CHARACTERIZATION OF THE HRPJ AND HRPU OPERONS OF PSEUDOMONAS-SYRINGAEPV SYRINGAE PSS61 - SIMILARITY WITH COMPONENTS OF ENTERIC BACTERIA INVOLVED IN FLAGELLAR BIOGENESIS AND DEMONSTRATION OF THEIR ROLE IN HARPIN(PSS) SECRETION

The hrp/hrmA gene cluster of Pseudomonas syringae pv. syringae Pss61 h as been shown to form a minimum genetic unit sufficient to enable nonp athogenic bacteria, such as Escherichia coli, to elicit the hypersensi tive response associated with disease resistance. The biochemical func tions of most of these genes have not been established. The nucleotide sequence of a 4.3-kb SstI-Bg/II fragment carrying hrp apparent transl ational units V, VI, and VII revealed one partial open reading frame ( ORF) and five complete ORFs producing 35,126-, 48,866-, 17,308-, 20,48 2-, and 26,363-Da gene products (hrpJ3, J4, J5, U1, U2, respectively). The production of these proteins was confirmed by using T7 RNA polyme rase-directed expression. The partial ORF was found to be identical to the C terminus of HrpJ2, The absence of apparent transcriptional term inators and promoters between hrpI (hrpJ2), hrpJ3, hrpJ4, and hrpJ5 to gether with the observation that the HrpL-dependent hrpJ promoter dire cts expression of hrpJ3-J5 indicates that these genes form a single op eron controlled by the HrpL-dependent hrpJ promoter. A second HrpL-dep endent promoter consensus sequence was also identified upstream of hrp U1 and demonstrated to function as a HrpL-dependent promoter, thus ind icating that hrpU1, hrpU2, and additional downstream genes may be part of a second operon. The deduced product of hrpJ3 exhibits similarity to FliG of Salmonella typhimurium, a cytoplasmic protein that regulate s flagellar rotation and biogenesis. HrpJ4 shares extensive similarity with the FliI family of ATPase-like proteins and retains the known fu nctional domains conserved among this family of proteins. HrpJ5 has pr operties similar to the S. typhimurium FliJ. Neither HrpU1 nor HrpU2 e xhibit significant similarity to known proteins. Secretion of Harpin(P ss) by E. coli ICIC4100 transformants carrying pHIR11::TnphoA derivati ves was blocked in hrpJ4, J5, and U2 mutants. In view of the previousl y reported similarity of HrpJ2 to the LcrD superfamily that includes F lhA, these results predict that the gene products of the hrpJ and hrpU operons form an inner membrane complex for translocation of proteins similar to that used by the flagellar biogenesis system of S. typhimur ium.