Transcript Comparative genomics and metabolic reconstruction of

Evolution
of bacterial regulatory systems
Mikhail Gelfand
Institute for Information Transmission Problems, RAS
BGRS-2004, Novosibirsk
Early analyses (BGRS’98, 00, 02)
“Making good predictions with bad rules”
Basic assumption: regulons are conserved
=>
Consistency check: sites upstream of orthologous
genes are correct; false positives are scattered at
random
• Validation of individual sites
• Validation of signals: candidate signals for
orthologous factors are correct if similar
Multiple genomes:
taxon-specific regulation;
multiple interacting systems;
evolution of regulation
• Evolution of orthologous regulatory sites
• Co-evolution of transcription factors
and their binding signals
• Evolution of regulons
(sets of co-regulated genes)
• Evolution of regulatory systems
Это – ряд наблюдений. В углу – тепло.
Взгляд оставляет на вещи след.
Вода представляет собой стекло.
Человек страшней, чем его скелет.
Иосиф Бродский
A list of some observations. In a corner, it’s warm.
A glance leaves an imprint on anything it’s dwelt on.
Water is glass’s most public form.
Man is more frightening than its skeleton.
Joseph Brodsky
Conservation of non-consensus
positions in orthologous sites
regulatory site LexA  lexA
consensus nucleotides are in caps
Escherichia coli
Salmonella typhi
Yersinia pestis
Haemophilus influenzae
Pasteurella multocida
Vibrio cholerae
TgCTGTATATActcACAGcA
aACTGTATATActcACAGcA
agCTGTATATActcACAGcA
atCTGTATAcAatacCAGTt
TtCTGTATATAataACAGTt
cACTGgATATActcACAGTc
wrong consensus?
PurR  purL
Escherichia coli
Salmonella typhi
Yersinia pestis
Haemophilus influenzae
Pasteurella multocida
Vibrio cholerae
A C G C A A A C Gg T T t C G T
A C G C A A A C Gg T T t C G T
A C G C A A A C Gg T T t C G T
A t G C A A A C G T T T G Ct T
A C G C A A A C G T T Tt C G T
A C G C A A A C Gg T T G C t T
PurR  purM
Escherichia coli
Salmonella typhi
Yersinia pestis
Haemophilus influenzae
Pasteurella multocida
Vibrio cholerae
t C G C A A A C G T T T G Ct T
t C G C A A A C G T T T G Ct T
t C G C A A A C G T T T G Cc T
t C G C A A A C G T T T G Ct T
t C G C A A A C G T T T G Ct T
A C G C A A A C G T T Tt C c T
Non-consensus positions are more conserved
than synonymous codon positions
1,2
Conservation
1
0,8
LexA non-cons.
Syn.2-fold
0,6
Syn. 3-fold
Syn. 4-fold
0,4
0,2
0
0
0,1
0,2
0,3
0,4
0,5
Distance between genomes
0,6
Non-consensus positions may be more
conserved than consensus positions
Relative conservation
1,6
1,4
1,2
1
Non-consensus
0,8
Consensus
0,6
0,4
0,2
0
1 2
3 4 5
6 7
8 9 10 11 12 13 14 15 16 17 18 19 20
Signal position (LexA)
Regulators and their signals
• Subtle changes at close evolutionary
distances
• Changes in spacing / geometry of dimers
• Correlation between contacting
nucleotides and amino acid residues
• Cases of conservation at surprisingly
large distances
Zinc repressors
nZUR-
GATATGTTATAACATATC
nZUR-
GAAATGTTATANTATAACATTTC
GTAATGTAATAACATTAC
TTAACYRGTTAA
pZUR
TAAATCGTAATNATTACGATTTA
AdcR
Alignment of nZUR binding signals
GTAATGTAA
TAACATTAC (alpha – most genera)
GATATGTTA
TAACATATC (alpha – Rhodobacter)
GAAATGTTATANTATAACATTTC (gamma)
GaaATGTtA-----TAACATttC (consensus of consensi)
CRP/FNR family of regulators
TGTCGGCnnGCCGACA
CooA
Desulfovibrio
TTGTGAnnnnnnTCACAA
FNR
Gamma
TTGATnnnnATCAA
HcpR
Desulfovibrio
TTGTgAnnnnnnTcACAA
Correlation between contacting
nucleotides and amino acid residues
•
•
•
•
DD
DV
EC
YP
VC
DD
DV
EC
YP
VC
CooA in Desulfovibrio spp.
CRP in Gamma-proteobacteria
HcpR in Desulfovibrio spp.
