Talk by Jonathan Eisen on Phylogenomics of DNA repair at Lake Arrowhead Small Genomes meeting in 2000.

1. TIGR

2. Topics of Discussion
• DNA Repair
• Why study evolution of repair?
• Evolution of specific pathways with examples
from recent genome projects (e.g., A. thaliana,
Vibrio cholerae, Shewanella putrefaciens,
Buchnera aphidicolum symbiont)
• Big picture – evolutionary origins of repair
TIGR

3. Damage is not just to DNA
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4. TIGR

5. General Mechanisms of Resistance to
Cellular Damaging Agents
• Damage protection/prevention
• Damage tolerance
• Repair and recovery
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6. Classes of DNA Repair
• Direct repair
– Photoreactivation
– Alkylation transfer
– DNA ligation/non-homologous end joining
• Excision repair
– Base excision repair
– Mismatch excision repair
– Nucleotide excision repair
• Recombinational repair
TIGR

7. Excision Repair Outline
NUCLEOTIDE BASE EXCISION
and
M ISM ATCH EXCISION
Damage
N-glycosylase
Re cognition
Endonucle ase AP e ndo
* *
Exonucle ase ,
He licase , Exonucle ase ,
Polyme rase Polyme rase
Ligase
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8. Recombination Outline
RecBCD Generation of
RecE,T single-strand
RecQ,J overhang
Rad50, M 11, XRS2
RE
RecA Initiation,
RecFOR alignment
Rad 52
RecA
Rad51,55,57 Strand invasion
DNA synthesis
RuvABC
RecG,RUS?
Branch migration
Rad54? and resolution
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9. “Nothing in biology makes sense
except in the light of evolution.”
T. H. Dobzhansky (1973)
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10. Why Study Evolution and Repair?
• Repair variation leads to differences in evolutionary
patterns within and between species.
• Evolutionary analysis can identify mutation/repair biases.
• Evolutionary studies can improve our understanding of
repair proteins and pathways.
• Comparisons of repair genes can be used to infer
evolutionary history.
• Information on mutation processes improves sequence and
phylogenetic analysis.
• Evolutionary analysis is required to infer the origins and
history of repair processes.
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11. Steps in Phylogenomic Analysis
• Create database of genes of interest
• Presence/absence of homologs in complete genomes
• Phylogenetic trees of each gene family
• Infer evolutionary events (gene origin, duplication, loss and
transfer)
• Refine presence/absence (orthologs, paralogs, subfamilies)
• Functional predictions and functional evolution
• Analysis of pathways
TIGR

