duplication?

Species 1 Species 2 Species 3 1 2 3 4 5 6 1A1B 2A 2B 3A 3B

ACTUAL EVOLUTION (ASSUMED TO BE UNKNOWN) Eisen, 1998, Genome Duplication Res. 8: 163-167. TIGRTIGR More Questions II:

Long Lost Cousins

TIGRTIGR Two UvrA Subfamilies

• Some species have a second UvrA-like gene • Some results on UvrA2 in D. radiodurans • Unclear what UvrA2s do

TIGRTIGR Eisen and Wu, 2002, Theor. Pop. Biol. 61: 481-487. Evolution of UvrA Family

A. ABC Transporters B. UvrA Subfamily UvrA H. influenzae

NrtDC UvrA E. coli

UvrA N. gonorrhoaea

OppDF UvrA R. prowazekii UUP UvrA S. mutans UvrA S. pyogenes

UvrA S. pneumoniae

NodI UvrA B. subtilis

LivF UvrA M. luteus

UvrA M. tuberculosis XylG UvrA M. hermoautotrophicumUvrA1 UvrA H. pylori UvrA1 UvrA C. jejuni

UvrA P. gingivalis

UvrA C. tepidum UvrA2 Duplication in UvrA uvra1 D. radiodurans family PstB UvrA T. thermophilus

UvrA T. pallidum MDR UvrA B. burgdorefi HlyB UvrA T. maritima UvrA A. aeolicus TAP1 UvrA Synechocystis sp. UvrA2 S. coelicolor CFTR, SUR DrrC S. peuceteus UvrA2 UvrA2 D. radiodurans

TIGRTIGR UvrC vs. Cho

• Cho in E. coli is involved in an alternative NER pathway (see Moolenar et al. 2002)

• What do the other relatives of Cho do?

Van Houten et al, 2002, TIGRTIGR PNAS 99: 2581-2583. Three V. cholerae Phr Homologs

Phr.S thyp PHR E. coli MTHF type ORFA00965 phr.neucr Phr.Tricho CPD Phr.Yeast Phr.B firm Photolyases phr.strpy phr.haloba PHR STRGR pCRY1.huma phr.mouse phr2.human 6-4 phr2.mouse phr.drosop Photolyases phr3.Synsp ORF02295.Vibch phr.neigo ORF01792.Vibch Phr.Adiant Phr2.Adian Phr3.Adian Blue phr.tomato CRY1 ARATH phr.phycom Light CRY2 ARATH PHH1.arath Receptors PHR1 SINAL phr.chlamy PHR ANANI phr.Synsp 8-HDF type PHR SYNY3 phr.Theth Photolyase Rh.caps TIGRTIGR Alkylation Repair Genes

Ada E. coli Ada H. infl Ogt E. coli Ogt H. infl Ogt Gram+ Ogt D. radio MGMT Euks AlkA Gram+ AlkA E. coli

AlkA Domain (O6-Me-G glycosylase) Ogt Domain (O6-Me-G alkyltransferase) Ada Domain (transcriptions regulator)

TIGRTIGR Eisen and Hanawalt, 1999, Mut. Res. 435: 171-213. More Questions III:

Does 1+1 = 1? Or 2?

TIGRTIGR Wolbachia pipientis wMel

TIGRTIGR Wu et al, 2004, PLOS Biology 2: 327-341 MutL Duplication in Wolbachia

TIGRTIGR Eisen, unpublished. Agrobacterium tumefaciens

Wood et al., 2001, Science 294: 2317-2323 TIGRTIGR Agrobacterium tumefaciens has multiple duplications

Chromosome1 DnaE, LigA, Xth Chromosome2 DnaE, LigA Plasmid1 DnaE, LigA, Xth Plasmid2 DnaE?, LigA

TIGRTIGR Wood et al., 2001, Science 294: 2317-2323 Duplication of UVDE in Anthrax

Schizosaccharomyces pombe Neurospora crassa perfringens Bacillus cereus Bacillus anthracis UVDE2 Bacillus halodurans Bacillus anthracis UVDE1 radiodurans Nostoc sp. PCC 7120

TIGRTIGR Eisen,unpublished. More Questions IV:

When a good repair pathway is hard to find

TIGRTIGR P. falciparum DNA Repair

• P. falciparum initial annotation showed the presence of a DNA Ligase IV homolog but not a DNA Ligase I.

