Oxygen-responsive genetic circuits constructed in Synechocystis sp. PCC 6803.
As photoautotrophic prokaryotes, cyanobacteria are promising platforms for producing value-added bioproducts. However, few regulatory genetic parts and devices (e.g., inducible promoters and regulatory circuits) have been developed for these potential hosts. Furthermore, the devices that have been created respond only to a single input. To address these issues, we developed an inducible genetic circuit that generates heterologous proteins in response to oxygen, an environmental signal. To test its performance and utility in Synechocystis sp. PCC 6803, a model cyanobacterial strain, we connected this circuit to either heterologous nifHDK genes, which encode oxygen-sensitive nitrogenase's structural proteins, or a fluorescent protein gene. The circuit was transcriptionally activated to generate nifHDK transcripts or fluorescent output only in low oxygen conditions. We expanded the oxygen-responsive circuit into a more complex circuit by building a two-input AND gate, which allows Synechocystis to specifically control expression of the fluorescent reporter in response to two signals, low oxygen and high anhydrotetracycline. To our knowledge, the AND gate is the first complex logic circuit built in a cyanobacterial strain. This work expands the synthetic biology tools available for complex gene expression in cyanobacteria, increasing their potential as biotechnology platforms.
Department of Energy, Environmental and Chemical Engineering, Washington University in St. Louis, St. Louis, Missouri.
Cheryl M
CM
Immethun
Department of Energy, Environmental and Chemical Engineering, Washington University in St. Louis, St. Louis, Missouri.
Kenneth M
KM
Ng
Department of Energy, Environmental and Chemical Engineering, Washington University in St. Louis, St. Louis, Missouri.
Drew M
DM
DeLorenzo
Department of Energy, Environmental and Chemical Engineering, Washington University in St. Louis, St. Louis, Missouri.
Ben
B
Waldron-Feinstein
Department of Energy, Environmental and Chemical Engineering, Washington University in St. Louis, St. Louis, Missouri.
Ying-Chiang
YC
Lee
Department of Energy, Environmental and Chemical Engineering, Washington University in St. Louis, St. Louis, Missouri. tsmoon@wustl.edu.
Tae Seok
TS
Moon
D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D017930:Genes, Reporter; D005827:Genetics, Microbial / Q000379:methods*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008967:Molecular Biology / Q000379:methods*; D009591:Nitrogenase / Q000235:genetics / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism*; D046939:Synechocystis / Q000187:drug effects* / Q000235:genetics*; D058615:Synthetic Biology / Q000379:methods; D014158:Transcription, Genetic; D015533:Transcriptional Activation / Q000187:drug effects*
D008164:Luminescent Proteins; D009591:Nitrogenase; D010100:Oxygen
cyanobacteria; genetic circuit; genetic context; oxygen sensor; synthetic biology