12. Nucleotide Excision Repair
Pathway Biochemical Activity(s). |-------------------------------------Bacteria------------------------------------| |-----Archaea------| |--Eukarya---|
Protein Name(s)
Bacter ial NER
UvrA Binds damaged DNA + + + + + + + + + + + + + + + - - - - -
UvrB Helicase, 3' incision endonuclease + + + + + + + + + + + + + + + - - - - -
UvrC 5' incision endonuclease + + + + + + + + + + + + + + + - - - - -
UvrD Excision helicase + + + + + ++ + + + ++ + + + + ++ - - - + +
MFD Transcription repair coupling + + + + + + - - + + + + - + - - - - - -
Eukaryotic NER
Recognition
Rad14 (XPA) Binds damaged DNA - - - - - - - - - - - - - - - - - - + + +
RFA1/RPA1 ssDNA binding w/ RFA2,3 - - - - - - - - - - - - - - ± - - - + + +
RFA2/RPA2 ssDNA binding w/ RFA1,3 - - - - - - - - - - - - - - - - - - + ++ +
RFA3/RPA3-human ssDNA binding w/ RFA1,2 - - - - - - - - - - - - - - - - - - - + +
RFA3/RPA3-yeast ssDNA binding w/ RFA1,2 - - - - - - - - - - - - - - - - - - + +
Initiation
Rad3 (XPD) (ERCC2) TFIIH component – helicase - - - - - - - - - - - - - - - - - ± + + +
Rad25 (XPB) (ERCC3) TFIIH component – helicase - - - - - - - - - + - + - + - - + + + + +
SSL1 (p44) TFIIH component - - - - - - - - - - - - - - - - - - + + +
TFB1 (p62) TFIIH component - - - - - - - - - - - - - - - - - - + + +
TFB2 (p52) TFIIH component - - - - - - - - - - - - - - - - - - + + +
TFB3 (MAT1) TFIIH component - - - - - - - - - - - - - - - - - - + + +
TFB4 (p34) TFIIH component - - - - - - - - - - - - - - - - - - + + +
CCL1 (CyclinH) TFIIH component - - - - - - - - - - - - - - - - - - + + +
Kin28 (CDK7) TFIIH component - protein kinase - - - - - - - - - - - - - - - - - - + + +
Incision
Rad2 (XPG) (ERCC5) 3' incision (flap endonuclease) - - - - - - - - - - - - - - + + + + + + +
Rad10 (ERCC1) 5' incision endonuclease w/ Rad1 - - - - - - - - - - - - - - - - - - + + +
Rad1 (XPF) (ERCC4) 5' incision endonuclease w/ Rad10 - - - - - - - - - - - - - + + + + + + +
Specificity
Rad4 (XPC) Repair of inactive DNA - - - - - - - - - - - - - - - - - - + + +
Rad23 (HHRAD23) Repair of inactive DNA - - - - - - - - - - - - - - - - - - + ++ +
Rad7 Repair of inactive DNA - - - - - - - - - - - - - - - - - - + +
Rad16 Repair of inactive DNA - - - - - - - - - - - - - - - - - - + + +
Rad26 (CSB) (ERCC6) Transcription-repair coupling - - - - - - - - - - - - - - - - - - + + +
CSA (ERCC8) Transcription-repair coupling - - - - - - - - - - - - - - - - - - ± + +
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13. Recombinational Repair
Pathway Biochemical Activity(s). |-------------------------------------Bacteria------------------------------------| |-----Archaea------| |--Eukarya---|
Protein Name(s)
Initiation
RecBCD pathway
RecB ExoV H elicase + + + - - - - - - + + - - + - - - - - -
RecC ExoV Nuclease + + + - - - - - - + ±+ - - + - - - - - -
RecD ExoV Helicase + + + - ± ± - - - + ±+ - - + - - - - - -
RecF pathway
RecF Assists RecA filamentation + + - - + + - - + + - + - + - - ± - ± ±
RecJ 5'-3' ssDNA exonuclease + + + + + + - - + - + + + + - - - - - -
RecO Binds ssDNA, assists RecF? + + + - + + - - + + - - - + - - - - - -
RecR ATP binding, assists RecF? + + + ±+ + + - - + + - + + + - - - - - -
RecN ATP binding + + + + + + - - + + - + + + - - ± - - -
RecQ 3'-5' DNA helicase + + + - ± + - - + - - + - + - - - - + ++ +
RecE pathway
RecE/ExoVIII 5'-3' dsDNA exonuclease + - - - - - - - - - - - - + - - - - - -
RecT Binds ssDNA, promotes pairing + - - - + + - - - - - - - + - - - - - -
SbcBCD pathway
SbcB/ExoI 3'-5' ssDNA exonuclease + + - - - - - - - - - - - + - - - - - -
SbcC dsDNA exonuclease (w/ sbcD) + - - - ±+ + - - + - + + + + ± ± ± ± ± ± ±
SbcD dsDNA exonuclease (w/ sbcC) + - - - - + - - + - + + + + ± ± ± ± ± ± ±
AddAB Pathway
AddA/RexA Exonuclease + helicase w/ AddB - - + - + + - - - - - + - + - - - - - -
AddB/RexB Exonuclease + helicase w/ AddA - - + - + + - - - - - - - + - - - - - -
Rad52 pathway
Rad52, Rad59 n/a - - - - - - - - - - - - - - - - - - ++ + +
M re11/Rad32 Nuclease w/ Rad50 ± - - - ± ± - - ± - ± ± ± ± + + + + + + +
Rad50 Nuclease w/ M re11 ± - - - ± ± - - ± - ± ± ± ± + + + + + + +
Recom binas e
RecA, Rad51 DNA binding, strand exchange + + + + + + + + + + + + + + + + + + ++ ++ ++
Br anch m igr ation/r esolution
Branch migration
RuvA Binds junctions. Helicase w/ RuvB + + + + + + + + + + + + - + - - - - - -
RuvB 5'-3' junction helicase w/ RuvA + + + + + + + + + + + + - + - - - - - -
RecG Resolvase, 3'-5' junction helicase + + + + + + - - + + + + + + - - - - - -
Resolvases
RuvC Junction endonuclease + + + + - - - - + + - + - + - - - - - -
RecG Resolvase, 3'-5' junction helicase + + + + + + - - + + + + + + - - - - - -
Rus Junction endonuclease + - - - - - - - - ±+ - - ±+ + - - - - - -
CCE1 Junction endonuclease - - - - - - - - - - - - - - - - - - + +
O ther r ecom bination pr oteins
Rad54 n/a - - - - - - - - - - - - - - - - - - + + +
Rad55 n/a - - - - - - - - - - - - - - - - - - + + +
Rad57 n/a - - - - - - - - - - - - - - - - - - + + +
Xrs2 Assists Rad50/M RE11? - - - - - - - - - - - - - - - - - - + +
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14. Evolution of Specific Pathways
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15. Photoreactivation and Photolyases
• All photoreactivation is carried out by enzymes in the photolyase
family
• Two main classes of photolyases – class I and class II – are
distantly related to each other and likely the result of an ancient
duplication
• PhrI and PhrII missing from most species for which complete
genomes are available.
• Many cases of functional change (e.g., CPD -> 6-4) and some are
not even involved in DNA repair
• Many of the eukaryotic proteins appear to be of an organellar
ancestry
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16. Uses of Evolution : Photoreactivation
• All known enzymes that perform photoreactivation are part of
a single large photolyase gene family
• Some members of the family do not function as photolyases,
but instead work as blue-light receptors
• If a species does not encode a member of the photolyase gene
family, it likely does not have photoreactivation capability
• If a species encodes a photolyase, one cannot conclude it has
photolyase activity
• Position of photolyase homologs within photolyase tree helps
predict what activities they have
TIGR

