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Programming Cells

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

#include <iostream> main() { int input;cout << “Please Enter 0 or 1”;cin >> input;if (input = = 1)

cout << “Hello iGEM"; else

cout << “Goodbye”;

return 0; }

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

PromoterRegion

Chemical A

Activator

Gene

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

PromoterRegion

Chemical A

Activator

Gene

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

PromoterRegion

Chemical AActivator

Gene

mRNA

Protein

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

PromoterRegion

Repressor

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

PromoterRegion

Chemical A

Repressor

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

PromoterRegion

Chemical ARepressor

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Goal:Can we Program cells in a manner analogous to how we nowprogram computers?

Programming Cells

PromoterRegion

Repressor

Chemical A

mRNA

Protein

Cellular Pseudocode

{read in input; //light, chemical, etc

if (input is equal to “Chemical A”)“Turn on Gene 1”;

else“Gene 1 remains off”;

}

Programming Cells

But Really We would like to “write more complex cellular programs”

dozens of lines of code (or even 100s)

Multiple input typesMultiple output typesCrosstalkLoopsCounters

Perform a Variety of Activities

Engineering BiologyEngineering

-Framework for Design

We would like a toolbox of Modular Genetic Parts-Standardized Parts (BioBricks)-Swap components- Put together in new ways to perform new function -Portability - Transfer into new organisms or strains -Ability to program

Parts Devices Systems

Library ofPartshttp://parts.mit.edu

Build DevicesWith the Parts

Link Devices Together

Hierarchy:

http://parts.mit.edu

BioBrick Parts Assembly Strategy

The Cell is a Complex System

Schematic of an E. coli cell, by D. Goodsell, Scripps

Dynamic System

Although we do know the complete set of genes for many organisms

We don’t know exactly how everything works together (Goal of Systems Biology)

Synthetic BiologyResearch at the Interface

What can we do with reprogrammed cells:

1. Harness for production (metabolic engineering)-Introduce New Pathways

-Malarial Drug (artemisinin)

2. Coordinate Behavior of CellsTarget cells to tissues or other cell typesRespond to disease states or disease cells (biosensor, target cell death)2-D Patterns

3. Bacteria to Build or Fabricate Systems

Sensors

Sensors-respond to external commands

-Can be used to turn genes on and off-Control motility, etc

3. Environment Responsive Promoter

1. Cytoplasmic Regulatory Proteins

4. Regulatory RNAs

2. Two-Component Systems

Sensors

Cytoplasmic Regulatory Proteins- Inducers – Usually a small molecule – pass through cell membrane

binds to a cytoplasmic regulatory protein1. Turn on an activator2. Turns off a repressor

Graded population induction(All cells behave similar)

Intermediate Induction Difficult

Basal activity

Maximal Induced State

Dynamic Range of Induction

SensorsTwo Component Systems -Membrane Bound Sensor with Kinase Domain

-Responds to different stimuli (light, temp, chemicals)-Phosphorylates a Response Regulator (Triggers Transcription, binds promoter)

Sensor Domain

Kinase Domain P

NarX

tar

SensorsEnvironmental Response Promoters - pH, temp, Oxygen, UV light

-might not know the protein elements involved (but know result)

Sensor Domain

Kinase Domain P

NarX

tar

PromoterRegion

UV light DegradecI repressor

System used in “Tumor Killing Bacteria”-Anaerobic Inducible Promoter

SensorsRegulatory RNAs

RNA aptamers – Change Conformation when bound to small molecule, protein, or peptide

Potentially can be used to regulate any gene

Off conformation(ligand not bound)

On conformation(ligand bound)

On ConformationBinds to target transcriptAnd inhibits transcription(Antisense)

Schematic by C. Smolke

Genetic CircuitsGenetic Circuits

Enable Cells to- Process Input Signals- Make Logical Decisions- Cell-Cell Comunication

Switch-Used to turn on Gene Expression (once input is above a threshold)

Forms:Transcriptional Activators or RepressorsPre-transcription

Genetic CircuitsInverter – a switch that produces a reciprocal response (logic gate)

onrepressor

off

Toggle Switch – can exist in two states-where one or the other repressor is fully expressed-switch can be flipped between states

Genetic Circuits

GFP in a single cell over time (Elowitz and Leibler)

Dynamic Circuits- Oscillator – Cascade

- 3 Repressors

Genetic CircuitsCell – Cell Communication

Sender Cell Receiver Cell

acyl-homoserine lactone (AHL)

Figure from Basu … Weiss

Chemical Signal

ActuatorsActuators-To control the outputMechanical Device for Moving a SystemInvasion of Malignant Cells (hypoxic environment triggers invasion of cells)

Y. pseudotuberculosisinvasin

In Conclusion

We now have the tools to build new and exciting devices within biological cells

Where we can construct new parts, new devices, and new systems

We can build on previous work in Synthetic Biology

Develop novel uses for this technology (Medical applications)

Share with others through iGEM

Thanks!