Signal integration in bacterial two-component regulatory systems PRESENTED BY: SEPIDEH BENVARI SUPERVISED BY: DR. ASLANIMEHR Qazvin university of medical science Journal Club & MSc Seminar
Signal integration in bacterial
two-component regulatory
systems
PRESENTED BY: SEPIDEH BENVARI
SUPERVISED BY: DR. ASLANIMEHR
Qazvin university of medical scienceJournal Club & MSc Seminar
Free-living organisms modulate their gene expression patterns in response to environmental cues.
This modulation requires sensors to detect chemical and/or physical signals, and regulators to
bring about changes in the levels of gene products.
Adaptability is a crucial characteristic of bacteria that are able to prosper in a wide variety of
environmental conditions.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
Regulation of Gene
Expression in bacteria
1. Gene copy number
2. Transcriptional control
2.1. Promoters
2.2. Terminators, attenuators and anti-terminators
2.3. Induction and repression: regulatory proteins
2.4. Two-component regulatory systems
2.5. Global regulatory systems
2.6. Quorum sensing
3. Translational control
3.1. Ribosome binding
3.2. Codon usage
3.3. Stringent response
3.4. Regulatory RNA
4. Phase variation
produced by: sepideh benvari supervised by: Dr. Aslanimehr
In bacteria, extracellular signals are transduced into the cell predominantly by two-
component systems.
Although two-component signaling systems are found in all domains of life, they are most
common by far in bacteria, particularly in Gram-negative and cyanobacteria.
They are much less common in archaea and eukaryotes; although they do appear in yeasts,
filamentous fungi, and slime molds, and are common in plants.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
The number of two-component systems present in a bacterial genome is highly
correlated with genome size as well as ecological niche; bacteria that occupy niches with
frequent environmental fluctuations possess more histidine kinases and response
regulators.
New two-component systems may arise by gene duplication or by lateral gene transfer,
and the relative rates of each process vary dramatically across bacterial species. In
most cases, response regulator genes are located in the same operon as their cognate
histidine kinase.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
The average number of two-component systems in a bacterial genome has been estimated as
around 30, or about 1-2% of a prokaryote's genome. A few bacteria have none at all -
typically endosymbionts and pathogens - and others contain over 200.
Two-component signal transduction systems enable bacteria to respond to a wide variety of
stimuli, including nutrients, cellular redox state, changes in osmolarity, quorum signals,
antibiotics, temperature, chemoattractants, pH and more.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
two-component systems
In general, these systems comprise an integral membrane protein called a histidine
protein kinase (HPK) and a separate cytoplasmic protein called a response regulator
(RR).
The HPK has two domains. The input domain is usually found on the outside of the
cell, in an ideal position to detect environmental signals. In contrast, the transmitter
domain is located on the cytoplasmic face of the cell membrane, positioned to interact
with the RR.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
When a stimulus causes a conformational change in HPK,
the HPK autophosphorylates at a conserved histidine
residue and subsequently transfers this phospho- group
to the response regulator. In this form the RR is able to
bind to DNA to regulate transcription of the target genes.
The RR also consists of two domains: a receiver domain
containing an aspartate residue which accepts the
phospho- group, and an output domain which can bind to
DNA.
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Phosphorelays
Phosphorelays are a more complex version of the TCS in which a sensor kinase first
transfers the phosphoryl group to a response regulator possessing the domain with
the conserved aspartate but no output domain.
The response regulator subsequently transfers the phosphoryl group to a histidine-
containing phosphotransfer protein, and it is the latter protein that serves as a
phosphodonor to the terminal response regulator, which possesses an output domain
mediating a cellular response.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
The vast majority of response regulators are active
only when phosphorylated. Therefore, any condition
or product that affects the phosphorylated state of a
response regulator will impact its ability to exert its
biological functions.
