Identification of Two Novel hrp -Associated Genes in the hrp Gene Cluster of Xanthomonas oryzae pv. oryzae

Abstract

We have cloned a hrp gene cluster from Xanthomonas oryzae pv. oryzae. Bacteria with mutations in the hrp region have reduced growth in rice leaves and lose the ability to elicit a hypersensitive response (HR) on the appropriate resistant cultivars of rice and the nonhost plant tomato. A 12,165-bp portion of nucleotide sequence from the presumed left end and extending through the hrpB operon was determined. The region was most similar to hrp genes from Xanthomonas campestris pv. vesicatoria and Ralstonia solanacearum. Two new hrp-associated loci, named hpa1 and hpa2, were located beyond the hrpA operon. The hpa1 gene encoded a 13-kDa glycine-rich protein with a composition similar to those of harpins and PopA. The product of hpa2 was similar to lysozyme-like proteins. Perfect PIP boxes were present in the hrpB and hpa1 operons, while a variant PIP box was located upstream of hpa2. A strain with a deletion encompassing hpa1 and hpa2 had reduced pathogenicity and elicited a weak HR on nonhost and resistant host plants. Experiments using single mutations in hpa1 and hpa2 indicated that the loss of hpa1 was the principal cause of the reduced pathogenicity of the deletion strain. A 1,519-bp insertion element was located immediately downstream of hpa2. Hybridization with hpa2 indicated that the gene was present in all of the strains of Xanthomonas examined. Hybridization experiments with hpa1 and IS1114 indicated that these sequences were detectable in all strains of X. oryzae pv. oryzae and some other Xanthomonas species.The hrp ("harp") genes encode type III secretory pathways and are required by many phytopathogenic bacteria to elicit a hypersensitive response (HR) on nonhost or resistant host plants and for pathogenesis on susceptible hosts. The HR is a rapid localized death of the host cells that occurs upon pathogen infection and, together with the expression of a complex array of defense-related genes, is a component of plant resistance. The hrp genes were first identified in Pseudomonas syringae pv. phaseolicola, a bean pathogen (38). Since then, hrp genes from a variety of plant pathogenic bacteria, including Erwinia, Pseudomonas, Ralstonia, and Xanthomonas, have been characterized (for reviews, see references 2, 9, and 11). The specific functions of the hrp pathway in pathogenesis are not known. However, type III secretion pathways of animal and plant pathogens have been demonstrated to mediate the secretion of virulence factors into the extracellular melieu. Some of the proteins ultimately end up in the host cell cytoplasm (reviewed in references 11, 25, and 37). In mediating the interaction of the bacterium and the host plant, the hrp pathway presumably acts to prevent or inhibit a general resistance response or otherwise enhance the colonization of the plant by the bacteria.Given the importance of the type III systems to pathogenicity, it can be expected that analysis of the systems in different species will provide insight into the adaptation of the species to their respective host plants. The hrp gene cl...

Expression of hpa1 Gene Encoding a Bacterial Harpin Protein in Xanthomonas oryzae pv. oryzae Enhances Disease Resistance to Both Fungal and Bacterial Pathogens in Rice and Arabidopsis

“…They are secreted from bacteria through type III secretion system and are known to be located at intercellular space of plant tissues (Alfano and Collmer, 1997). Harpin proteins have been identified from diverse Gram-negative plantpathogenic bacteria such as Erwinia amylovora (Kim and Beer, 1998;Wei et al, 1992), Pseudomonas syringae (Charkowski et al, 1998;He et al, 1993;Kvitko et al, 2007), Ralstonia solanacearum (Arlat et al, 1994), and Xanthomonas species (Kim et al, 2003;Noel et al, 2002;Zhu et al, 2000). For example, Hpa1 is the harpin protein that is produced and secreted from X. oryzae pv.…”

Molecular determinants of disease and resistance in interactions of Xanthomonas oryzae pv. oryzae and rice

“…vesicatoria hrp region as a probe, an X. oryzae pv. oryzae hrp region has also been isolated [7]. The 12.2-kb DNA fragment covers the region corresponding to hrpA, hrpB, hrpC and hrpD of X. campestris pv.…”

“…Complementation of the Tn5 mutants with the wild-type hrp region restored the ability to cause disease on rice and to elicit HR on tomato, indicating that hrp gene-encoded TTSS is also required for the pathogenicity and HR of X. oryzae pv. oryzae [7].…”

“…oryzae pv. oryzae interaction, with the cloning of the two rice resistance (R) genes, Xa1 and Xa21 [4,5], the bacterial avirulence (avr) genes, avrXa5, avrXa7 and avrXa10 [6], the hypersensitive response and pathogenicity (hrp) genes [7] and many other virulencerelated genes [8][9][10][11][12][13][14]. Studies on the pathogen population structure using molecular genotyping methods have provided information on pathogen evolution.…”

Host and Pathogen Factors Controlling the Rice-Xanthomonas oryzae Interaction

“…1). Xanthomonas oryzae pv oryzae (Xoo) is the causal agent and a member of the g-proteobacteria and, like many other proteobacteria, depends, in part, on a type III secretion system for pathogenicity (Swings et al, 1990;Zhu et al, 2000). The bacterium invades the xylem tissue, either through wounds or water pores, leading to systemic infection.…”