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A strategy for building an amplified transcriptional switch to detect bacterial contamination of plants

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Abstract

We have designed and tested a transcriptional autofeedback loop that could be used to engineer plants to sense the presence of bacteria. The signal amplification circuit was built based on the biological switch responsive to the presence of bacterial flagellin. Several flagellin- and E. coli-inducible Arabidopsis promoters were cloned and tested in transient expression assays in Arabidopsis and lettuce protoplasts using a flagellin-based peptide. These were investigated either as direct drivers of a reporter gene, or as a component of a transcriptional autofeedback loop. Arabidopsis promoters from the xyloglucan endotransglucosylase/hydrolase 18 (ATXTH18) and cytochrome P450 family CYP82C3 monooxygenase worked well as biological switches. These promoters were incorporated into our feedback loop system for signal amplification. The inclusion of a transcriptional repressor reduced basal expression, thereby increasing fold-amplification of signal detection and fine-tuning the positive autofeedback loop regulation.

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Acknowledgments

We thank Christopher Defraia for Arabidopsis growth protocols and Donna Williams for help with the confocal microscope. This project was funded by grant W911SR-07-C-0084 to WBG and EC from the Florida Biodefense Research Consortium.

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Authors and Affiliations

  1. Department of Microbiology and Cell Science, Program of Plant Molecular and Cellular Biology, University of Florida, Bldg. 981, 1052 Museum Road, P.O. Box 110700, Gainesville, FL, 32611-0700, USA

    Eva Czarnecka, F. Lance Verner & William B. Gurley

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  1. Eva Czarnecka
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  2. F. Lance Verner
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  3. William B. Gurley
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Correspondence to Eva Czarnecka or William B. Gurley.

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Czarnecka, E., Verner, F.L. & Gurley, W.B. A strategy for building an amplified transcriptional switch to detect bacterial contamination of plants. Plant Mol Biol 78, 59–75 (2012). https://doi.org/10.1007/s11103-011-9845-2

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