• Yet Ligase I has more of a core function in other , while Ligase IV is involved in NHEJ DNA Ligase Tree

Arabidopsis thalianaGP9651815g Drosophila melanogasterGP72929 Homo sapiensSPP49917DNL4 HUMAN Gallus gallusGP15778121dbjBAB6 Xenopus laevisGP18029886gbAAL5 Candida albicansSPP52496DNLI C Ligase IV Saccharomyces cerevisiaeGP1151 Schizosaccharomyces pombeGP700 Camelpox virusGP18483081gbAAL7 Variola major virusGP439074gbA Cowpox virusGP20153167gbAAM136 Vaccinia virusGP2772802gbAAB96 VIRUS vaccinia 9791118refNP 06 Vaccinia virus strain Tian Tan Monkeypox virusGP17529940gbAAL Homo sapiensSPP49916DNL3 HUMAN Mus musculusGP1794221gbAAC5300 Viral Ligases Xenopus laevisGP18029884gbAAL5 lumpy skin disease virusGP1514 Swinepox virusGP18448623gbAAL6 Myxoma virusGP6523988gbAAF1502 Rabbit fibroma virusGP392838gb Fowlpox virusGP453602embCAA828 Drosophila melanogasterGP72996 Arabidopsis thalianaSPQ42572DN Oryza sativaGP16905197gbAAL310 Crithidia fasciculataGP312384e Caenorhabditis elegansSPQ27474 Drosophila melanogasterGP72916 Homo sapiensSPP18858DNL1 HUMAN Mus musculusSPP37913DNL1 MOUSE Rattus norvecusSPQ9JHY8DNL1 RA Xenopus laevisSPP51892DNL1 XEN Ligase I Plasmodium falciparumGP1815859 Schizosaccharomyces pombeSPP12 Saccharomyces cerevisiaeSPP048 Aeropyrum pernixSPQ9YD18DNLI A Acidianus ambivalensSPQ02093DN Sulfolobus solfataricusSPQ980T Sulfolobus shibataeSPQ9P9K9DNL Sulfolobus tokodaiiSPQ976G4DNL aeolicusGP2983805gbAAC Aquifex aeolicusSPO67398DNLI A Pyrobaculum aerophilumGP409906 uncultured crenarchaeote 74A4G Thermoplasma acidophilumSPQ9HJ Thermoplasma volcaniumOMNINTL0 Methanosarcina acetivorans str Archaeoglobus fuldusSPO29632DN A METAC 19916535gbAAM05952.1 D Pyrococcus abyssiSPQ9V185DNLI Archaeal Ligases Pyrococcus horikoshiiSPO59288D Pyrococcus furiosusSPP56709DNL Thermococcus kodakaraensisGP10 Thermococcus fumicolansSPQ9HH0 kandleri AV19GP19 jannaschiiSPQ576 sp.SPQ9HR35DNLI Streptomyces coelicolorSPQ9FCB Lymantria dispar nucleopolyhed TIGRTIGR Eisen, unpublished DNA Ligase IV SubTree

Arabidopsis thaliana Drosophila melanogaster Homo sapiens Gallus gallus Xenopus laevis Candida albicans Saccharomyces cerevisiae Schizosaccharomyces pombe TIGRTIGR Eisen, unpublished DNA Ligase I SubTree Arabidopsis thaliana Oryza sativa Crithidia fasciculata Caenorhabditis elegans Drosophila melanogaster Homo sapiens Mus musculus Rattus norvecus Xenopus laevis Plasmodium falciparum Schizosaccharomyces pombe Saccharomyces cerevisiae TIGRTIGR Eisen, unpublished No NHEJ in P. falciparum?

• No Ligase IV • None of the other “normal” NHEJ genes for eukaryotes • Does it use only homologous recombination or is there another NHEJ pathway?

TIGRTIGR See Gardner et al 2002 Wolbachia pipientis wMel

TIGRTIGR Wu et al, 2004, PLOS Biology 2: 327-341 Wolbachia Evolves Rapidly

Wu et al., 2004 TIGRTIGR Insect Symbiont Tree

TIGRTIGR Wu et al. In preparation Intracellular and Evolutionary Rate

• Many intracellular have higher evolutionary rates than free-living relatives

• Two main theories to explain this: – loss of DNA repair – altered population genetics (e.g., bottlenecks)

TIGRTIGR Wolbachia Has Similar Repair Genes to More Slowly Evolving Species

Repair Category Gene Name

MMR mutS1, mutL1, mutL2 NER uvrABCD BER fpg, mpg, nth AP Endo xth Rec recA, ruvABC, recG, recFJR Other ligA, ssb, radA