17. Phr.S thyp
PHR E. coli
O R FA0 0 9 6 5* * * * * * * * *
p hr.neucr
M T H F ty pe
Phr.Tricho Class I CPD
Phr.Yeast Photoly ases
Phr.B firm
p hr.strp y
p hr.haloba
PHR STRGR
p C RY1.huma
p hr.mouse
p hr2.human
p hr2.mouse 6-4
p hr.drosop Photoly ases
phr3.Synsp
O R F0 2 2 9 5.V ib ch* * * * * * * *
p hr.neigo
O RF0 1 7 9 2 .V ib ch* * * * * * *
Phr.Adiant
Phr2.Adian
Phr3.Adian
p hr.tomato Blue
C RY1 ARATH
p hr.phycom
Light
C RY2 ARATH
Receptors
PHH1.arath
PHR1 SINAL
p hr.chlamy
PHR ANANI
p hr.Synsp
8-H DF ty pe
PHR SYNY3
CPD
TIGR p hr.Theth
Rh.cap s Photoly ases

18. Photolyases in A. thaliana
phr.chlamy
cry2.tomat
PHH1.CRY2.
PHR1 SINAL
Cry3.Adian Crys Group with
CRY1/hy4.A
phr.Brevib
Phr.Cordi. α-Proteobacteria
ORF05094.C
Phr.Rhoca
Phr.Bacfi
Phr.Entfa
Phr.Strpy
Phr.Pseae.
Phr.Yerpe.
Phr.Ecoli
Phr.S thyp
Phr.Salty
Phr.Shepu.
A00965.Vib
Phr.Yeast
Other Bacteria
Phr.Neucr
Phr.Tricho
Phr.SYNY3
Phr.Anani
Phr.Synsp
Phr.Theth
Phr.Mycav.
Phr.Mycsm
CT12574.Fl
ARATH3 MSJ
Phr.fly
pCRY1.huma
PhrL.Mouse
Eukaryotic
PhrL2.huma
PhrL2.Mous
295.Vibch
Phr2.Synsp
ARATH2 T30
ARATH5 F6A Cyano/Plastid
1792.Vibch
TIGR Phr.Neime
Phr.Neigo
Phr.Halha
Phr.Strgr

19. Alkyltransferases
• All known alkyltransferases are members of a single
gene family
• Found in most but not all species
• Likely present in LUCA
• Ada protein in E. coli originated by fusion between
an alkyltransferase and a transcription-regulatory
domain
• Gram-positive bacteria have the Ada domain fused to
an alkylation glycosylase instead of alkyltransferase
TIGR

20. Alkylation Repair Genes
Ada E. coli
Ada H. infl
Ogt E. coli
Ogt H. infl
Ogt Gram+
Ogt D. radio
M M E
G T uks
AlkA Gram+
AlkAE. coli
AlkA Domain (O6-Me-G glycosylase)
Ogt Domain (O6-Me-G alkyltransferase)
Ada Domain (transcriptions regulator)
TIGR

21. DNA Ligases
• Two major ligase families
• Ligase I
– NAD dependent
– Found in all bacteria and only in bacteria
• Ligase II
– ATP dependent
– Found in all Archaea and eukaryotes
– Found in some bacteria
– Duplicated in many eukaryotes
TIGR

22. DNA Ligases in A. thaliana
ARATH1 F4N21.14
YEAST-GP-600039
YEAST-SW-DNLI YEAST
YEAST-GP-3515
ARATH1 F13F21.31
ARATH1 T23G18.1
ARATH1 T6D22.10
CELEG C29A12.3
DROMECG560
AERPE-gi|5104764.
AQUAE-gi|2983805
DROME-CG17227
ARATH5 MUL3 11
YEAST-SW-DNL4 YEAST
DROMECG12176
ARCFU-gi|2648829
METJA-gi|1590924
METTH-gi|2622703
ARCFU-gi|2649996
TIGR PYRHO-gi|3258051
PYRFUPf 1527421

23. Mismatch Excision Repair
• Core of process highly homologous between bacteria and
eukaryotes (all use MutS and MutL homologs).
• Eukaryotes encode multiple MutS and MutL homologs, not all
of which are involved in mismatch repair.
• Two major MutS groups– MutS-I proteins involved in MMR
and MutS-II proteins involved in chromosome segregation.
• MutS1 and MutL missing from many bacteria, especially
pathogens. Other MMR proteins also defective in some.
• Few homologs in Archaea – some encode MutS2, none encode
MutS1, and some may encode MutL.
• Some evolutionary and functional relationships to restriction-
modification systems (MutH, MED1, Vsr).
TIGR

24. 9
9
0
5 MH
S 6
1 0
0
79 MH
S 3
1 0
0
1 0
0 MH
S 2
M tS
u -I
M ism tch
a
95
1 0
0
MH
S 1 R a
ep ir
2
9
5
6
M tS
u 1
6 /8
1 9
Proposed
duplication
55
1 0
0 MH
S 5
M tS
u -II
8
9
5
6
MH
S 4
C ro o m
h m so e
C sso er &
ro v
S reg tio
eg a n
TIGR
60
74 M tS
u 2

