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Gene Network Model and Quorum Sensing in Pseudomonas Aeruginosa ELE 580B- Cellular and Biochemical Computing Project Presentation Hidekazu OKI & Canturk.

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Presentation on theme: "Gene Network Model and Quorum Sensing in Pseudomonas Aeruginosa ELE 580B- Cellular and Biochemical Computing Project Presentation Hidekazu OKI & Canturk."— Presentation transcript:

1 Gene Network Model and Quorum Sensing in Pseudomonas Aeruginosa ELE 580B- Cellular and Biochemical Computing Project Presentation Hidekazu OKI & Canturk ISCI

2 Project Workplan Quorum Sensing mechanisms in P.A. Gene network for P.A. Biochemical Reactions Possible Simulation Techniques Simulation and results

3 Pseudomonas Aeruginosa Lethal, opportunistic, Gram negative human pathogen Gram negative LasB elastase, LasA Elastase, Alkaline Protease  degrade Elastin (lung & blood vessels) ExotoxinA  inhibit protein synthesis Uses cell-cell signaling – quorum sensing – to overcome host defense Communal behaviour

4 Quorum Sensing Generic Quorum Sensing Mechanism: (xxx-HSL) Binding Occurs only at high AI concentrations

5 Quorum Sensing Signaling Molecules Gram ‘ - ’  HSL ring & Fatty acid side chain Different side chains  Different AIs Gram ‘ + ’  Oligo Peptides Gram ‘ + ’

6 Quorum Sensing in PA 2 Hierarchical xxxI-xxxR systems: 1) las system 2) rhl system Las System: lasI  LasI  3-oxo-C12-HSL (PAI1) lasR  LasR LasR/3-oxo-C12-HSL  lasA, lasB, aprA, toxA, etc. & lasI  rhlR – hierarchy!!

7 Quorum Sensing in PA Rhl System: rhlI  RhlI  C4-HSL (PAI2) rhlR  RhlR RhlR/C4-HSL  rhlAB operon, lasA, lasB, aprA & other genes & rhlI PQS AutoInducer: Additional link between las-rhl LasR  PQS  lasB & rhlI

8 Quorum Sensing in PA Informal Description: lasI lasR LasI PAI1 LasR PQS LasR rhlI rhlR RhlI PAI2 RhlR PAI1 can bind to RhlR and block it!

9 PA Gene Network Is it important to make a detailed circuit like the circuit? All the promoters, repressors, activators, specified explicitly We care about i/p-o/p and cause-effect relations Our Model: I/p  Gene  O/p Protein (  Secondary o/p) Details of i/p strength hidden in affinities of chemical reactions

10 PA Gene Network All mentioned genes + las system inputs + additional downstream genes LasR/PAI1  excitatory on rhlR PAI1  inhibitory on RhlR alkaline phosphatase exotoxinA LasR/{3-oxo-C12-HSL} {3-oxo-C12-HSL} Vfr lasR lasI LasR LasI LasR lasA lasB LasA LasB aprA toxA xcpP xcpR ? ? ? ? GacA + RsaL - RhlR/{C4-HSL} {C4-HSL} rhlR rhlI RhlR RhlI RhlR {3-oxo-C12-HSL} RhlR/{3-oxo-C12-HSL} RhlR  Next Slide PQS +

11 PA Gene Network All mentioned genes + additional downstream genes alkaline phosphatase RhlR/{C4-HSL} RhlR lasA rhlAB LasA RhlAB aprA lasB ? rpoS  s s  s s LasB pyocyanin ? lecA cyanide cytoxic lectin

12 PA Biochemical Reactions Reactions that describe the core of the quorum sensing mechanism 1) LasR/PAI1 complex: R1: Concentration of LasR A1: Concentration of PAI1 C1: Concentration of Lasr/PAI1

13 PA Biochemical Reactions 2) RhlR/PAI2 complex: R2: Concentration of RhlR A2: Concentration of PAI2 C2: Concentration of RhlR/PAI2 3) RhlR/PAI1 complex: C3: Concentration of RhlR/PAI1

14 PA Biochemical Reactions 4) LasR: b R1 : Degradation rate of LasR V R1 : Maximum production rate of LasR K R1 : Affinity between C1 and lasR promoter! R 10 : LasR basal production rate

15 PA Biochemical Reactions 5) RhlR: b R2 : Degradation rate of RhlR V R2 : Maximum production rate of RhlR K R2 : Affinity between C1 and rhlR promoter R 20 : RhlR basal production rate

16 6) RsaL: S : RsaL concentration b S : Degradation rate of RsaL V S : Maximum production rate of RsaL K S : Affinity between C1 and rsaL promoter S 0 : RsaL basal production rate PA Biochemical Reactions

17 7) PAI1: b A1 : Degradation rate of PAI1 V A1 : Maximum production rate of PAI1 K A1 : Affinity between C1 and lasI promoter K S1 : Affinity between RsaL and lasI promoter A 10 : PAI1 basal production rate A 1ex : Extracellular PAI1 PA Biochemical Reactions

18 8) PAI2: b A2 : Degradation rate of PAI2 V A2 : Maximum production rate of PAI2 K A2 : Affinity between C2 and rhlI promoter A 20 : PAI2 basal production rate A 2ex : Extracellular PAI2 PA Biochemical Reactions

19 Simulation Methodology Deterministic, Single-Cell model Numerical Integration of Ordinary Differential Equations. C Program simulator. Time step = 0.01 hours. Total simulated time varied from 100 hours to 10,000 hours.

20 Simulation Results (1) Low concentration of extra-cellular PAI1 causes cell to remain in inactive state. LasR/PAI1 complex concentration is low

21 Increasing extra-cellular concentration of PAI1 beyond 2.0 causes the system to eventually reach active state. Simulation Results (2)

22 Simulation Results (3) Final Steady-State concentrations vary sharply depending on the extra-cellular PAI1 concentration: (KR1 = 4, KA1 = 0.4) (KR1= 5.0, KA1 = 0.6 )

23 Index of Terms Gram negative: … cell wall of Gram-negative bacteria is a thinner structure with distinct layers. There is an outer layer which is more like a cytoplasmic membrane in composition with the typical trilaminar structure. Gram Positive: … are characterised by having as part of their cell wall structure eptidoglycan as well as polysaccharides and/or teichoic acids. Ba ck Ba ck

24 References David's Paper (lecture 11) --> about quorum and PA --> slides come from this web in the lecture 11 pres M. Miller and B Bassler, “ Quorum Sensing in Bacteria ”, Annual Review of Microbiology, 55: , > Rweiss reading list paper -- > WEB page about PA and quorum L. Passador and B. Iglewski, "Quorum Sensing and Virulence Gene Regulation in Pseudomonas Aeruginosa", Virulence mechanisms of bacterial pathogens, 1995 lecture 7 slides --> the lambda cct and the determinstic vs stochastic simulation models Fagerlind, Magnus. “ The role of regulators on the expression of quorum-sensing signals in Pseudomonas aeruginosa ” A thesis of 20p in molecular computational biology for the degree of Bachelor of Science at the University of Skovde. Oct, 2001 Albus, Anne M., etal. “ Vfr Controls Quorum Sensing in Pseudomonas Aeruginosa ” Journal of Bacteriology, June 1997, p

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