Two component systems Flashcards
What signals do bacteria respond to? (8)
ENVIRONMENTAL CUES
Temperature Acidic pH Ions Population density (QS) Osmolarity Presence of antimicrobials Oxygen availability Nutrient availability
Two-component systems
histidine sensor kinase detects an environmental signal and autophosphorylates and passes to an aspartate on the RR. The RR is usually a DNA binding protein or a TF
Sometimes, intermediate proteins facilitate a phosphorelay
Phosphatase activity resets the system - In many cases, TCSs auto-regulate themselves
they are very specific to the particular system
Same genes can be activated by multiple 2CS: allows the same set of genes to be activated by multiple environmental stressors
2CS in ECOLI: EnvZ/OmpR
in ecoli
OmpF and OmpC are channels made of porin proteins; allow transport of small molecules
balance between OmpF & OmpC is critical, and corresponds with the osmolarity of the environment
OmpC pores are slightly smaller; favoured during growth at high osmotic pressures
Larger OmpF pores are favoured in more dilute environments, allowing solutes to diffuse into the cell more readily
EnvZ is the
sensor kinase that detects the osmolarity between the cell and the environment
at higher osmolarity, it autophosphorylates
passes phosphate to OmpR
OmpR binds DNA at its CTD
OmpR activates or represses ompC or ompF (outer membrane porins)
when OmpR is not active = LOW osmolarity = ompF on (larger porin to scavenge nutrients)
when OmpR is phosphorylated = HIGH omsolarity = ompC on (smaller porin) AND ompF repressed
2CS in SALMONELLA
PhoP/Q & PmrA/B
both connected by PmrD which is activated by the PhoP/Q system. PmrD protects PmrA from dephosphorylation by PmrB, activating PmrA
PhoP/Q contributes to resistance to bile salts and is activated in the GI tract
PhoP/Q and PmrA/B both confer resistance to antimicrobial peptides
APs are + charged, and have high affinity for the neg charged bacterial cell envelope. PhoPQ/PmrAB modify the lipid A region by the addition of aminoarabinose which reduces the net negative charge and lowers the affinity for APs
How does PhoQ respond to multiple stimuli?
Magnesium ions form a bridge between the negatively-charged membrane and a negatively-charged domain of the PhoQ protein. Low magnesium concentrations, acidic pH and the presence of antimicrobial peptides can all impact on this magnesium bridge, leading to PhoQ activation.
Low Mg2+ concentrations result in loss of bridges = ACTIVATION
Acidic pH alters conformation of the periplasmic domain = ACTIVATION
Positively-charged antimicrobial peptides displace Mg2+ = ACTIVATION
How might PhoQ help intracellular survival of salmonella?
Hypothesis:
Phagolysosomes have low Mg2+ concentrations, thus the PhoP/Q system is activated during phagocytosis
PhoP/Q activation induces Mg2+ transport mechanisms & LPS modifications that promote intracellular survival
PhoP/Q is primarily triggered by acidic pH but also magnesium
Extra reading
The pmrCAB operon produces three protein products: a phosphoethanolamine (pEtN) phosphotransferase (PmrC), a response regulator (PmrA) and a sensor kinase (PmrB).
PmrAB regulates over 20 confirmed and possibly up to
100 genes in Salmonella
Orthologs of the Salmonella PmrAB TCS exist in several Gram-negative pathogens. In Yersinia spp., Escherichia coli and Pseudomonas aeruginosa, PmrA also
activates genes involved in the modification of LPS
Activation of PmrAB can occur by direct or indirect means
Direct PmrAB activation is thought to be mediated by PmrB through its periplasmic domain. Known activating signals for Salmonella are ferric (Fe+3) iron, aluminum (Al+3), low pH (e.g. pH 5.5) and vanadate.
Indirect activation of PmrAB occurs through the PhoP/Q
TCS. PhoP/Q activates the expression of pmrD.
PmrD produces a 9.6 kDa product that regulates PmrA
activity at a post-transcriptional level, as PmrD binds to
and stabilizes PmrA in its phosphorylated form
One of the primary roles of PmrAB activation is LPS
modification. These modifications include both
Ara4N and pEtN additions to the lipid A and pEtN
addition to the LPS core. These modifications mask phosphate groups with positively charged moieties, affecting the electrostatic interaction of certain
cationic APs with the bacterial cell surface dramatically
Extra reading
The pmrCAB operon produces three protein products: a phosphoethanolamine (pEtN) phosphotransferase (PmrC), a response regulator (PmrA) and a sensor kinase (PmrB).
PmrAB regulates over 20 confirmed and possibly up to
100 genes in Salmonella
Orthologs of the Salmonella PmrAB TCS exist in several Gram-negative pathogens. In Yersinia spp., Escherichia coli and Pseudomonas aeruginosa, PmrA also
activates genes involved in the modification of LPS
Chemotaxis is regulated by a two-component system
CheA-CheY
regulates the chemotactic response through direct interaction with flagellar motor proteins, influencing direction of rotation
Two-component systems involved in chemotaxis differ:
The sensor kinase (CheA) is cytoplasmic
The response regulator (CheY) interacts directly with the motor proteins of the flagellar basal body
Proteins of the chemotaxis signalling pathway
The cytoplasmic CheA sensor kinase; interacts with chemoreceptors (temporal sensing)
When activated, CheA phosphorylates CheY (response regulator) and CheB (a methyl esterase)
Phosphorylated CheY interacts with the FliM protein of the flagellar motor
The chemotaxis signalling pathway
methyl-accepting chemotaxis proteins (MCPs)
Moving up a concentration gradient of attractant (i.e. towards attractant)
Attractant is bound to MCP, which inhibits CheA autophosphorylation
> CheY is inactive > flagellar rotation remains CCW (forward run)
Moving down a concentration gradient (i.e. away from attractant)
If attractant levels fall, MCP-bound attractant decreases, activating CheA autophosphorylation
> Activation of CheY (CheY-P) > Interacts with FliM
> CW rotation (tumbling)
CheZ dephosphorylates CheY-P to reset the system (CCW rotation)
SoxR/S - evading the respiratory burst
senses the redox active compounds, causes a conformational change in the sox R dimer which will bing and this causes transcriptional activation of sox S, and sox S is another transcription activator which then transcribles superoxide dismutase, acrA and acrB (efflux pump system which can remove the radical species)
GRAS/GRAR Resistance to antimicrobial peptides in G+
(glycopeptide resistance association)
GraS probably functions as a sensor protein kinase which is autophosphorylated at a histidine residue and transfers its phosphate group to GraR. Confers resistance to vancomycin and polymyxin B
glycopeptides are cationic amps (CAMPs)
Lysosome contains glycopeptides that will damage G+ GRAS/R TCS controls the transcription of the DLT operon (LTA of G+ve cell wall is negatively charged) upregulated transcription of DTA causing +ve alanine to be added on to the LTA (lipoteic acid) of the G+ve cell wall
