Julius B. Lucks/Bibliography/Kobayashi-PNAS-101-2004

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Notes on [1]

  • couple engineered gene networks to cells natural regulatory circuitry
    • E. coli genetic toggle switch interfaced with
      • SOS signaling for DNA damage
      • transgenic quorum sensing path from Vibrio fischeri
  • engineer cells to form biofilms in response to DNA damage
  • engineer cells to activate protein synthesis when pop reaches critical density
  • synthetic biol refs (1,2)
  • gene circuits for memory storage and logic operations (19-22) [2],[3]
    • most designed to respond to external stimuli
  • should we really design the first language implementation modularly? Perhaps the goal of modularity is too complex for now - best to get a monolithic design (like the linux kernel), but a small one done first, while research on modularity continues.
    • in contrast to this paper's belief that should generate 'plug-and-play' solutions
      • modular approach aimed at eventually constructing general components for constructing programmable cells
  • most work in bio-computing that I have heard of concentrate on treating the cell as a programmable circuit board at best. Thus to solve a computational problem, you have to construct a "hardware" circuit that solves that problem. To me this does not mean programmable. I have in mind something more along the lines of an implementation that involves a mini-kernel + memory device + language interpreter, all wrapped up into one (for the time being), that you can write code for. To "program" this first simple system would be to do something like write assembly code. Eventually this system could be modularized by the real implementers out there to consist of a stack that is (bottom to top) system devices : kernel : interpreter, much like a modern CPU architecture. Biocomputing "languages" of the future would generate the low level "bio-assembly" code that the first systems would use as its primary language. High-level languages could be built upon the low-level languages, and the history of the modern computer can be played out again, with the biological cell at the bottom rather than the silicon semi-conductor. Obviously there are lots of challenges here.
  • simplest plug-and-play programmable cell has 3 parts
    1. Biosensor module (senses input signal)
    2. engineered regulatory network (performs the computation)
    3. output interface (converts computation output into a biological response we can 'see')
  • in this work, a toggle switch is used for number 2, and interfaced with the signaling pathways listed above
  • GFP is quantifiable biological response
  • elements
    • regulators: pTMSa, pTMSb1, pTMSb2 (toggle switches)
    • sensors: SOS pathway, pAHLa/b (AHL inducible plasmid - quorum)
    • outputs: plasmid pCIRa/b (GFP reporters), plasmid pBFR (biofilm formation)
  • full description of genes and plasmids in supporting info
  • 1st step in engineering gene network into a cell is to understand network's dynamic properties
  • toggle switch used uses lacI and lambda CI - lacI epressed from modified lamda PL promoter (repressed by cI), cI expressed from Ptrc (repressed by lacI) - see (4) and fig 2A for details
    • RecA is know to cleave cI, so can trigger a flip of the switch by triggering RecA - this is how to integrate this with the DNA damage response (which triggers RecA)
    • To get an output, put whatever output response want in front of PL* so that it is repressed when cI active, and activated when cI chewed up by RecA - see Figure 3
  • switch can flip by noise induced transition (fluctuations in repressor conc. for example) - (48,49)
    • this causes bimodal population of cells
      • this genetic toggle robust - noise transitions rare (4)
      • mathematical analysis in suppl info - transition can be flipped by increase cI decay rate (recA - see (51)), or LacR basal synthesis rate (LuxR induced by AHL (quorum sensing) - binds to P_LuxI - put lacI downstream of this - see Fig 5)
  • this toggle prevents flopping back once flipped (no way to start synthisis of cI once repressed) - later generations 'remember' having incurred DNA damage in a previous generation
  • 2 other DNA damage detecting papers - (37,38)
  • use a biofilm 'readout' of this circuit by putting relevant traA gene in front of PL* - get same readout of DNA damage (Fig 4)
  • AHL - gram negative bacterial signal to coordinate activity (29-31)
  • could program population control by replacing GFP reporter with a killer gene (34)
  • switch that is turned on at high pop density useful for industrial production of growth-inhibiting product - only turns on when enough cells to produce it effectively
  • 'In many cases, the properties of the system must be optimized rather than those of individual components.' - see supplementary info
    • suggest selected evolution to get novel dynamical behaviours (54,55)


  1. Kobayashi H, Kaern M, Araki M, Chung K, Gardner TS, Cantor CR, and Collins JJ. Programmable cells: interfacing natural and engineered gene networks. Proc Natl Acad Sci U S A. 2004 Jun 1;101(22):8414-9. DOI:10.1073/pnas.0402940101 | PubMed ID:15159530 | HubMed [Kobayashi-PNAS-101-2004]
  2. Simpson ML, Sayler GS, Fleming JT, and Applegate B. Whole-cell biocomputing. Trends Biotechnol. 2001 Aug;19(8):317-23. PubMed ID:11451474 | HubMed [Simpson-TrendsBiotechnol-19-2001]


  3. Weiss R., Basu S., Hooshangi S., Kalmbach A., Karig D., Mehreja R., Netravali I. Genetic circuit building blocks for cellular computation, communications, and signal processing. PDF [Weiss-NatComput-2-2003]
All Medline abstracts: PubMed | HubMed