1. DARPA BioComp PI Meeting, 2001 “The Eukaryotic Cell Cycle as a Test Case for Modeling Cellular Regulation in a Collaborative Problem Solving Environment” PI: John J. Tyson (Biology, Virginia Tech) CoPIs: Cliff Shaffer & Layne Watson (CS, Virginia Tech) With collaborators at Rockefeller University University of Kentucky Medical School Budapest University of Technology and Economics

2. Cdc2 Cdc13 START S cell division G2 FINISH G1 DNA replication Metaphase Anaphase cyclin-dependent kinase cyclin

3. Three Objectives ModelingModeling Experimental ValidationExperimental Validation Software ToolsSoftware Tools

4. DoD Relevance The cell cycle is central to all processes of biological growth, development and reproduction. Wound healingWound healing Nerve regenerationNerve regeneration Radiation damageRadiation damage Eukaryotic pathogensEukaryotic pathogens Tissue cultureTissue culture CancerCancer

5. Kurt Kohn (1999) Mol Biol Cell

8. Getting in Touch with Your Inner Yeast

9. Clb5 MBF P Sic1 SCF Sic1 Swi5 Clb2 Mcm1 Unaligned chromosomes Cln2 Clb2 Clb5 Cdc20 Cdh1 Cdc20 APC PPX Mcm1 SBF Esp1 Pds1 Cdc20 Net1 Net1P Cdc14 RENT Cdc14 Cdc15 Tem1 Bub2 CDKs Esp1 Mcm1 Mad2 Esp1 Unaligned chromosomes Cdc15 Lte1 Budding Cln2 SBF ? Cln3 Bck2 and growth Sister chromatid separation DNA synthesis

10. 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, …

11. Clb/Cdk1 activity A + Cln2 B+Cdc20 A/B G1 S/G2/M Start Finish

12. Predictions: Budding Yeast 1. Rate constants 2. Hysteresis Mendenhall (U Kentucky) Cross (Rockefeller U) 3. Mutants Cross & Mendenhall

14. Predictions: Frog Eggs 1. Rate constants (1993) 2. Hysteresis (1993) 3. Bifurcation diagram (1998) Confirmed Kumagai & Dunphy (1995) Confirmed Moore (unpubl) Sible & Sha (VA Tech) Under test Sible & Sha (VA Tech) 4. Cdk2/Cyclin E (in ppn) Under test Sible (VA Tech)

15. Software Requirements Experimental Database Wiring Diagram Differential Equations Parameter Values Analysis Simulation Visualization-Translation Experimental Database

16. G1 M cell mass Bifurcation diagram Cdc20 Cdk1 Clb2,5 Cln2 Sic1 Cdh1 Cdk1 activity

17. Parameter Estimation DatabaseSimulation Prop 1 Prop 2......Comparator Good fit Bad fit... Error Function (parameters)

18. Parameter Estimation trust region Levenburg Marquardt Jones’ direct search global optimizer

20. Milestones Year One Year Two Year Three Modeling Full model of bud yeast Sensitivity analysis Cdk2/cyclin E in frog Morpho checkpoint  -factor signalling Revisions Comprehensive model of budding yeast Comprehensive model of frog egg Experiment Absolute prot concen in budd yeast Hysteresis in frog extr and in budd yeast Kinetic & thermody properties Hopf & SNIPER bifns in frog extr Mutants Complex bifns in budd yeast & frog Checkpoint signals Software Model builder Steady-state finder Comparator Param optimization in frog egg model Numer bifn analysis Run management Simul analysis Param optimization in yeast cell model Working PSE for cell cycle modeling

21. Deliverables Year One Year Two Year Three Modeling 2 publications budding yeast budding yeast frog egg develop frog egg develop 1 publication morpho checkpt morpho checkpt 2 tech reports 3 publications  -factor signalling  -factor signalling compreh yeast compreh yeast compreh frog compreh frog Experiment 2 publications yeast (Cross) yeast (Cross) frog (Sible) frog (Sible) 1 tech report (Menden) Some combination of publications and technical reports on experiments 3 publications irrev trans in yeast irrev trans in yeast kinet & thermo par kinet & thermo par bifns in frog extr bifns in frog extr Software 3 software tools model builder model builder steady-state finder steady-state finder param optimizer param optimizerPublications Software tools for run management run management bifn analysis bifn analysis comparisons comparisons deter global search deter global searchPublications Suites of integrated software tools for a complete cell-cycle problem-solving- environment

22. Personnel VT Faculty VT Students Collaborators Modeling Tyson (25%) Chen (25%) Ciliberto (25%) Yi (25%) Novak (20%) Pataki (100%) Experiment Sible (15%) Sha (50%) Cross (25%) Liu (100%) Menden. (25%) GRA (100%) Software Shaffer (15%) Watson (15%) Ramak. (10%) Zwolak (100%) Vass (100%) Allen (100%)