J Integr Plant Biol. ›› 2020, Vol. 62 ›› Issue (8): 1065-1079.DOI: 10.1111/jipb.12891

Special Issue: Biotechnology

• New Technology • Previous Articles     Next Articles

Split Nano luciferase complementation for probing protein-protein interactions in plant cells

Feng-Zhu Wang1†, Nannan Zhang1,2†, Yan-Jun Guo1†, Ben-Qiang Gong1 and Jian-Feng Li1*   

  1. 1Guangdong Provincial Key Laboratory of Plant Resources, State Key Laboratory of Biocontrol, MOE Key Laboratory of Gene Function and Regulation, School of Life Sciences, Sun Yat‐sen University, Guangzhou 510275, China
    2Guangdong Provincial Key Laboratory of Sugarcane Improvement and Biorefinery, Guangdong Bioengineering Institute, Guangzhou 510316, China

    These authors contributed equally to this work.
    *Correspondence: Jian-Feng Li(lijfeng3@mail.sysu.edu.cn)
  • Received:2019-11-02 Accepted:2019-11-15 Online:2019-11-22 Published:2020-08-01


Deciphering protein‐protein interactions (PPIs) is fundamental for understanding signal transduction pathways in plants. The split firefly luciferase (Fluc) complementation (SLC) assay has been widely used for analyzing PPIs. However, concern has risen about the bulky halves of Fluc interfering with the functions of their fusion partners. Nano luciferase (Nluc) is the smallest substitute for Fluc with improved stability and luminescence. Here, we developed a dual‐use system enabling the detection of PPIs through the Nluc‐based SLC and co‐immunoprecipitation assays. This was realized by coexpression of two proteins under investigation in fusion with the HA‐ or FLAG‐tagged Nluc halves, respectively. We validated the robustness of this system by reproducing multiple previously documented PPIs in protoplasts or Agrobacterium‐transformed plants. We next applied this system to evaluate the homodimerization of Arabidopsis CERK1, a coreceptor of fungal elicitor chitin, and its heterodimerization with other homologs in the absence or presence of chitin. Moreover, split fragments of Nluc were fused to two cytosolic ends of Arabidopsis calcium channels CNGC2 and CNGC4 to help sense the allosteric change induced by the bacterial elicitor flg22. Collectively, these results demonstrate the usefulness of the Nluc‐based SLC assay for probing constitutive or inducible PPIs and protein allostery in plant cells.

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