The CsgD protein of Escherichia coli: a relay between bacterial biofilm formation and gene expression. Gualdi, L., Brombacher, E.*, Bertagnoli, S. and Landini P. Dipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano; *Swiss Federal Institute of Environmental Technology (EAWAG), Switzerland
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The CsgD protein of Escherichia coli: a relay between bacterial biofilm formation and gene expression. Gualdi, L., Brombacher, E.*, Bertagnoli, S. and.
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The CsgD protein of Escherichia coli: a relay between bacterial biofilm formation and gene expression.
Gualdi, L., Brombacher, E.*, Bertagnoli, S. and Landini P.
Dipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano;
*Swiss Federal Institute of Environmental Technology (EAWAG), Switzerland
Biofilm formationBiofilm formation:
• 1. Adhesion factors: e.g. flagella in Escherichia coli, Pseudomonas
aeruginosa;
• 2. Extracellular polysaccharides
• 3. Cell density; e.g. “Quorum sensing”
In Enterobacteria such as Escherichia coli, Salmonella enterica
presence of a specific adhesion factor
CURLI FIBERS
Adhesion Microcolony Maturation
Function of curli (thin aggregative fimbriae):
Cell aggregation and clumping
Ability to adhere to a solid surface
Co-regulated with other adhesion genes
CurliCurli:: Long intertwined structureProteinaceousInvolved in adhesion to solid surfaces and in cell-cell aggregation
Locus Genetic structure and organization well conserved in Enterobacteria.
However, silent in many E. coli lab strains.csgBA Operon: encodes curli structural subunits
CsgA major subuint CsgB nucleator protein
csgDEFG Operon: CsgD transcription activator of csgBA operon CsgEFG assembly and transport of curli subunit
Overexpression of the yoaD gene inhibits cell aggregation in a curli-proficient E. coli strain
This result would be consistent with a PDE role for the YoaD protein (inhibition of cellulose biosynthesis), but….
Why would both postitive (csgBA, adrA) and negative (yoaD) factors for bacterial cell aggregation be regulated by the same mechanism?
yoaDPlac
Timing is everything…..
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=adrA expression (DGC) =yoaD expression (PDEA)
A feedback control for cellulose biosynthesis
• CsgD activates the adrA gene, resulting in di-c-GMP accumulation and cellulose biosynthesis
• At the onset of stationary phase, the yoaD gene is also activated to counteract the effect of AdrA and reduce cellulose biosynthesis, possibly to reduce glucose consumption
CsgD may act on intracellular cy-di-GMP pool
Adapted from Camilli and Bassler, Science 2006
CsgD
“Global impact” by the CsgD protein on gene expression
Cytoplasm Outer membrane
pT7-7 pT7-7pT7-CsgD pT7-CsgD
Protein Function Regulation
WrbA Enhancer of trp repressor protein binding to DNA
rpoS-dependent
PflB Pyruvate formate lyase I
(anaerobic metabolism)
Induced anaerobically
GadA Glutamate decarboxylase
(resistance to acid)
rpoS-dependent
CsgG Involved in assembly or transport protein for curli; novel lipoprotein
rpoS-dependent
Dps Unspecifically binds and protects DNA from oxidative damage mediated by hydrogen peroxide
rpoS-dependent
rpoS regulonHightly expressed in
biofilm-forming strains
CsgD seems to activate expression of rpoS-dependent proteins
- + - + CsgD
1 2 3 4
WT rpoS
1 2 3 4
WT rpoS
- + - + CsgD
Indeed, CsgD-dependent alteration in protein expression requires a functional rpoS gene
How does CsgD affect S-dependent expression?
iraP
CsgD
iraP
yaiB= unknown gene regulated by CsgD
yaiB now annotated as iraP and identified as a factor for S stabilization
1 2 3 4 5 6 7
6xHis-S
35 KDa
50 KDa
- + - + - + CsgD
WT iraP rpoS
30 KDa
CsgD affects S intracellular concentrations in a manner dependent on IraP
Presence of rpoS-dependent proteins in cell extracts correlate with S cellular levels
- + - + - + CsgD
1 2 3 4 5 6
WT iraP rpoS
CsgD iraP
csgDEFG
[S][ES]
Genes belonging to rpoS regulon
IraP
CsgD
- Fimbriae and cellulose csg, ymdA, adrA
-Signalling system c-di-GMPadrA, (gsk), yoaD
S-dependent genes
Curli
- Metabolism pyrBI, gat, metA
- Porin ompF, ompT
- Iron-sensing fecR, fhuE
- Cold-shock csp, infA
Activation Repression
Cellulose
Cell aggregation, Surface attachmentMetabolic adaptation to biofilm growth conditions, activation of stress responses(via the rpoS regulon)