TIGRTIGR Wu et al., 2004 Insect Symbiont Repair Buchnera Buchnera Buchnera Wigglesworthia Blochmannia Baumannia (APS) (BP) (SG) Base Excision Repair Ung gi15616802 gi27904674 - gi32491032 gi33519990 GEBCORF00107 MutY gi15617145 gi27904974 gi21672797 - gi33519714 - Fpg - - - - - GEBCORF00321 Nfo gi15616757 gi27904631 gi21672420 - - GEBCORF00528 Xth - - - gi32491120 gi33519889 -

Nucleotide Excision Repair UvrD - - - gi32491021 - GEBCORF00274

Transcription-Repair Coupling Factor Mfd gi15616905 gi27904769 - gi32490848 - -

Mismatch Repair MutL gi15617161 gi27904987 gi21672810 - - GEBCORF00079 MutS gi15617028 gi27904861 gi21672681 - - GEBCORF00349 MutH - gi27904531 - - - GEBCORF00038

8-oxo-dGTP Hydrolysis MutT gi15616821 - gi21672482 - - -

Pyrimidine Dimer Repair Phr gi15616910 gi27904772 - gi32490931 - -

Homologous Recombination RecG - - - - - GEBCORF00259 RuvA - - - - - GEBCORF00452 RuvB - - - - - GEBCORF00451 RuvC - - - gi32490863 - GEBCORF00453 RecB gi15617053 gi27904885 gi21672704 gi32491015 gi33519733 GEBCORF00043 RecC gi15617052 gi27904884 gi21672703 gi32491014 gi33519731 GEBCORF00042 RecD gi15617054 gi27904886 gi21672705 gi32491016 gi33519732 GEBCORF00044 RecA - - - gi32490984 - GEBCORF00346 RecJ - - - gi32491188 - GEBCORF00298 RmuC - - - - gi33520059 GEBCORF00163 TIGRTIGR Wu et al. In preparation Insect Nutritional Symbiont Tree Mut RecA LS - + - - + - + - + + + +

TIGRTIGR Wu et al. In preparation Organelle Derived Genes in the A. thaliana Nuclear Genome

Pathway Genes Mismatch Repair MutS2 Base Excision Repair Fpg, Tag? Excision Mfd Repair Recombination RecA, RecG Direct Repair Phr Other Lon

TIGRTIGR Eisen and Britt, 2000 More Questions V:

One of these things just does not belong here

TIGRTIGR Other Unusual Patterns

• RecBCD but no RecA • Mfd but no UvrABCD • No Ung in • Rad25 in some bacteria • No MSH4, 5 in many eukaryotes • No CSB in some eukaryotes • Tons of repair genes in Mimivirus

TIGRTIGR

• NER – UvrABC in some – Rad1, Rad2, Rad25 etc in others – Some species with both • MMR – Many species with MutS2 only – Some with multiple MutLs, MutS1s – Three MutSs in

TIGRTIGR Even More Questions?

You ain’t seen nothing yet

TIGRTIGR Biased Sampling of Genomes

• Most bacterial genomes come from three phyla • Most phyla have no genome sequences • Even worse sampling for eukaryotes and archaea

Hugenholtz 2002 TIGRTIGR TM6 OS-K • At least 40 Termite Group phyla of OP8 bacteria Chlorobi Marine GroupA WS3 Gemmimonas OP9 Synergistes Deferribacteres Chrysiogenetes NKB19 Chlamydia OP3 Spriochaetes Coprothmermobacter OP10 TM7 Deinococcus- Dictyoglomus Thermudesulfobacteria OP1 OP11 TIGRTIGR Proteobacteria TM6 OS-K • At least 40 Acidobacteria Termite Group phyla of OP8 Nitrospira Bacteroides bacteria Chlorobi Fibrobacteres Marine GroupA • Genome WS3 Gemmimonas sequences are Firmicutes Fusobacteria mostly from Actinobacteria OP9 Cyanobacteria three phyla Synergistes Deferribacteres Chrysiogenetes NKB19 Verrucomicrobia Chlamydia OP3 Planctomycetes Spriochaetes Coprothmermobacter OP10 Thermomicrobia Chloroflexi TM7 Deinococcus-Thermus Dictyoglomus Aquificae Thermudesulfobacteria Thermotogae OP1 TIGRTIGR OP11 Proteobacteria TM6 OS-K • At least 40 Acidobacteria Termite Group phyla of OP8 Nitrospira Bacteroides bacteria Chlorobi Fibrobacteres Marine GroupA • Genome WS3 Gemmimonas sequences are Firmicutes Fusobacteria mostly from Actinobacteria OP9 Cyanobacteria three phyla Synergistes Deferribacteres Chrysiogenetes • Some other NKB19 Verrucomicrobia Chlamydia phyla are OP3 Planctomycetes Spriochaetes only sparsely Coprothmermobacter OP10 sampled Thermomicrobia Chloroflexi TM7 Deinococcus-Thermus Dictyoglomus Aquificae Thermudesulfobacteria Thermotogae OP1 TIGRTIGR OP11 Proteobacteria TM6 OS-K • At least 40 Acidobacteria Termite Group phyla of OP8 Nitrospira Bacteroides bacteria Chlorobi Fibrobacteres Marine GroupA • Genome WS3 Gemmimonas sequences are Firmicutes Fusobacteria mostly from Actinobacteria OP9 Cyanobacteria three phyla Synergistes Deferribacteres Chrysiogenetes • Some other NKB19 Verrucomicrobia Chlamydia phyla are OP3 Planctomycetes Spriochaetes only sparsely Coprothmermobacter OP10 sampled Thermomicrobia Chloroflexi TM7 • Solution: Deinococcus-Thermus Dictyoglomus Aquificae sequence Thermudesulfobacteria Thermotogae OP1 more phyla TIGRTIGR OP11 Solution II: Environmental Sequencing