25. Ancient Duplication in MutS Family
A. B.
B ug of r
. r d rei
b
S y gns
po ee
5
Tp li u
. ald m
Bs bls
. ut i
i
Dr do ua s
. ai dr n
Sn s
y. p Mt 2
uS Aaoi u
. e lc s
Aa oc s
. el u
i
M e iai m
.gn l u
t
Dr d d r n
. a i ua s
o 4
3 M nu oi e
.p e mna
Bb r d rei
. ug of r Spo e e
. y gns
Spo e e
. y gns
Bs bii
. u tl s
Bs bls
. ut i
i Gn
ee Sns
y. p
Dpc to
ul a n
i i
Sn s
y. p Hpl r
. yoi
M tS
u 1
Gn
ee 2 Ng n rh e e
. oor oa
Dpc to
ul a n
i i Aa oc s
. el u
i
1 Hi funa
. nl e z e
Dr d d r n
. a i ua s
o
Bb r d rei
. ug of r Ec l
. oi
TIGR

26. Parallel Loss of MutLS
Lost in mycoplasmal lineage (present in B. subtilis and S.
pyogenes)
Lost in M. tuberculosis lineage (found in some other highGC
Gram-positives)
Lost in H. pylori / C. jejuni lineage (present in many other
Proteobacteria)
Possibly lost in Euryarchaeota lineage
Defective in many “wild” E. coli and S. typhimurium strains
Loss of genes may give an advantage in some conditions by
increasing mutation rate or recombination rate between
species.
TIGR

27. Nucleotide Excision Repair
• Bacterial and eukaryotic systems are not-homologous,
despite having very similar mechanisms
• Most of the eukaryotic and bacterial proteins originated
within each of these domains
• Some of the eukaryotic proteins are shared with Archaea
(Rad1, Rad2, Rad25).
• All free-living bacteria encode UvrABCD. B. aphidicolum
encodes Mfd but not UvrABCD.
• UvrABC also found in one Archaea.
• Some functional and evolutionary relationships with drug
resistance and transport
TIGR

28. 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 UvrA1
UvrA M. hermoautotrophicum
UvrA H. pylori
UvrA1 UvrA C. jejuni
UvrA P. gingivalis
UvrA2 Dup lication
UvrA C. tepidum
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
TIGR

29. UvrA Evolution
UvrA1C UvrA1N UvrA2C UvrA2N
Gene Duplication
UvrAC UvrAN
Tandem Duplication
ABC2 ABC1 UvrA
Diversification of ABC family
ABC
TIGR

30. Base Excision Repair Glycosylases
• Distribution patterns highly uneven but some glycosylases
have been found in all species
• Some are ancient enzymes, probably presence in LUCA (e.g.,
MutY-Nth), others more recent (e.g., TagI).
• Many families are distantly related to each other (e.g., Ogg,
AlkA, MutY-Nth)
• Many cases of gene duplication, loss and possibly transfer,
especially from organellar genomes to nucleus
• Orthologs frequently have different specificity
TIGR

31. A. thaliana TAG homologs
C. crescentus
A. thaliana_ 5 K23L20 1
A. thaliana_ 3 MBK21.7
A. thaliana_ 1 F23A5.15
A. thaliana_ 1 T24D18.7
A. thaliana_5 MTI20 23
A. thaliana_1 F9E10.6
V. cholerae
H. influenzae
E.coli
M. tuberculosis
N. meningitidis A
TIGR N. meningitidis B

32. AP Endonucleases
• All species encode either Nfo or Xth homologs. Some encode
both.
• Only Nfo: mycoplasmas, Aquifex, M. jannascii, yeast
• Only Xth: many bacteria, A. fulgidus, humans (so far)
• Both: E. coli, B. subtilis, M. tuberculosis, M.
thermoautotrophicum
• Both Nfo and Xth are likely ancient.
• Many cases of gene loss of one or the other, but never both
TIGR

33. Recombinational Repair
• RecA homologs found in all free-living species (B.
aphidicolum encodes RecBCD but not RecA)
• Most recombination initiation pathways are of recent origin
– RecBCD, RecE within Proteobacteria/Gram-positives
– RecF within bacteria
– AddAB within low-GC gram-Positives
– SbcCD may be of ancient origin (possibly homologous to
MRE11/Rad50)
• Resolution pathways also somewhat recent origin
– CCE1 within eukaryotes
– RuvABC, RecG near origin of bacteria
– Rus within bacteria (phage origin?)
• Many cases of gene loss in initiation, resolution pathways.
TIGR