FNR in Gamma-proteobacteria
COOA
COOA
CRP
CRP
CRP
HCPR
HCPR
FNR
FNR
FNR
ALTTEQLSLHMGATRQTVSTLLNNLVR
ELTMEQLAGLVGTTRQTASTLLNDMIR
KITRQEIGQIVGCSRETVGRILKMLED
KXTRQEIGQIVGCSRETVGRILKMLED
KITRQEIGQIVGCSRETVGRILKMLEE
DVSKSLLAGVLGTARETLSRALAKLVE
DVTKGLLAGLLGTARETLSRCLSRMVE
TMTRGDIGNYLGLTVETISRLLGRFQK
TMTRGDIGNYLGLTVETISRLLGRFQK
TMTRGDIGNYLGLTVETISRLLGRFQK
Contacting residues: REnnnR
TG: 1st arginine
GA: glutamate and 2nd arginine
TGTCGGCnnGCCGACA
TTGTGAnnnnnnTCACAA
TTGTgAnnnnnnTcACAA
TTGATnnnnATCAA
The correlation holds for other factors
in the family
Factor
CRP
VFR
CLP
FNR & ANR
FNR
FNR & FixK
DNR & Nnr
FNR
PrfA
NtcA
CysR
CooA
HcpR*
HcpR*
HcpR*
HcpR*
HcpR*
HcpR*
HcpR*
ArcR
CprK
FlpA&B
Organisms
Consensus
Specific aa
Enterobacteria&Vibrio&Pasteurellaceae
TTGTGAnnnnnnTCACAA
R
E
R
Pseudomonas sp.
TTGTGAnnnnnnTCACAA
R
E
R
Xanthomonas&Xylella sp.
nTGTGAnnnnnnTCACAn
R
E
R
Gamma-proteobacteria
nnTTGATnnnnATCAAnn
V
E
R
Beta-proteobacteria
nnTTGATnnnnATCAAnn
L
E
R
Alpha-proteobacteria
nnTTGATnnnnATCAAnn
I/L E
R
Pseudomonas &Paracoccus
nnTTGATnnnnATCAAnn
P
E
R
Bacillus sp.
nTGTGAnnTAnnTCACAn
R
E
R
Listeria
nnTTAACAnnTGTTAAnn
S
S
R
Cyanobacteria
ntGTAnCnnnnGnTACan
R
V
R
Cyanobacteria
?
R
V
R
Desulfovibrio sp. and R.rubrum
nTGTCGGCnnGCCGACAn
R
Q
T
Desulfovibrio sp.
TTGTgAnnnnnnTcACAA
R
E
R
Desulfuromonas acetoxidans, Desulfotalea
atTTGAccnnggTCAAat
psychrophila
S/P E
R
Clostridia, Bacteroides, Thermotogales,
ctGTAACawwtCTTACag
Fusobacteria, Treponema
R
P
R
~P. gingivalis
nTGTCGCnnnnGCGACAn
R
A
R
~C. difficile
nnGGATnnnnnnATCCnn
R
S
R
~T.tengcongensis, D.halfniensa nTGTGAnnnnnnTCACAn
R
E
R
~Acidithiobacillus ferrooxidans
nCTTGATTnnAATCAAGn
P
E
R
Bacillus, Enterococcus sp.
nTGTGAnATATnTCACAn
R
E
A/S
Desulfitobacterium dehalogenas nnTTAnTGnnCAnTAAnn
H
V
R/K
Lactococcus lactis
nnTTGATnnnnATCAAnn
P
E
R
Metabolic system
catabolic repression
virulence
phytopathogenicity
response to anaerobiosis
response to anaerobiosis
nitrogen fixation
denitrification
response to anaerobiosis
virulence
nitrogen metabolism
sulfate utilization
CO utilization
prismane & sulfate reduction
prismane
prismane
prismane
prismane
prismane
prismane
arginine catabolism
halorespiration
?
Inducer
cAMP
cAMP
? (not cAMP)
O2,NO
O2
O2
NO, NO2
O2-low conditions
?
2-oxoglutarate
sulfate?
CO
?
?
?
?
?
?
?
O2
aromatics
Eh, O2
SMc04260 11
The LacI
family of
transcriptional
regulators
RAFR_ECOLI 12
RRC03428
13
10 SMb21598
SMb20324
9 BS_YvdE
8 MALR_STAXY
SMc03060
14
EC_MalI
7 SACR_LACLA
BS_RbsR
16
SMc04401
REF00345
6
SMb21272 15
RKP03067
SMc02975 17
REF00754
mlr2242 18
SCRR_SALTY
BS_CcpA
19
20
EC_FruR
21
5 TTE0201
(each branch
represents a
subfamily)
RKP05215
4 PA1949
22
SMb21650
3 EC_PurR
GALR_STRTR
23
EC_EbgR
2 EC_RbsR
1 EC_LacI
43 EC_YcjW
EC_TreR
24
VCA0654
25
SCRR_STAXY 26
42
41
RPU04121
SMb21372
STM3696
29
40
27
STM2345
RKP05499
36
39 PA2320
RSc1790
34
38 RRC03254
EC_IdnR
EC_GntR
EC_AscG
30
STM1555
31
EC_CytR
EC_GalR
EC_GalS
32
37
CSCR_ECOLI
33
35 SMc03165
XCC2369
PA2259
BS_KdgR
28
D-galactose & galactosides
maltose & trehalose
sucrose
D-fructose
D-ribose
D-xylose
… and their signals
BirA: regulator of biotin biosynthesis and
transport in eubacteria and archaea
Profile 1: Gram-positive bacteria, Archaea
Profile 2: Gram-negative bacteria
Evolution of regulons
and regulatory systems
• conserved cores
• taxon-specific marginal members
• migration of genes between interacting
regulatory systems
• taxon-specific cross-regulation
• genome-specific operons and genomic loci
• complete change of regulatory
mechanisms
Genome loci for hyaluronate utilization
in invasive Streptococcus spp.
