Type I-F CRISPR-PAIR platform for multi-mode regulation to boost extracellular electron transfer in Shewanella oneidensis
Yaru Chen,
Meijie Cheng,
Hao Song,
Yingxiu Cao
Affiliations
Yaru Chen
Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin 300072, China
Meijie Cheng
Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin 300072, China
Hao Song
Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin 300072, China; Corresponding author
Yingxiu Cao
Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin 300072, China; Corresponding author
Summary: Bio-electrochemical systems are based on extracellular electron transfer (EET), whose efficiency relates to the expression level of numerous genes. However, the lack of multi-functional tools for gene activation and repression hampers the enhancement of EET in electroactive microorganisms (EAMs). We thus develop a type I-F CRISPR/PaeCascade-RpoD-mediated activation and inhibition regulation (CRISPR-PAIR) platform in the model EAM, Shewanella oneidensis MR-1. Gene activation is achieved (3.8-fold) through fusing activator RpoD (σ70) to Cas7 when targeting the prioritized loci upstream of the transcription start site. Gene inhibition almost has no position preference when targeting the open reading frame, which makes the design of crRNAs easy and flexible. Then CRISPR-PAIR platform is applied to up-/down-regulate the expression of six endogenous genes, resulting in the improved EET efficiency. Moreover, simultaneous gene activation and inhibition are achieved in S. oneidensis MR-1. CRISPR-PAIR platform offers a programmable methodology for dual regulation, facilitating in-depth EET studies in Shewanella spp.
