Transcript Evolution of regulatory interactions in bacteria

Life
without
Fur
Life without FUR: evolutionary
reconstruction of transcriptional
regulation of iron homeostasis in
alpha-proteobacteria
Mikhail Gelfand
Research and Training Center of Bioinformatics,
Institute for Information Transmission Problems, RAS
Genome Dynamics: From Replication to Post-Translation and Turnover
HHMI, 11-14 March 2007
Regulation of iron homeostasis
(the Escherichia coli paradigm)
Iron:
• essential cofactor (limiting in many environments)
• dangerous at large concentrations
FUR (responds to iron):
• synthesis of siderophores
• transport (siderophores, heme, Fe2+, Fe3+)
• storage
• iron-dependent enzymes
• synthesis of heme
• synthesis of Fe-S clusters
Similar in Bacillus subtilis
Regulation of iron homeostasis in α-proteobacteria
[- Fe]
[+Fe]
[ - Fe]
[+Fe]
RirA
RirA
Irr
Irr
FeS
heme
degraded
Siderophore
uptake
2+
3+
Fe / Fe
uptake
Iron uptakesystems
Fur
[- Fe]
Iron storage
ferritins
FeS
synthesis
Heme
synthesis
Iron-requiring
enzymes
[ironcofactor]
Fur
IscR
Fe
FeS
Transcription
factors
FeS status
of cell
[+Fe]
Experimental studies:
• FUR/MUR: Bradyrhizobium, Rhizobium and Sinorhizobium
• RirA (Rrf2 family): Rhizobium and Sinorhizobium
• Irr (FUR family): Bradyrhizobium, Rhizobium and Brucella
Comparative genomics of regulatory systems
• Standard methods:
–
–
–
–
BLAST
Construction of phylogenetic trees to identify orthologs
Functional annotation by similarity
Co-localization patterns
• Analysis of regulation:
– Phylogenetic footprinting
(Conserved motifs upstream of orthologs)
– Consistency filtering
(true sites upstream of orthologs;
false positives scattered at random)
Distribution of
transcription
factors in
genomes
FUR/MUR branch of the FUR family
Fur
sp|
Escherichia coli: P0A9A9
ECOLI
Pseudomonas aeruginosa : sp|Q03456
PSEAE
NEIMA
Fur in g- and b- proteobacteria
Neisseria meningitidis : sp|P0A0S7
HELPY Helicobacter pylori : sp|O25671
P54574
BACSU Bacillus subtilis : sp|
SM mur
Sinorhizobium meliloti
Mesorhizobium sp. BNC1 (I)
MBNC03003179
BQ fur2
Bartonella quintana
BMEI0375
Brucella melitensis
EE36 12413 Sulfitobacter sp. EE-36
MBNC03003593Mesorhizobium sp. BNC1 (II)
Rhodobacterales bacterium HTCC2654
RB2654 19538
Agrobacterium tumefaciens
AGR C 620
RHE_CH00378 Rhizobium etli
Rhizobium leguminosarum
RL mur
Nham 0990 Nitrobacter hamburgensis X14
Nwi 0013
Nitrobacter winogradskyi
Rhodopseudomonas palustris
RPA0450
Bradyrhizobium japonicum
BJ fur
Roseovarius sp.217
ROS217 18337
Jannaschia sp. CC51
Jann 1799
Silicibacter pomeroyi
SPO2477
STM1w01000993Silicibacter sp. TM1040
MED193 22541 Roseobacter sp. MED193
OB2597 02997Oceanicola batsensisHTCC2597
Loktanella vestfoldensisSKA53
SKA53 03101
Rhodobacter sphaeroides
Rsph03000505
Roseovarius nubinhibensISM
ISM 15430
PU1002 04436Pelagibacter ubiqueHTCC1002
GOX0771 Gluconobacter oxydans
Zmomonas
y
mobilis
ZM01411
Saro02001148 Novosphingobium aromaticivorans
Sphinopyxis alaskensis RB2256
Sala 1452
ELI1325
Erythrobacter litoralis
Oceanicaulis alexandrii HTCC2633
OA2633 10204
PB2503 04877 Parvularcula bermudensis HTCC2503
CC0057
Caulobacter crescentus
Rhodospirillum rubrum
Rrub02001143
Magnetospirillum magneticum (I)
Amb1009
Magnetospirillum magneticum(II)
Amb4460
Fur in e- proteobacteria
Fur in Firmicutes
Mur
in a-proteobacteria
Regulator of manganese
uptake genes (sit, mntH)
Fur
in a-proteobacteria
Regulator of iron uptake
and metabolism genes
Irr
a-proteobacteria
Erythrobacter litoralis
Caulobacter crescentus