Warner Brothers, Inc. shotgun

sequence

TIGRTIGR Duplication and Loss of UVDE

Schizosaccharomyces pombe Neurospora crassa Clostridium perfringens Bacillus subtilis Bacillus cereus Bacillus anthracis UVDE2 Bacillus halodurans Bacillus anthracis UVDE1 Nostoc sp. PCC 7120 TIGRTIGR 0.1

TIGRTIGR Eisen, unpublished 0.1

Haloarcula_marismortui_ATCC_43

Nostoc_sp._PCC_7120_gi17228758 Deinococcus_radiodurans_gi1580 Parachlamydia_sp._UWE25_gi4644

Desulfotalea_psychrophila_LSv5

Mimivirus_gi55819554-NC_006450 Cryptococcus_neoformans_var._n Schizosaccharomyces_pombe_gi19 Thermus_thermophilus_HB27_gi46 Clostridium_perfringens13_gi18 Clostridium_perfringens_SM101 B_anthracisSterne_gi49188182-N B_anthracisAmes_gi30265371-NC_ B_anthracis''_gi47530914-NC_00 B_thuringiensi_gi49481769-NC_0 B_cereus_ZK_gi52140215-NC_0062 B_cereus_ATCC_10987_gi42784523 B_cereus_ATCC_14579_gi30023378 B_thuringiensi_gi49481707-NC_0 B_anthracisAmes_gi30260424-NC_ B_anthracis''_gi47525505-NC_00 B_anthracisSterne_gi49183267-N Eisen, unpublished B_cereus_ZK_gi52144992-NC_0062 B_cereus_ATCC_10987_gi42779349 B_cereus_ATCC_14579_gi30018497 TIGR B_halodurans_gi15615160-NC_002 TIGR Bacteriophage_Aeh1_gi38640041- 0.1

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Better Bioinformatics

TIGRTIGR Non Homology Based Functional Prediction: Phylogenetic Profiles

TIGRTIGR Non-Homology Prediction: Phylogenetic Profiles

• Step 1: Search all genes in organisms of interest against all other genomes • Ask: Yes or No, is each gene found in each other species • Cluster genes by distribution patterns (profiles)

TIGRTIGR MutL and MutS1 Always Together

Table 3. Presence of MutS Homologs in Complete Genomes Sequences

Species # of MutS Which MutL Homologs Subfamilies? Homologs

Bacteria Escherichia coli K12 1 MutS1 1 Haemophilus influenzae Rd KW20 1 MutS1 1 Neisseria gonorrhoeae 1 MutS1 1 Helicobacter pylori 26695 1 MutS2 - genitalium G-37 - - - Mycoplasma pneumoniae M129 - - - Bacillus subtilis 169 2 MutS1,MutS2 1 pyogenes 2 MutS1,MutS2 1 tuberculosis - - - Synechocystis sp. PCC6803 2 MutS1,MutS2 1 pallidum Nichols 1 MutS1 1 Borrelia burgdorferi B31 2 MutS1,MutS2 1 Aquifex aeolicus 2 MutS1,MutS2 1 Deinococcus radiodurans R1 2 MutS1,MutS2 1

Archaea Archaeoglobus fulgidus VC-16, DSM4304 - - - Methanococcus janasscii DSM 2661 - - - Methanobacterium thermoautotrophicum ΔH 1 MutS2 -

