DARPA BioComp PI Meeting, 2001DARPA BioComp PI Meeting, 2001

““The Eukaryotic Cell Cycle as a Test CaseThe Eukaryotic Cell Cycle as a Test Casefor Modeling Cellular Regulation in a for Modeling Cellular Regulation in a

Collaborative Problem Solving Environment”Collaborative Problem Solving Environment”

PI: John J. Tyson (Biology, Virginia Tech)PI: John J. Tyson (Biology, Virginia Tech)CoPIs: Cliff Shaffer & Layne Watson (CS, Virginia Tech)CoPIs: Cliff Shaffer & Layne Watson (CS, Virginia Tech)

With collaborators atWith collaborators atRockefeller UniversityRockefeller University

University of Kentucky Medical SchoolUniversity of Kentucky Medical SchoolBudapest University of Technology and EconomicsBudapest University of Technology and Economics

Cdc2

Cdc13

START

S

cell d

ivision

G2

FIN

ISH

G1

DNAreplication

Metaphase

Anaphase

cyclin-dependentkinase

cyclin

Three ObjectivesThree Objectives

•ModelingModeling•Experimental ValidationExperimental Validation•Software ToolsSoftware Tools

DoD RelevanceDoD Relevance

The cell cycle is central to all processes of The cell cycle is central to all processes of biological growth, development and biological growth, development and

reproduction.reproduction. • Wound healingWound healing• Nerve regenerationNerve regeneration• Radiation damageRadiation damage• Eukaryotic pathogensEukaryotic pathogens• Tissue cultureTissue culture• CancerCancer

Kurt Kohn (1999) Mol Biol Cell

Getting in

Touch with

Your Inner Y

east

Getting in

Touch with

Your Inner Y

east

Clb5MBF

P Sic1SCF

Sic1Swi5

Clb2Mcm1

Unaligned chromosomes

Cln2Clb2

Clb5

Cdc20 Cdc20

Cdh1

Cdh1

Cdc20

APC

PPX

Mcm1

SBF

Esp1Esp1 Pds1

Pds1

Cdc20

Net1

Net1P

Cdc14

RENT

Cdc14

Cdc14

Cdc15

Tem1

Bub2

CDKs

Esp1

Mcm1 Mad2

Esp1

Unaligned chromosomes

Cdc15

Lte1

Budding

Cln2SBF

?

Cln3

Bck2

and

growth

Sister chromatid separation

DNA synthesis

Table 6. Properties of clb, sic1, and hct1 mutants

mass at birth

mass at

SBF 50%

mass at

DNA repl.

mass at bud ini.

mass at division

TG1

(min)

changed

parameter

Comments

1 wild type

(daughter) 0.71 1.07

(71’) 1.15 (84’)

1.15 (84’)

1.64 (146’)

84 CT 146 min (time of occurrence of event)

2 clb1 clb2

0.71 1.07 1.16 1.16 No mit k's,b2 = 0

k"s,b2 = 0 Surana 1991 Table 1, G2 arrest.

3 clb1 clb2

1X GAL-CLB2 0.65 1.10 1.19 1.19 1.50 105 k's,b2 = 0.1

k"s,b2 = 0 Surana 1993 Fig 4, 1X GAL-CLB2 is OK, 4X GAL-CLB2 (or 1X GAL-CLB2db) causes telophase arrest.

4 clb5 clb6 0.73 1.07

(65’) 1.30 (99’)

1.17 (80’)

1.70 (146’)

99 k's,b5 = 0 k"s,b5 = 0

Schwob 1993 Fig 4, DNA repl begins 30 min after SBF activation.

5 clb5 clb6

GAL-CLB5 0.61 0.93 0.92 0.96 1.41 73 k's,b5 = 0.1

k"s,b5 = 0 Schwob 1993 Fig 6, DNA repl concurrent with SBF activation in both GAL-CLB5 and GAL-CLB5db.

6 sic1 0.66 1.00

(73’) 0.82 (37’)

1.06 (83’)

1.52 (146’)

38 k's,c1 = 0 k"s,c1 = 0

Schneider 1996 Fig 4, sic1 uncouples S phase from budding.