Novosphingobium aromaticivorans
Zymomonas mobilis
Oceanicaulis alexandrii
Sphinopyxis alaskensis
Gluconobacter oxydans
Rhodospirillum rubrum
Parvularcula bermudensis -
Magnetospirillum magneticum
Identified Mur-binding sites
of a - proteobacteria
-
FUR
and
MUR
boxes
Bacillus subtilis
Mur
Escherichia coli
Sequence logos for
the known
Fur-binding sites
in Escherichia coli
and Bacillus subtilis
Irr branch of the FUR family
Fur
Escherichia coli : P0A9A9
sp|
ECOLI
Pseudomonas aeruginosa : sp|Q03456
PSEAE
NEIMA
Fur in g- and b- proteobacteria
Neisseria meningitidis : sp|P0A0S7
HELPY Helicobacter pylori : sp|O25671
sp|
BACSU Bacillus subtilis : P54574
Fur in e- proteobacteria
Fur in Firmicutes
a-proteobacteria
Mur / Fur
Agrobacterium tumefaciens
AGR C 249
Sinorhizobium meliloti
SM irr
Rhizobium etli
RHE CH00106
Rhizobium leguminosarum (I)
RL irr1
RL irr2 Rhizobium leguminosarum (II)
Mesorhizobium loti
MLr5570
MBNC03003186 Mesorhizobium sp. BNC1
BQ fur1 Bartonella quintana
Brucella melitensis (I)
BMEI1955
Brucella melitensis (II)
BMEI1563
BJ blr1216 Bradyrhizobium japonicum (II)
RB2654 182 Rhodobacterales bacterium HTCC2654
Loktanella vestfoldensis SKA53
SKA53 01126
Roseovarius sp.217
ROS217 15500
Roseovarius nubinhibens ISM
ISM 00785
OB2597 14726 Oceanicola batsensis HTCC2597
Jann 1652 Jannaschia sp. CC51
Rsph03001693Rhodobacter sphaeroides
Sulfitobacter sp. EE-36
EE36 03493
STM1w01001534 Silicibacter sp. TM1040
Roseobacter sp. MED193
MED193 17849
SPOA0445
Silicibacter pomeroyi
Rhodobacter capsulatus
RC irr
RPA2339
Rhodopseudomonas palustris (I)
RPA0424*
Rhodopseudomonas palustris (II)
Bradyrhizobium japonicum (I)
BJ irr*
Nwi 0035* Nitrobacter winogradskyi
Nham 1013* Nitrobacter hamburgensis X14
PU1002 04361
Pelagibacter ubique HTCC1002
Irr in a-proteobacteria
regulator of iron
homeostasis
Irr boxes
Rhizobiaceae plus
Bradyrhizobiaceae
Rhodobacteriaceae
Rhodospirillales
RirA/NsrR family (Rhizobiales)
IscR family
Summary: regulation of
genes in functional
subsystems
Rhizobiales
Bradyrhizobiaceae
Rhodobacteriales
The Zoo (likely
ancestral state)
Reconstruction of history
Frequent
co-regulation
with Irr
Strict division
of function
with Irr
Appearance of the
iron-Rhodo motif
Experimental validation
• RirA: sites and binding motif
in Rhisobium legumisaurum
(site-directed mutagenesis).
Andy Johnston lab (University of East Anglia)
• Microarray study if the Bradyrhizobium
japonicum FUR– mutant:
regulatory cascade FUR  irr:
Mark O’Brian group (SUNY, Buffalo)
All logos and
Some Very Tempting
Hypotheses:
• Cross-recognition of FUR
and IscR motifs in the
ancestor.
• When FUR had become
MUR, and IscR had been
lost in Rhizobiales,
emerging RirA (from the
Rrf2 family, with a rather
different general
consensus) took over their
sites.
• Iron-Rhodo boxes are
recognized by IscR:
directly testable
More stories
• Regulation of methionine metabolism in Firmicutes
(from S-boxes to T-boxes and transcriptional
factors)
• T-box regulon in Firmicutes
(duplications, bursts, changes of specificity)
• Regulation of respiration in gamma-proteobacteria
(rewiring of regulatory cascades and shuffling of
regulons)
• Emerging global regulators in Enterobacteriaceae
(how FruR has become CRA, and how duplicated
RbsR has become PurR)
Acknowledgements
• Dmitry Rodionov (IITP,
now at Burnham Institute, La Jolla, CA)
• Andrew Johnston and
Jonathan Todd
(University of East Anglia, UK)
• Howard Hughes Medical Institute
• Russian Academy of Sciences
program “Molecular and Cellular Biology”