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

outline

• Biological background

• The building blocks of a programmed cell

• Examples

• summary

The central dogma

Transcription factors control gene expression

• Activators:

• Repressors

A B

A B

Activator A

RNA pol

RNA pol ON

Repressor A

RNA pol

OFF

Gene B

Gene B

inputoutput

chemo taxisFood gradient Direction of movement

Sugar

Metabolism

The composition of sugars in the environment

Activation of lac operon

Cells need to compute in order to survive

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

A modular programmed cell

input

engineered regulatory network

requirments

•the regulatory network can read the input

•The cell can read the output

Sensory

moduleoutput

Kobayashi, Kærn, Araki, Chung, Gardner, Cantor & Collins,( PNAS 2004).

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

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

Phage λ uses a toggle switch to transfer between 2 cycles

Phage lambda toggle switch uses the E.coli SOS system

Lytic phase Lysogenic phase

DNA damage

RecAUV

radiation

Phage genes

OFF

Phage genes

ON

SOS

repressor

Interfacing the genetic switch with SOS pathway and an output module

regulatory module-

genetic switch

GFP*LP

output module - GFP

ON

DNA damage

SS DNA

UV RECA Sensory module-

SOS system

DNA

E.Coli cell

The system is applied using plasmidsPlasmid - an extra-chromosomal DNA molecule which is capable of replicating

GFP

output module

Regulatory module

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

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.

Example 2 : population control

luxI/luxR system – the engineered regulatory module

You, Cox, Weiss & Arnold ( 2004 NATURE)

Construction of the population control program

Cells divide

More I More R * More E

Less cells

Less ILess R*Less E

Output module

Regulatory moduleinput

Results : The system reaches a steady state

the cell density in the active programmed cell is 10 fold lower.

Wt cells Programmed cells

Concentration of the killer protein in the active programmed cell is 1000 fold higher

Results : The killer protein concentration

Programmed cellswt cells

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