An engineered promoter driving expression of a microbial avirulence gene confers recognition of TAL effectors and reduces growth of diverse Xanthomonas strains in citrus

Xanthomonas citri subsp. citri (X. citri), causal agent of citrus canker, uses Transcription Activator-Like Effectors (TALEs) as major pathogenicity factors. TALEs, which are delivered into plant cells through the type III secretion system (T3SS), interact with Effector Binding Elements (EBEs) in host genomes to activate expression of downstream susceptibility genes to promote disease. Predictably, TALEs bind EBEs in host promoters via known combinations of TALE amino acids to DNA bases known as the TALE code. We introduced 14 EBEs, matching to distinct X. citri TALEs, into the promoter of the pepper Bs3 gene (ProBs31EBE ) and fused this engineered promoter with multiple EBEs (ProBs314EBE ) to either the GUS reporter gene or the coding sequence (cds) of the pepper gene, Bs3. TALE induced expression of the Bs3 cds in citrus leaves resulted in no visible HR. Therefore, we utilized a different approach in which ProBs31EBE and ProBs314EBE were fused to the Xanthomonas gene, avrGf1, which encodes a bacterial effector that elicits a hypersensitive response (HR) in grapefruit and sweet orange. We demonstrate in transient assays that activation of ProBs314EBE by X. citri TALEs is T3SS dependent and that expression of AvrGf1 triggers HR and correlates with reduced bacterial growth. We further demonstrated that all tested virulent X. citri strains from diverse geographic locations activated ProBs314EBE . TALEs are essential for virulence of X. citri strains and, because the engineered promoter traps are activated by multiple TALEs, this concept has potential to confer broad-spectrum, durable resistance to citrus canker in stably transformed plants. This article is protected by copyright. All rights reserved.