34. Xen.bov ie
Xen.nemat
Pr.v ulgari
Pr.mirabil
Ent.agglo
Y .pestis
S.marcesce
E.coli
Shig.flex
Shig.sonn
Shepu.tig
V ib.angui
V ib.choler
γ
Ps.oleov or
Ps.margina
Ps.fluores
Ps.putid
Ps.aerugi
Ps.aePA M
A z.v inelan
M BBA D17T F * * * * * *
A c.calcoac
A c.sp.A DP
Past.haem
H.influenz
Past.multo
A ctinobaci
A er.salmon
Xa.ory za
Xa.citri
Xa.campes
B.pertussi
Ps.cepaci
Chrom.v ino
M thmon.cla
M thphy .met
M thbac.fla
β
Nitrosomon
L.pneumop
Ne.gonorr
Ne.meningi
T.ferroox i
R hb.phase
R h.legumin
A .tumefaci
R h.melilot
Br.abortus
Blastochlo
α
R hps.palu
A ceto.pol
A ceto.alt
Gluc.ox y d
A q.magnet
Zy m.mobili
Caul.cresc
Prcs.denit
R ho.sphae
R ho.capsu
2M y x .x anth
1M y x .x anth
δ
He.py lori
TIGR ε
He.py lori2
Cmp.jejuni
Cmp.fetus
0.1

35. A05970
MucB
U muCs ImpB
******
RumB DinP3
RumB R391
RulB
DinP1
DinP2
UvrX
TIGR

36. Big Picture: Evolutionary Origin
TIGR

37. Likely Ancient Repair Processes/Proteins
Process Proteins
Mismatch repair MutL, MutS
AP endonuclease Xth, Nfo
Recombinase RecA/RadA/Rad51
Alkylation reversal Ogt/MGMT
Photolyase PhrII, PhrI
dGTP/GTP clean up MutT
Base excision glycosylases MutY/Nth, AlkA, Ung?
Recombination endonuclease SbcC/Rad50, SbcD/MRE11
Other SMS, Lon, UmuC
TIGR

38. Originated within Bacteria
Process Proteins
Mismatch repair MutH, Vsr
Alkylation reversal Ada (fusion of Ogt)
Base excision Fpg-Nei, TagI
Recombination initiation RecFJNOR, AddAB,
RecBCD, RecET, SbcB
Recombination resolution RecG, RuvABC, Rus
Nucleotide excision repair UvrABCD
Transcription-coupled repair MFD
Induction LexA
Other SSB, LigaseI
TIGR

39. Originated within Eukaryotes
Process Proteins
Mismatch repair duplications of MutS, mutL
Base excision 3MG?
Recombinase duplication of RecA
Recombination initiation duplications of RecQ
Recombination resolution CCE1
Nucleotide excision repair Most XPs, TFIIH, etc.
Transcription-coupled repair CSA, CSB
Induction P53
Non-homologous end joining XRCC4, Kus, DNA-PKcs
Other RFAs, Rad52-59, XRS2
TIGR

40. Originated in Eukaryote-Archaea Lineage
Process Proteins
Base excision Ogg
Nucleotide excision repair Rad1, Rad2, Rad25?
Ligation LigaseII
TIGR

41. Ambiguous Origin
TIGR

42. Repair Genes in Archaea
• All species: RecA,MRE11, Rad50, MutY-
Nth, Ogt, Rad2, Lig-II, PCNA
• UvrABCD in M. thermoautotrophicum
• PhrI and PhrII in some species
• Variety of glycosylases in some species
• No Ung homologs in any species, but
alternative glycosylases have Ung activity
• Rad1 in many species.
• New Holliday junction resolvase
TIGR

43. TIGR

44. 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
TIGR

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

46. Unusual Features of D. radiodurans
DNA Repair Genes
Process Genes
Nucleotide excision repair Two UvrAs
Base excision repair Four MutY-Nths
Recombination RecD but not RecBC
Replication Four Pol genes
dNTP pools Many MutTs, two RRases
Other UVDE
TIGR

47. Gain and Loss of Repair Genes BACTERIA ARCHAEA EUKARYOTES
Human
Mycge
Mycpn
Yeast
Bacsu
Synsp
Borbu
Neigo
Trepa
Metth
Haein
Helpy
Metjn
Arcfu
Strpy
Ecoli
-Ogt -AlkA -Nfo -AlkA
-PhrI -Ogt -AlkA
-AlkA -PhrI -Ogt -Ung -Xth -Rad25
-AlkA -Nfo -RecFRQN -Rad25?
Rus
+ -Nfo -TagI -RecQ +P53
-Vsr -RuvC
UmuD
+ -Nfo -SbcD? dRecQ
-SbcCD -Dut +Rad7
+Nei? -Rec -Lon dRad23
-LexA -SMS +CCE1
+RecE -SbcCD -LexA +MAG?
tRecT? -UmuC -LexA +Spr tTagI ? tRad25
t3MG
-PhrI -PhrII
-PhrI -Ogt -PhrI -Ogg tUvrABCD
Ada
+ -PhrII -AlkA -Ogt
MutH
+ -PhrII?
-AlkA -Xth -AlkA -Ung
SbcB
+ -Fpg -MutLS -Nfo
-Fpg
-Nfo -RecFJORQN -Nfo -Dut
-MutLS -Mfd -RecO -Lon -PhrI
-RecFORQ -SbcCD -LexA -Ung?
-PhrII -SbcCD -RecG -UmuC -MutLS
-LexA -Dut -RecQ?
Vsr
+ -UmuC -PriA -Dut
RecBCD?
+ -TagI+RecT -LexA -UmuC
-SMS
-MutT RFAs
+
-PhrII +TFIIH
-RuvC +Rad4,10,14,16,23,26
CSA
+
Rad52,53,54
+
+TagI? dPhr DNA-PK, Ku
+
SNF2
d
TagI?
+ dMutS
+Fpg dMutL
UvrABCD
+ dRecA
Mfd
+
RecFJNOR
+ Ung?
+
RuvABC
+ SSB,
+
+RecG Rad1
+ +Dut?
LigI
+ +Rad2 from mitochondria
LexA
+ +Rad25?
SSB
+ Ogg
+
+PriA LigII
+
+Dut?
PhrI, PhrII
+
+Ogt
+Ung, AlkA, MutY-Nth
+AlkA
+Xth, Nfo?
+MutLS?
+SbcCD
+RecA
+UmuC
TIGR
+MutT
+Lon
dMutSI/MutSII
dRecA/SMS
dPhrI/PhrII

