1. Programmed cells: Interfacing natural and engineered gene networks Kobayashi, Kærn, Araki, Chung, Gardner, Cantor & Collins,( PNAS 2004). You, Cox, Weiss & Arnold ( 2004 NATURE) Noam Vardi

2. outline Biological background The building blocks of a programmed cell Examples summary

3. The central dogma

4. Transcription factors control gene expression Activators: Repressors A B AB Activator A RNA pol ON Repressor A RNA pol OFF Gene B

5. outputinput Direction of movement Food gradientchemo taxis Activation of lac operon The composition of sugars in the environment Sugar Metabolism Cells need to compute in order to survive

6. Sugar metabolism genes in E.coli The lac genes are transcribed if and only if: 1.Absence of glucose 2.The relevant sugar is present AND gate Expression of alternative sugar genes

7. A modular programmed cell input engineered regulatory network requirments the regulatory network can read the input The cell can read the output Sensory module output Kobayashi, Kærn, Araki, Chung, Gardner, Cantor & Collins,( PNAS 2004).

8. The SOS pathway in E.coli DNA damage RecA SOS repressor SOS genes UV radiation Example 1 : interfacing the S.O.S pathway with a genetic switch

9. A genetic toggle switch All or none reaction – when one promoter is ON, the second one is OFF Each promoter is inhibited by the repressor transcribed from the opposite promoter Phage lambda’s Bi-stable system A Promoter A B promoter B

10. Phage λ uses a toggle switch to transfer between 2 cycles

11. Phage lambda toggle switch uses the E.coli SOS system Lytic phaseLysogenic phase DNA damage RecA UV radiation Phage genes OFF Phage genes ON SOS repressor

12. Interfacing the genetic switch with SOS pathway and an output module regulatory module- genetic switch GFP output module - GFP ON DNA damage SS DNA UV RECA Sensory module- SOS system

13. DNA E.Coli cell The system is applied using plasmids Plasmid - an extra-chromosomal DNA molecule which is capable of replicating GFP output module Regulatory module

14. Example 1 - results Treating cells with UV The genetic switch keep the system active long time after the S.O.S signal is gone and the DNA is repaired. Possible application Detecting DNA damage

15. Quorum sensing - the way bacteria communicate Quorum Sensing - a phenomenon whereby the accumulation of signaling molecules enable a single cell to sense the number of bacteria. Vibrio fischeri.

16. Example 2 : population control luxI/luxR system – the engineered regulatory module You, Cox, Weiss & Arnold ( 2004 NATURE)

17. Construction of the population control program Cells divide More IMore R*More E Less cells Less ILess R*Less E Output module Regulatory module input

18. Results : The system reaches a steady state the cell density in the active programmed cell is 10 fold lower. Wt cellsProgrammed cells

19. Concentration of the killer protein in the active programmed cell is 1000 fold higher Results : The killer protein concentration Programmed cells wt cells

20. summary The unity of the genetic code allows us to transfer circuits between organisms The properties of the output is determined by the characteristic of the network DNA damage detecting circuit (example 1) Population density control circuit (example 2) examples