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Cell Programming: Implementing Logical Functions Using Genetic Information

Explore the innovative concept of programming cells to implement configurable Boolean logic functions in 2 variables and compile larger circuits via intercellular signaling. Learn about the biological background, gene expression mechanisms, and design of regulatory networks for artificial gene regulation. Discover methodologies, practical significance, and biological realizations through genetic manipulation, transcription, translation, and regulatory networks.

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Cell Programming: Implementing Logical Functions Using Genetic Information

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  1. The Programming of a Cell By L Varin and N Kharma Biology and Computer Engineering Departments Concordia University

  2. Motivation • Cells have advantages over silicon: • They have/can have built-in interfaces, to sense and produce many biological substances • They are easy to mass produce, store and distribute • They are generally more robust than man-made systems • They are optimizable via (real) evolution The Programming of a Cell

  3. Motivation – precisely • The aim is to produce a cell that implements a configurable Boolean logic function in 2 var’s • Ultimately, we would like to use intercellular signalling to compile larger circuits using many smaller ones A mechanical AND gate The Programming of a Cell

  4. Outline • Motivation & Outline • Biological Background • Problem Statement • Alternative Methods • Biological Realization • Practical Significance The Programming of a Cell

  5. Biological Background:Flow of genetic information DNA RNA Protein We can easily manipulate DNA Gene expression Transcription Translation The Programming of a Cell

  6. Biological Background:Gene expression (promoters) +1 RNA Pol -10 Box TATAA -35 Box TTGTCA RNA Core promoter = Binding site for RNA Polymerase In this configuration transcription is ON The Programming of a Cell

  7. Biological Background:Gene expression (promoters) +1 X R -10 box -35 box operator R = Repressor In this configuration RNA Polymerase cannot bind transcription is OFF The Programming of a Cell

  8. Biological Background:Gene expression (synthetic gene) Modular structure • Construct a promoter • Insert an operator • Select a coding sequence (output) Output -10 box -35 box operator The Programming of a Cell

  9. Biological Background:Biological regulatory network • The lactose operon of E. coli lacI repressor X R -35 O -10 Transcription is OFF R Active repressor The Programming of a Cell

  10. Biological Background:Biological regulatory network • The lactose operon of E. coli lacI repressor RNA Pol -35 O -10 X Transcription is ON R Inactive repressor = inducer (lactose) The Programming of a Cell

  11. Biological Background:Artificial regulatory network • Select an output gene • Select a promoter • Select an operator-repressor system • Assemble the parts together The Programming of a Cell

  12. Biological Background:Artificial regulatory network - lactose X OFF lacI l C1 repressor Repressed by lac repressor Lac promoter ON l promoter Green Fluorescent protein Repressed by C1 repressor The Programming of a Cell

  13. Biological Background:Artificial regulatory network + lactose R X l C1 repressor Repressed by lac repressor Lac promoter C1 X C1 l promoter Green Fluorescent protein Repressed by C1 repressor OFF The Programming of a Cell

  14. Problem Statement ++ • Synthesize a cell that can be configured to implement any one of 16 different Boolean functions in 2 variables • Such a project will involve 4 phases: 1 Designing a regulatory network 2 Constructing a configurable cell 3 Configuring the cell 4 Using the cell The Programming of a Cell

  15. Methodology 1: Using Repressilators A B Output R1 Anti-sense DNA R2 The Programming of a Cell

  16. A B’ A B A’ B A’ B’ Output Output Output Output R5 R1 R7 R3 Anti-sense DNA Anti-sense DNA Anti-sense DNA Anti-sense DNA R2 R8 R4 R6 Methodology 1: Full Picture The Programming of a Cell

  17. Methodology 2: Using Excision • A Boolean function in 2 variables has 16 possible truth tables • They all involve 1-4 (3) different terms of 2 variables A B Output 00 ? 01 ? 11 ? 10 ? The Programming of a Cell

  18. Methodology 2: Chosen Path • Design A regulatory network implementing the 4 terms and allowing for subsequent excision of any term • Construct The regulatory network by embedding 4 gene networks corresponding to the 4 terms in a real organism (e.g. e.coli) • Configure The cell by excising those gene networks corresponding to the unwanted terms • Use The configured cell by adding the inducers (variables) it is designed to respond to, and monitoring the output The Programming of a Cell

  19. Biological Realization • 2 variables A and B • A = lactose • B = arabinose • 1 promoter • 4 repressors • 1 ouput gene (Green Fluorescent Protein) • 4 terms (A B), (A B), (A B), (A B) The Programming of a Cell

  20. Biological Realization Output (GFP) (A B) = Lac operon operator (bound by LacI repressor) = Arabinose operon operator (bound by AraR repressor) In the absence of A and B X LacI AraR Output (GFP) The Programming of a Cell

  21. Biological Realization (A B) LacI AraR X Output (GFP) In the presence of A and B ( lactose and arabinose) LacI AraR X X Output (GFP) The Programming of a Cell

  22. Biological Realization Output (GFP) (A B) = l Pr operator (bound by C1 repressor) = Arabinose operon operator (bound by AraR repressor) X LacI C1 In the absence of A and in presence of B AraR X Output (GFP) The Programming of a Cell

  23. Biological Realization Output (GFP) (A B) = lactose operon operator (bound by lacI repressor) = l Prm operator (bound by CRO repressor) X AraR CRO In the presence of A and in absence of B LacI X Output (GFP) The Programming of a Cell

  24. Biological Realization Output (GFP) (A B) = l Pr operator (bound by C1 repressor) X AraR CRO = l Prm operator (bound by CRO repressor) X LacI C1 In the absence of A and in absence of B Output (GFP) The Programming of a Cell

  25. Biological Realization Output (GFP) (A B) (A B) (A B) (A B) Output (GFP) Output (GFP) Output (GFP) The Programming of a Cell

  26. Practical Significance Limited ConfigurableDecision Logic Inputs tied to a particular application: lactose, arabinose etc. Outputs tied to a particular application: GFP etc. The Programming of a Cell

  27. Practical Significance Application-specific outputs Application-specific inputs Input Interface Output Interface Extended Decision Logic Standardized Signals The Programming of a Cell

  28. Summary • A specific Boolean logic function in 5 variables has been recently realized in living cells, but never a configurable bio-logic device  theoretic value • We believe we have found a simple means of realizing a configurable 2-input Boolean function in an e.coli cell simple methodology • Both the logic functionality and the practical value of the work can be considerably enhanced with the use of intercellular signaling  broader vision • First experiments (for the Method 2) will start in January 2008  and we’ll update you! The Programming of a Cell

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