48. Repair Studies in Different Species
(determined by Medline searches as of 1998)
Humans 7028
E. coli 3926
S. cerevisiae 988
Drosophila 387
B. subtilits 284
S. pombe 116
Xenopus 56
C. elegans 25
A. thaliana 20
Methanogens 16
Haloferax 5
Giardia 0
TIGR

49. Evolution of Repair Summary
• Mycoplasmas have lost many repair genes which may
explain high mutation rate.
• Mismatch repair genes absent in many pathogens (is high
mutation rate advantageous?)
• Whole pathways frequently lost as units (e.g., MutLS).
• May be able to predict pathway interactions by correlated
loss of genes.
• Archaeal genomes have few homologs of bacterial or
eukaryotic repair proteins.
• Some eukaryotic repair proteins have likely mitochondrial
and plastid ancestry
• Many ancient duplications (MutS, SNF2, UvrC).
• Some unusual distributions (XPB, UvrABCD)
TIGR

50. TIGR

51. Acknowledgements
TIGR NIEHS
•Craig Venter •Ben Van Houten
•Claire Fraser
•John Heidelberg Louisiana State University
•Owen White •John Battista
•Steve Salzberg
Other
Stanford •J. Laval
•Phil Hanawalt •F. Taddei
•Rick Myers •A. Britt
•D. Crowley •J. Miller
U.C. Berkeley Funding
•Michael Eisen •DOE, OBER
•A. J. Clark •NIH
•NSF
TIGR

52. TIGR

53. Unusual Distributions
• XP-B like gene in some bacteria and some Archaea.
• LigaseII in M. tuberculosis, B. subtilis, and A. aeolicus
• UvrABCD in M. thermoatuotrophicum
• Mycoplasmas and some low GC gram positives do not have
any Holliday junction resolving homologs (RuvC, RecG,
Rus)
• Mycoplasmas are the only species without MutY-Nth
homologs
• MutS2 unevenly distributed among bacteria, Archaea
• Genes in RecF pathway not always present as a unit
• Uracil glycosylase missing from Archaea and some bacteria
TIGR

54. Big Picture: Duplication and
Loss
TIGR

55. Genes Lost in Mycoplasmal Lineage
Process Protein
Base excision repair MutY/Nth, AlkA
Recombination initiation RecF pathway, SbcCD
Recombination resolution RecG, RuvC
Mismatch repair MutLS
Transcription coupled repair MFD
Induction LexA
Direct repair PhrI, Ogt
AP endonuclease Xth
Other MutT, Dut, PriA, SMS
TIGR

56. Parallel Loss of MutLS
Lost in mycoplasmal lineage (present in B. subtilis and S.
pyogenes)
Lost in M. tuberculosis lineage (found in some other highGC
Gram-positives)
Lost in H. pylori lineage (present in many other Proteobacteria)
Possibly lost in Euryarchaeota lineage
Defective in many “wild” E. coli and S. typhimurium strains
Loss of genes may give an advantage in some conditions by
increasing mutation rate or recombination rate between
species.
TIGR

57. Need for Experimental Studies in Archaea
• No novel repair genes cloned in Archaea. All
repair genes show homology to repair genes in
other species.
• Many novel repair genes found in bacteria and
eukaryotes because of experimental work in
these species.
• Since novel repair pathways appear to evolve
frequently in bacteria and eukaryotes, there is
a need for more genetic and experimental
studies of repair in Archaea.
TIGR

58. Repair Genes in all Archaea
Process Protein
Nucleotide excision repair Rad2, Rad1 ±
Recombination RecA, Mre11, Rad50
Replication PolB, PCNA
Ligase Ligase II
Base excision repair MutY-Nth
dNTP pools MutT family
Alkyltransferase Ogt in all species
TIGR

59. DNA Repair Gene Summary
• Most of the standard eukaryotic DNA repair
genes are found
• Some likely plastid repair genes are found
• Some duplications relative to other species
TIGR

60. Acknowledgements
• Genome duplications: S. Salzberg, J. Heidelberg, O.
White, A. Stoltzfus, J. Peterson
• Genome sequences and analysis: J. Heidelberg, T.
Read, H. Tettelin, K. Nelson, J. Peterson, R.
Fleischmann, D. Bryant
• Horizontal transfers: K. Nelson, W. F. Doolittle
• TIGR: C. Fraser, J. Venter, M-I. Benito, S. Kaul,
Seqcore
• $$$: DOE, NSF, NIH, ONR
TIGR