7 sic1 GAL-SIC1 0.80 1.07 1.38 1.17 1.86 94 k's,c1 = 0.1 k"s,c1 = 0

Verma 1997 Fig3B, Nugroho & Mendenhall 1994 Fig 2, most cells are viable.

8 hct1 0.73 1.08 1.17 1.18 1.69 82 k"d,b2 = 0.01 Schwab 1997 Fig 2, viable, size like WT, Clb2 level high

throughout the cycle. 9 sic1 hct1

0.71 No SBF 0.72 No bud No mit k's,c1 = 0

k"d,b2 = 0.01 Visintin 1997, telophase arrest.

10 sic1 GAL-CLB5

first cycle second cycle

0.71 0.52

0.74

0.73

No repl

0.76

1.20

k's,b5 = 0.1 k"s,b5 = 0 k's,c1 = 0

Schwob 1994 Fig 7C, inviable. First cycle OK, DNA repl advanced; but pre-repl complexes cannot form and cell dies after the first cycle.

d CDK dt = k1 - (v2’ + v2” . Cdh1 ) . CDK

d Cdh1dt =

(k3’ + k3” . Cdc20A) (1 - Cdh1) J3 + 1 - Cdh1 -

(k4’ + k4” . CDK . M) Cdh1 J4 + Cdh1

d IEPdt = k9

. CDK . M . (1 – IEP ) – k10 . IEP

d Cdc20T

dt = k5’ + k5” (CDK . M)4

J54 + (CDK . M)4 - k6

. Cdc20T

d Cdc20A

dt = k7

. IEP (Cdc20T - Cdc20A) J7 + Cdc20T - Cdc20A

- k8

. MAD Cdc20A

J8 + Cdc20A - k6

. Cdc20T

Differential equations Parameter values

k1 = 0.0013, v2’ = 0.001, v2” = 0.17,

k3’ = 0.02, k3” = 0.85, k4’ = 0.01, k4” = 0.9,

J3 = 0.01, J4 = 0.01, k9 = 0.38, k10 = 0.2,

k5’ = 0.005, k5” = 2.4, J5 = 0.5, k6 = 0.33,

k7 = 2.2, J7 = 0.05, k8 = 0.2, J8 = 0.05,

…

Clb/Cdk1activity

A + Cln2B+Cdc20

A/B

G1

S/G2/M

Start

Finish

Predictions: Budding YeastPredictions: Budding Yeast

1. Rate constants1. Rate constants

2. Hysteresis2. Hysteresis

Mendenhall (U Kentucky)Mendenhall (U Kentucky)

Cross (Rockefeller U)Cross (Rockefeller U)

3. Mutants3. Mutants Cross & MendenhallCross & Mendenhall

Predictions: Frog EggsPredictions: Frog Eggs

1. Rate constants (1993)1. Rate constants (1993)

2. Hysteresis (1993)2. Hysteresis (1993)

3. Bifurcation diagram (1998)3. Bifurcation diagram (1998)

ConfirmedConfirmed Kumagai & Dunphy (1995)Kumagai & Dunphy (1995)

ConfirmedConfirmed Moore (unpubl)Moore (unpubl)

Sible & Sha (VA Tech)Sible & Sha (VA Tech)

Under testUnder test Sible & Sha (VA Tech)Sible & Sha (VA Tech)

4. Cdk2/Cyclin E (in ppn)4. Cdk2/Cyclin E (in ppn)Under testUnder test

Sible (VA Tech)Sible (VA Tech)

Software RequirementsSoftware Requirements

Experimental DatabaseExperimental Database

Wiring DiagramWiring Diagram

Differential EquationsDifferential Equations Parameter ValuesParameter Values

AnalysisAnalysis SimulationSimulation

Visualization-TranslationVisualization-Translation

Experimental DatabaseExperimental Database

0 1 20.0

0.5

1.0

G1

M

cell mass

Bifurcation diagram

Cdc20

Cdk1

Clb2,5

Cln2

Sic1Cdh1

Cd

k1 a

ctiv

ity

Parameter EstimationParameter Estimation

DatabaseDatabaseSimulationSimulation

Prop 1Prop 1Prop 1Prop 1

Prop 2Prop 2Prop 2Prop 2

............

