Comparative Genomics Identifies a Novel Conserved Protein, HpaT, in Proteobacterial Type III Secretion Systems that Do Not Possess the Putative Translocon Protein HrpF.

is the causal agent of bacterial leaf streak, the most common bacterial disease of wheat and barley. To cause disease, most xanthomonads depend on a highly conserved type III secretion system, which translocates type III effectors into host plant cells. Mutagenesis of the conserved type III secretion gene confirmed that the type III secretion system is required to cause disease on the host plant barley and to trigger a non-host hypersensitive response (HR) in pepper leaves. Type III effectors are delivered to the host cell by a surface appendage, the Hrp pilus, and a translocon protein complex that inserts into the plant cell plasma membrane. Homologs of the HrpF protein, including PopF from and NolX from rhizobia, are thought to act as a translocon protein. Comparative genomics revealed that strains harbor a noncanonical gene cluster, which rather shares features with type III secretion systems from , , , and than other spp. Surprisingly, none of these bacteria, except , encode a homolog of the HrpF translocon. Here, we aimed at identifying a candidate translocon from . Notably, genomes from strains that lacked // instead encode another gene, called , adjacent to and co-regulated with the type III secretion system gene cluster. An insertional mutant in the gene, which is the first gene of a two-gene operon, -, was non-pathogenic on barley and did not cause the HR or programmed cell death in non-host pepper similar to the mutant. The mutant phenotypes were partially complemented by either or the downstream gene, , which has been described as a facilitator of translocation in . Interestingly, the mutant was also complemented by the gene from . These findings reveal that both HpaT and HpaH contribute to the injection of type III effectors into plant cells.