Programmable cells: Interfacing natural and engineered gene networks

Hideki Kobayashi, Mads Kærn, Michihiro Araki, Kristy Chung, Timothy S. Gardner, Charles R. Cantor, and James J. Collins

• create novel cellular behaviors and characteristics by coupling engineered gene networks to the cell’s natural regulatory circuitry

• Four examples• Detects and retains memory of DNA damage• Forms biofilm in response to DNA damage• Detects and retains memory of quorum sensing

molecules• Density dependent protein synthesis

Scope

Flow Cytometer (BD FcsCalibur)

Reporter: GFP

lacI -> lacR -> Ptrc -> cI -> cI -> PL

lacI -> lacR -> Ptrc -> cI -> cI -> PL

MMC – 15 hUV – 1-10 s

(With traA gene) (lacking the traA gene)

Replacegfp with traA

Biofilm was only observed if traA was expressed for > 4h

LuxI-LuxR quorum-sensing systems

acylated-homoserine lactone (AHL)

luxICDABEluxR

LuxR LuxI

Quorum sensing molecules

lacI -> lacR -> Ptrc -> cI -> cI -> PL

Density-dependent gene activation

Toggle

lacI -> lacR -> Ptrc -> cI -> cI -> PL

GFP

Discussions

• Programmable cells have been constructed by interfacing natural and engineered gene networks

• Programmable cells have been demonstrated using four different constructs with the toggle gene network as a building block

Something to think about…

DNA damage sensing

b)

Original design

Will the modified design work? If not, why not? If yes, how would it differ from (a)?

Modified design

Density-dependent gene activation

Q: What if we replace the lacI gene with gfp and forget about the regulatory circuit

gfp