ComparatorComparator

Good fitGood fit

Bad fitBad fit

......

Error Function (parameters)Error Function (parameters)

Parameter EstimationParameter Estimation

trust region Levenburg Marquardt

Jones’ direct search global optimizer

MilestonesMilestones

Year OneYear One Year TwoYear Two Year ThreeYear Three

ModelingModeling Full model of bud Full model of bud yeastyeast

Sensitivity analysisSensitivity analysis

Cdk2/cyclin E in frogCdk2/cyclin E in frog

Morpho checkpointMorpho checkpoint

-factor signalling-factor signalling

Revisions Revisions

Comprehensive Comprehensive model of budding model of budding yeastyeast

Comprehensive Comprehensive model of frog eggmodel of frog egg

ExperimenExperimentt

Absolute prot concen Absolute prot concen in budd yeastin budd yeast

Hysteresis in frog extr Hysteresis in frog extr and in budd yeastand in budd yeast

Kinetic & thermody Kinetic & thermody propertiesproperties

Hopf & SNIPER Hopf & SNIPER bifns in frog extrbifns in frog extr

MutantsMutants

Complex bifns in Complex bifns in budd yeast & frogbudd yeast & frog

Checkpoint signalsCheckpoint signals

SoftwareSoftware Model builderModel builder

Steady-state finderSteady-state finder

ComparatorComparator

Param optimization in Param optimization in frog egg modelfrog egg model

Numer bifn analysisNumer bifn analysis

Run managementRun management

Simul analysisSimul analysis

Param optimization Param optimization in yeast cell modelin yeast cell model

Working PSE for Working PSE for cell cycle modelingcell cycle modeling

DeliverablesDeliverables

Year OneYear One Year TwoYear Two Year ThreeYear Three

ModelingModeling 2 publications2 publications

budding yeastbudding yeast

frog egg developfrog egg develop

1 publication1 publication

morpho checkptmorpho checkpt

2 tech reports2 tech reports

3 publications3 publications

-factor -factor signallingsignalling

compreh yeastcompreh yeast

compreh frogcompreh frog

ExperimenExperimentt

2 publications2 publications

yeast (Cross)yeast (Cross)

frog (Sible)frog (Sible)

1 tech report 1 tech report (Menden)(Menden)

Some combination Some combination of publications and of publications and technical reports on technical reports on experimentsexperiments

3 publications3 publications

irrev trans in irrev trans in yeastyeast

kinet & thermo kinet & thermo parpar

bifns in frog extrbifns in frog extr

SoftwareSoftware 3 software tools3 software tools

model buildermodel builder

steady-state findersteady-state finder

param optimizerparam optimizer

Publications Publications

Software tools forSoftware tools for

run run managementmanagement

bifn analysisbifn analysis

comparisonscomparisons

deter global deter global searchsearch

PublicationsPublications

Suites of integrated Suites of integrated software tools for a software tools for a complete cell-cycle complete cell-cycle problem-solving-problem-solving-environmentenvironment

PersonnelPersonnel

VT FacultyVT Faculty VT StudentsVT Students CollaboratorsCollaborators

ModelingModeling Tyson (25%)Tyson (25%)

Chen (25%)Chen (25%)Ciliberto Ciliberto (25%)(25%)

Yi (25%)Yi (25%)

Novak (20%)Novak (20%)

Pataki (100%)Pataki (100%)

ExperimentExperiment Sible (15%)Sible (15%) Sha (50%)Sha (50%) Cross (25%)Cross (25%)

Liu (100%)Liu (100%)

Menden. Menden. (25%)(25%)

GRA (100%)GRA (100%)

SoftwareSoftware Shaffer Shaffer (15%)(15%)

Watson Watson (15%)(15%)

Ramak. Ramak. (10%)(10%)

Zwolak Zwolak (100%)(100%)

Vass (100%)Vass (100%)

Allen (100%)Allen (100%)