Consequently, the output of a response regulator is
determined not only by the presence of the specific
signals sensed by its cognate sensor kinase but also
by gene products that stimulate or inhibit its
phosphorylation.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
REGULATORY MECHANISMS
The sole purpose of two-component signal transduction systems is to allow for
regulation; the signaling pathway merely provides steps at which the flow of
information can be modulated.
A great diversity of regulatory mechanisms has been overlaid on the central
phosphotransfer/phosphorelay pathways, allowing optimization of signal
transmission for the specific needs of each system.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
Some systems output a graded response, such as
the EnvZ-OmpR system that mediates the
differential expression of porin genes ompF and
ompC. Others, such as the B. subtilis Spo system
that controls commitment to sporulation, output
an all-or-nothing response.
Regardless of the output, both types of systems
can involve a significant amount of regulation
and often involve a number of auxiliary protein
(TCS connectors) components. The primary
targets for regulation are the activities of the
HK and dephosphorylation of the RR.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
TCS connectors (which for the sake of brevity will also be called connectors) are an emerging group
of proteins that modulate the activity of sensor kinases and response regulators at the post-
translational level.
Because connector proteins are typically synthesized in response to signals that are different from
those sensed by the cognate sensor, they often establish regulatory links between otherwise
independent signal transduction pathways (in other words, they “connect” a TCS to the signal(s)
controlling a different regulatory system).
TCS connectors
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TCS connectors use a variety of mechanisms to alter response regulator output:
1. Inhibiting sensor kinase autophosphorylation
2. Promoting dephosphorylation of phosphorylated response regulators and His-containing
phosphotransfer proteins
3. Inhibiting response regulator dephosphorylation
4. Activating a sensor kinase
5. Inhibiting DNA binding by a response regulator
6. Inhibiting recruitment of RNA polymerase to promoters
7. Sequestering adaptor proteins that promote protein degradation
produced by: sepideh benvari supervised by: Dr. Aslanimehr
Inhibiting sensor kinase
autophosphorylation
TCS connectors can inhibit sensor autophosphorylation or promote dephosphorylation of
phosphorylated response regulators.
In the B. subtilis phosphorelay, the sensor kinase KinA is activated by an unknown signal,
which results in autophosphorylation from ATP and subsequent phosphotransfer to the
response regulator Spo0F. Spo0F passes on the phosphoryl group to the His-containing
protein Spo0B, which in turn transfers it to the terminal acceptor, the response
regulator Spo0A.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
The phosphorylated form of Spo0A acts as a
transcription factor, being the key activator of
sporulation genes. The connectors Sda and KipI
block activation of the phosphorelay by inhibiting
KinA autophosphorylation. The connectors RapA,
RapB, RapE, and RapH promote
dephosphorylation of the response regulator
Spo0F-P; the connectors Spo0E, YisI, and YnzD act
in a similar way on Spo0A-P.
The expression of connectors is controlled by
factors such as growth conditions, status of the
DNA replication machinery, and development of
competence (through the action of ComA and
ComK the key regulators of competence genes).
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Inhibiting response regulator
dephosphorylation
TCS connectors can promote activation of response regulators and sensor kinases.
The connector-mediated pathway from S. enterica. The low Mg2+ signal activates the
PhoP/PhoQ TCS, which triggers the expression of the connector PmrD. PmrD binds to
the phosphorylated form of the response regulator PmrA, thereby protecting it from
dephosphorylation.
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PmrA-P binds to DNA and regulates its target promoters. PmrA-P represses
transcription of the pmrD gene, thus establishing a negative feedback loop controlling
PmrD expression. The PmrA/PmrB TCS can be activated directly by the Fe3+ signal.
produced by: sepideh benvari supervised by: Dr. Aslanimehr
Inhibiting recruitment of RNA polymerase
to promoters
TCS connectors can inhibit binding of activated response regulators to DNA or prevent RNA polymerase from interacting
with a response regulator.
The ComA/ComP TCS from B. subtilis responds to the extracellular quorum-sensing signal, the peptide ComX. Upon
activation, ComA promotes transcription of the srf operon, which leads to development of the competent state.
