Identification of the Transcription Factors RAP2-13 Activating the Expression of CsBAK1 in Citrus Defence Response to Xanthomonas citri subsp. citri

Abstract

Citrus canker is a quarantined disease caused by the bacterial plant pathogen Xanthomonas citri subsp. citri (Xcc), which causes persistent surface damage, leaf and fruit drop, and tree decline in citrus plants. The citrus cultivar Citron C-05 (Citrus medica L.) is a disease-resistant genotype identified after years of screening at the National Center for Citrus Improvement (Changsha), which displays allergic, necrotic, and disease-resistant responses to Xcc. In this study, the BAK1 gene was identified in this cultivar to be a disease resistance gene involved in plant-microbe interaction between citrus and Xcc. Functional investigations of this gene revealed that both CsBAK1 (C. sinensis BAK1) or CmBAK1(C. medica BAK1) could inhibit the growth of Xcc to some extent when transiently expressed in the susceptible ‘Bingtang’ genotype of sweet orange. Critical regions of the CmBAK1 promoter sequence were identified by creating downstream deletions and exposing mutants to Xcc to determine effects on the resistance phenotype; a 426 bp region (−2000~–1574) was identified as a key functional region responsible for eliciting the hypersensitive response in plants. Through screening arrayed Citron C-05 cDNA libraries by yeast one-hybrid assays, a basic APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) transcription factor of CmRAP2-13 that binds directly to the 426 bp key sequence and activates expression of CmBAK1 was identified. Moreover, transcriptional analysis revealed an obvious increase in transcript levels of CsRAP2-13 in Citron C-05, American citron, and Finger citron. In this study, we present the identification of transcriptional activators that are found to interact with BAK1 proteins in response to Xcc. These results reveal a coordinated regulatory mechanism of RAP2-13, which may be involved in defence responses through the regulation of BAK1.