61. Evolution of Uracil Glycosylase
• Many non-homologous proteins have uracil-DNA
glycosylase activity (Ung, GPADH, MUG, cyclin)
• Therefore, absence of homologs of these genes
should not be used to infer likely absence of
activity
• However, presence of homologs of Ung and MUG
genes can be used to indicate presence of activity
because all homologs of these genes have this
activity
TIGR

62. Ambiguous Origin
Process Proteins
Base excision 3MG, GT MMR, Ung
Nucleotide excision repair Rad25
Recombination initiation RecQ
Other Dut
TIGR

63. Big Picture: Distribution Patterns
TIGR

64. Present in All Bacteria
Process Proteins
Nucleotide Excision Repair
Recombinase
Replication PolA,C
Single-strand DNA Binding SSB
Ligase LigaseI
TIGR

65. Present in All Free-Living Bacteria
Process Proteins
Nucleotide Excision Repair UvrABCD
Recombinase RecA
Replication PolA,C
Single-strand DNA Binding SSB
Ligase LigaseI
TIGR

66. Present in Most Bacteria
Process Protein
Nucleotide excision repair UvrABCD
Holliday junction resolution RuvABC
Recombination RecA; RecJ, RecG
Replication PolA,C; PriA; SSB
Ligase DnlJ
Transcription-coupled repair Mfd
Base excision repair Ung, MutY-Nth
AP endonuclease Xth
TIGR

67. Present in Bacteria or Eukaryotes
(But Not Both)
Process Bacteria Eukaryotes
Transcription-coupled repair CSB, CSA
Mismatch strand recognition MutH -
Nucleotide excision repair UvrABC XPs, TFIIH, etc.
Recombination initiation RecBCD, RecF KU, DNA-PK
Holliday junction resolution RuvABC CCE1
Base excision -
Inducible responses LexA P53
TIGR

68. Evolution of Alkyltransferases
• All known alkyltransferases share a conserved,
homologous alkyltransferase domain
• Therefore, if a species does not encode any protein
with this domain, it likely does not have
alkyltransferase activity
• If a species does encode an member of this gene
family, it likely has alkyltransferase activity
TIGR

69. Standard Eukaryotic Repair Genes
Pathway Genes
Mismatch Repair MSH2-6, MutLs
Base Excision Repair Ogg, MutY-Nth, Tag,
3MG, Ung
Nucleotide Excision XPA, Rad1, Rad2, Rad3,
Repair Rad10, Rad25, etc
Recombination MRE11, Rad50
Rad51
Direct Repair Phr, Dnl1
Other PCNA, Dut, Lon
TIGR

70. Missing Eukaryotic Repair Genes?
Pathway Genes
Mismatch Repair
Base Excision Repair
Nucleotide Excision XPA, Btf2, Btf3,
Repair Kin28
Recombination
Direct Repair Ogt
Other
TIGR

71. Bootstrap
MSH6.P ombe
55 MSH6.Yeast
35
30
38 100 GTBP .Arath IV.At4g02070
ARATH T10M13.8
93 93
ARATH AGAA.3 MSH6
MSH7.Arath
100 GTBP .Mouse
GTBP .Human
27 Y47G6A.11.Celegans
100 MSH3.Human
82
38 REP 1.Mouse
MSH3.Arath IV M7J2.90 MSH3
SWI4.pombe
MSH3.yeast
MUTS BORBU
MUTS TREP A
48 MutS.Cloac.blast
MutS.Clodi.blast
15 30 MutS.Synsp
26 MutS.Chlte.blast
96 MutS.P orgi.blast
100 100 MutS.Theaq
35 MutS.Theaq cald
MutS.Theth
MutS.Thema
66 100 MutS.Ecoli
12 60 MutS.Salty
82 MutS.Yerpe.blast
20 33 82 MutS.Vibch
100 97 MutS.Haein
83
43 100 MutS.Actin.blast
66
33 17 MutS.Actin.blast MutS1
100 MutS.P asmu.blast
72 MutS.Shepu.blast
31 61 MutS.Neime.TIGR
54 12 MutS.Neigo.blast
86 15 100 MutS.Azovi
MutS.P seae.blast
MutS.Thife.blast
24 100 MutS.Entfa.blast
98 77 MutS.Strmu.blast
97 MutS.Strpy.blast
78 HexA.Strpn
54 MutS.Staau.blast
MutS.Bacsu
100MutS.Ricpr
MUTS RHIME
89 MutS.Caucr.prelim
92 100 MutS.Aqupy
89 100MutS.Aquae
80 MutS.Chltr.?
MutS.Chlpn
MSH2.Rat
100 79 MSH2.Human
93
67 38 MSH2.Mouse
40
72
88 MSH2.Xenla
85 100 MSH2.Neucr
MSH2.Yeast
MSH2.Arath3
Mus1.Maize MSH2
MSH2.P ombe
SP E1.Drome
H26D21.2.Celeg
88 MSH1.Spombe MSH1
MSH1.Yeast
MSH1.Canal.blast
51 76 MSH4.Celeg
MSH4.Yeast
MSH4.Canal MSH4
MSH4.human
46 ARATH3 MQC12.24
MSH5.Yeast
100 MSH5.Human
53
Msh5.Mouse MSH5
MSH5.Celeg
mMutS.Saco.glaucum.
Muts2.Metth
100 100 MutS2.P yrho
100 MutS2.P yrab
100 MutS2.Helpy.TIGR
MutS2.Helpy99
MutS2.Camje
55 MutS2.Deira.TIGR
MutS2.Thema
58 MutS2.Bacsu
MutS2.Staau
17 45
23 77 Muts2.Entfa.blast
100 Muts2.Strmu.blast MutS2
49 27 87 39 MutS2.Strpy.Blast
50 917 MutS2.Strpn.blast
95 Muts2.Cloac.blast
26 Muts2.Clodi.blast
70 MutS2.Synsp
ARATH 5 MJP 23 7
Muts2.Chlte.blast
TIGR
MutS2.Borbu
MutS2.Aquae
Muts2.P orgi.blast
MutS2.Arath 1 F16G16.7