S. pyogenes, S. agalactiae
S. equi
S. pneumoniae TIGR4
IS
S. pneumoniae R6
IS
S. suis
IS
Respiration in gamma-proteobacteria
1. Three regulators, different regulatory cascades
Escherichia coli
(experimental data)
Haemophilus influenzae,
Pasteurella multocida,
A. actinomycetemcomitans
Haemophilus ducreyi,
Vibrio spp.
Respiration in gamma-proteobacteria
2. New genome/taxon-specific regulon members
Escherichia coli (known)
Yersinia pestis
Yersinia entercolitica
Pasteurella multocida
Actinobacillus actinomycetemcomitans
Haemophilus influenzae
Haemophilus ducreyi
Vibrio vulnificus
Vibrio parahaemolyticus
Vibrio cholerae
Vibrio fischeri
New, non-homologous regulon member
Fnr
Fnr
Fnr
—
Fnr
Fnr
—
—
Fnr
—
ArcA
—
ArcA
ArcA
ArcA
ArcA
ArcA
ArcA
ArcA
ArcA
—
—
NarP
NarP
—
NarP
—
—
—
—
Respiration in gamma-proteobacteria
3. New genome/taxon-specific regulon members, cont’d
Synthesis of molybdate cofactor
Yersinia pestis
Yersinia entercolitica
Pasteurella multocida
Actinobacillus actinomycetemcomitans
Haemophilus influenzae
Haemophilus ducreyi
Vibrio vulnificus
Vibrio parahaemolyticus
Vibrio cholerae
Vibrio fischeri
Fnr
Fnr
Fnr
Fnr
Fnr
Fnr
—
—
—
—
—
ArcA
ArcA
—
—
ArcA
—
—
—
ArcA
—
—
—
NarP
NarP
NarP
NarP
NarP
NarP
NarP
Zinc repressors - recapitulation
nZUR-
nZUR-
GATATGTTATAACATATC
GAAATGTTATANTATAACATTTC
GTAATGTAATAACATTAC
TTAACYRGTTAA
pZUR
TAAATCGTAATNATTACGATTTA
AdcR
Five
regulatory
systems for
methionine
biosynthesis
A.
SAMdependent
RNA
riboswitch
B. Met-tRNAdependent
T-box (RNA)
C,D,E.
repressors of
transcription
Three methionine regulatory systems in Grampositive bacteria: loss of S-box regulons
• S-boxes (riboswitch)
– Bacillales
– Clostridiales
– the Zoo:
•
•
•
•
•
•
ZOO
Petrotoga
actinobacteria (Streptomyces, Thermobifida)
Chlorobium, Chloroflexus, Cytophaga
Fusobacterium
Deinococcus
proteobacteria (Xanthomonas, Geobacter)
• Met-T-boxes (Met-tRNA-dependent attenuator)
– Lactobacillales
• MET-boxes (candidate transcription signal)
– Streptococcales
Lact.
Strep. Bac. Clostr.
Catabolism of gluconate in proteobacteria
Three regulatory systems
one global (FruR), two taxon-specific (GntR, PtxS)
β
Pseudomonas spp.
γ1
Instead of conclusions…
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Andrei A. Mironov (BGRS’98,00,02,04)
Anna Gerasimova (BGRS’02,04)
Olga Kalinina (BGRS’02,04)
Alexei Kazakov (BGRS’02,04)
Ekaterina Kotelnikova (BGRS’02,04)
Galina Kovaleva (BGRS’04)
Pavel Novichkov (BGRS’00,02,04)
Olga Laikova (BGRS’02,04)
Ekaterina Panina (BGRS’00)
(now at UCLA, USA)
Elizabeth Permina (BGRS’02,04)
Dmitry Ravcheev (BGRS’02,04)
Alexandra B. Rakhmaninova (BGRS’00)
Dmitry Rodionov (BGRS’00)
Alexey Vitreschak (BGRS’00,04)
(visiting LORIA, France)
• Howard Hughes
Medical Institute
• Ludwig Institute of
Cancer Research
• Russian Fund of Basic
Research
• Programs “Origin and
Evolution of the
Biosphere” and
“Molecular and
Cellular Biology”,
Russian Academy of
Sciences