Binding of ComA to DNA is inhibited by the connectors RapC, RapF, and RapH.
These connectors, in turn, are deactivated upon binding to the corresponding Phr
peptides. The action of ComA is also inhibited by the connector Spx, which disrupts
the interaction between ComA and RNA polymerase.
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POTENTIAL TARGETS FOR ANTIMICROBIAL
THERAPY
The search for structurally unique antibiotics that inhibit newmolecular targets has led researchers to
prokaryotic two-component systems.
Two-component systems are attractive for several reasons. First of all, they are widespread in bacteria and,
sofar, absent in mammals. Therefore, general HK or RR inhibitors could potentially be broad-spectrum
antibiotics.
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Perhaps the most attractive reason for targeting two-component systems is that they are used
by pathogenic bacteria to control the expression of virulence factors required for infectivity.
Several well-characterized virulence systems are the A. tumefaciens VirA/VirG system, the B.
pertussis BvgA/BvgS system, and the Salmonella typhimurium PhoP/PhoQ system. Interestingly, some bacteria have developed two-component systems that regulate
resistance to certain chemotherapeutics.
These include the vancomycin resistance systems in Enterococcus faecalis (VanR/VanS)
and Streptococcus pneumoniae (VncS/VncR), as well as the system associated with
tetracycline resistance in Bacteroides fragilis (RprX/RprY).
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Hexose phosphate is an important carbon source within the cytoplasm of host cells. Bacterial pathogens that invade, survive, and multiply within various host epithelial cells exploit hexose phosphates from the host cytoplasm through the hexose phosphate transport (HPT) system to gain energy and synthesize cellular components.
In Escherichia coli, the HPT system consists of a two-component regulatory system (UhpAB) and a phosphate sensor protein (UhpC) that tightly regulate expression of a hexose phosphate transporter (UhpT).
Staphylococcus aureus also can invade, survive, and multiply within various host epithelial cells. the HPT system in S. aureus that includes the hptRS (a novel two-component regulatory system), the hptA (a putative phosphate sensor), and the uhpT (a hexose phosphate transporter) genes.
We demonstrated that both hptA and hptRS are required to positively regulate transcription of uhpT in response to extracellular phosphates, such as glycerol-3-phosphate (G3P), glucose-6-phosphate (G6P), and fosfomycin.
disruption of the hptA, hptRS, or uhpT gene impaired the growth of bacteria when the available carbon source was
limited to G6P, impaired survival/multiplication within various types of host cells, and increased resistance to
fosfomycin.
the HPT system plays an important role in adaptation of S. aureus within the host cells and could be an important target
for developing novel antistaphylococcal therapies.
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Pseudomonas aeruginosa is an opportunistic human pathogen that causes severe, life-threatening infections in patients with cystic fibrosis (CF), endocarditis, wounds, or artificial implants.
During CF pulmonary infections, P. aeruginosa often encounters environments where the levels of calcium (Ca2) are elevated. P. aeruginosa responds to externally added Ca2 through enhanced biofilm formation, increased production of several secreted virulence factors, and by developing a transient increase in the intracellular Ca2 level, followed by its removal to the basal submicromolar level.
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we used microarray analysis to characterize the global transcriptional response of P. aeruginosa to elevated external Ca2 levels. we identified the TCS PA2656-PA2657 (here referred to as carSR, for calcium regulator sensor and regulator), whose transcription is highly induced by elevated Ca2 in planktonic cultures of P. aeruginosa PAO1.
Using deletion mutations and microarray analysis, we identified the regulatory targets of carSR, which include the hypothetical proteins PA0320 and PA0327. Further characterization of PA0320 and PA0327 indicate that they play roles in maintaining Ca2 homeostasis. PA0327 also influences the production of the virulence factor pyocyanin and swarming motility in a Ca2-dependent manner.
produced by: sepideh benvari supervised by: Dr. Aslanimehr