72. Bootstrap
81 E.coli
Shig.flexneri
Shig.sonnei
100
55
Ent.agglomerans
Y.pestis
61 87
45 S.marcescens
71
Xen.bovienii.edit
10 55
0
Xen.nematophilus
87 100
P r.vulgaris
γ
43 96 P r.mirabilis
Shepu.tigr
49
100 Vib.anguillarum
23 Vib.cholerae
66
%MP LA
Aer.salmonicida
Actinobacillus actinomycetemco
59 100 83 H.influenzae
75
1 00 P ast.multocida
P ast.haemolytica
25 P s.oleovorans
25 100
100 P s.marginalis
P s.fluorescens
93
P s.putida
96 P s.aeruginosa
100
P s.aeP AM7
46
7 Az.vinelandii
Ac.calcoaceticus
Ac.sp.ADP 1
100 Mthmon.clara
99 90 Mthphy.methylotrophus
65
Mthbac.flagellatum
β
100 30
30 Nitrosomonas.blast
8
35 Chrom.vinosum
L.pneumophila
100 T.ferrooxidans
10
67
Xa.oryzae
Xa.campestris
Xa.citri
25
B.pertussis
40 P s.cepacia
Ne.gonorrhoea
Ne.meningitis.TIGR
4 Blastochloris.viridis
Rhps.palustris
85
85
Rhb.phaseoli
78 Rh.leguminosarum
96 99 8 7
A.tumefaciens
54 Rh.meliloti
18 Br.abortus
α
44
94
3 36 Aq.magnetotacticum.edit
49
100 9 3 Aceto.polyoxogenes
Aceto.altoacetigenes
98
Gluc.oxydans
17 Zym.mobilis
84 P rcs.denitrificans
4 Rho.sphaeroides
Rho.capsulatus
Caul.crescentus
ε
99 Ric.prowazekii
18 reca.wolbachia.blast
He.pylori
10 0100
He.pylori2
56 82
Cmp.jejuni
δ
Cmp.fetus
100
32 1Myx.xanthus
10
95 8 0
2Myx.xanthus
blast.geobacter
blast.desulf
Tmtg.maritima
Spirochetes
93 Trep.pallidum
9944 Tr.denticolum.blast
Bor.borgdorferi
96
Lept.biflexa
Lept.interrogans
Cyanobacteria
39
Spir.platensis
68 Syn.7942
97 Ana.variabilis
9 94
Syn.7002
Syn.6803
A.thaliana
A.thaliana.chr3
Mycoplasmas
88 A.thaliana.chr2
51 100Ureo urolyticum
39 23 100Mycp.genitalium
21 Mycp.pneumoniae
68
P hytoplasma sp
Mycp.mycoides
97 Mycp.pulmonas
100
De.radiodurans
The.aquaticus
D/T
Green Non-Sulfur
The.thermophilus
30
Chlfx.aurantiacus
60
Dehalo.blast
Chlamydia
100
Chl.trachomatis
Chl.pneumoniae
Cory.glutamicum
48
Cory.pseudotuberculosis
99 Strpm.coelicolor
100 52 Strpm.lividans
93
Strpm.ambofaciens
High GC Gram +
96 100 Strpm.violaceus
5 2 57 Strpm.rimosus
100 Mycb.tuberculosis
9956 Mycb.bovis.sanger.fr2
36
99 Mycb.avium.blast
Mycb.leprae
Mycb.smegmatis
Amycolatopsis mediterranei
Bifido.breve
Green Sulfur
Chb.tepidum
100P orp.gingivalis
78Bact.fragilis
P rev.ruminocola
List.monocytogenes
64 60B.subtilis
73
Bac anthracis.blast
76 Ent faecalis.blast
Low GC Gram +
70 Staph.aureus
77 100 Strc.pneumoniae
88
100 Strc.parasanguis
TIGR
Strc.pyogenes
84 1Lct.lactis
Ach.laidlawii
reca.blast.carboxy
100Clost.perfringens
Cl.acetybutylicum.blast
Hydrogenobacteria
Aq.pyrophilus
Aq.aeolicus