Eukaryotes Saccharomyces cerevisiae 6 MSH1-6 3+ Homo sapiens 5 MSH2-6 3+ TIGRTIGR Co-Occurrence of Spo Genes

TIGRTIGR Co Occurrence of UvrABC

TIGRTIGR Co-Occurrence of Muk, SeqA, MutH

TIGRTIGR Answers II:

Novelty is good and bad

TIGRTIGR Deinococcus radiodurans

TIGRTIGR DNA Repair Genes in D. radiodurans Complete Genome

Process Genes in D. radiodurans

Nucleotide Excision Repair UvrABCD, UvrA2 Base Excision Repair AlkA, Ung, Ung2, GT, MutM, MutY-Nths, MPG AP Endonuclease Xth Mismatch Excision Repair MutS, MutL Recombination Initiation RecFJNRQ, SbcCD, RecD Recombinase RecA Migration and resolution RuvABC, RecG Replication PolA, PolC, PolX, phage Pol Ligation DnlJ dNTP pools, cleanup MutTs, RRase Other LexA, RadA, HepA, UVDE, MutS2

TIGRTIGR White et al, 1999 Problem:

List of DNA repair gene homologs in D. radiodurans genome is not significantly different from other bacterial genomes of the similar size

TIGRTIGR Proteobacteria TM6 OS-K • At least 40 Acidobacteria Termite Group OP8 phyla of Nitrospira Bacteroides bacteria Chlorobi Fibrobacteres Marine GroupA WS3 Gemmimonas Firmicutes Fusobacteria Actinobacteria OP9 Cyanobacteria Synergistes Deferribacteres Chrysiogenetes NKB19 Verrucomicrobia Chlamydia OP3 Planctomycetes Spriochaetes Coprothmermobacter OP10 Thermomicrobia Chloroflexi TM7 Deinococcus-Thermus Dictyoglomus Aquificae Thermudesulfobacteria Thermotogae OP1 OP11 TIGRTIGR Proteobacteria TM6 OS-K • At least 40 Acidobacteria Termite Group phyla of OP8 Nitrospira Bacteroides bacteria Chlorobi Fibrobacteres Marine GroupA • Most DNA WS3 Gemmimonas metabolism Firmicutes Fusobacteria studies in two Actinobacteria OP9 Cyanobacteria phyla Synergistes Deferribacteres Chrysiogenetes NKB19 Verrucomicrobia Chlamydia OP3 Planctomycetes Spriochaetes Coprothmermobacter OP10 Thermomicrobia Chloroflexi TM7 Deinococcus-Thermus Dictyoglomus Aquificae Thermudesulfobacteria Thermotogae OP1 TIGRTIGR OP11 Proteobacteria TM6 OS-K • At least 40 Acidobacteria Termite Group phyla of OP8 Nitrospira Bacteroides bacteria Chlorobi Fibrobacteres Marine GroupA • Most DNA WS3 Gemmimonas metabolism Firmicutes Fusobacteria studies in two Actinobacteria OP9 Cyanobacteria phyla Synergistes Deferribacteres Chrysiogenetes • Deinococcus NKB19 Verrucomicrobia Chlamydia is distant OP3 Planctomycetes Spriochaetes from any Coprothmermobacter OP10 well studied Thermomicrobia Chloroflexi TM7 Deinococcus-Thermus Dictyoglomus Aquificae Thermudesulfobacteria Thermotogae OP1 TIGRTIGR OP11 Proteobacteria TM6 OS-K • At least 40 Acidobacteria Termite Group phyla of OP8 Nitrospira Bacteroides bacteria Chlorobi Fibrobacteres Marine GroupA • Most DNA WS3 Gemmimonas metabolism Firmicutes Fusobacteria studies in two Actinobacteria OP9 Cyanobacteria phyla Synergistes Deferribacteres Chrysiogenetes • Deinococcus NKB19 Verrucomicrobia Chlamydia is distant OP3 Planctomycetes Spriochaetes from any Coprothmermobacter OP10 well studied Thermomicrobia Chloroflexi TM7 organism Deinococcus-Thermus Dictyoglomus Aquificae • Solution: Thermudesulfobacteria Thermotogae OP1 more studies TIGRTIGR OP11 $$$ J. C. Venter C. Fraser

M-I Benito Other people DOE ONR NSF N. Moran H. Tettelin N. Ward NIH F. Robb H. Ochman T.Read S. Salzberg J. Battista E. Orias J. Heidelberg D. Bryant R. Myers S. O’Neill O. White M. Eisen W. Martin I. Paulsen

P. Hanawalt

C. M. Cavanaugh TIGR

Mom and Dad

TIGRTIGR