Molecular analysis of secretion genes located on the syr-syp genomic island of Pseudomonas syringae pv. syringae strain B301D

Abstract

An RND (resistance-nodulation-cell division) transporter, called the PseC protein, was identified at the left border of the syr-syp genomic island of Pseudomonas syringae pv. syringae strain B301D. The PseC protein exhibited amino acid homology to a putative RND transporter of Ralstonia solanacearum with identities of 61% (i.e., PseC). The pseC mutant strain showed a larger reduction in syringopeptin secretion (67%) than syringomycin secretion (41%). A β-glucuronidase assay with a pseA::uidA reporter construct indicated that the GacS/A two-component system controls expression of the pseA gene. Expression of the sypA gene by mutant strain B301D-HK4 corresponded to approximately 13% of that by parental strain B301D, whereas the syrB1 gene expression by mutant strain B301D-HK4 was nearly 61%. Mutant strain B301D-HK4 was reduced in virulence by about 58% as compared to parental strain B301D. A drug-supersensitive acrB mutant of E. coli showed increased resistance to acriflavine and tetracycline upon heterologous expression of the pseA, pseB, and pseC genes. Thus, the PseC protein, an RND transporter, has an important role in secretion of syringomycin and syringopeptin. An ATP-binding cassette (ABC) transporter, called the PseF protein, was identified at the left border of the syr-syp genomic island. The PseF protein exhibited amino acid homology to a putative ABC transporter of E. coli W3104 with identities of 57.6% (i.e., PseF to MacB). The pseF mutant strain showed significant reduction in secretion of syringomycin (74%) and syringopeptin (71%). Expression of the sypA gene by mutant strain B301D-HK7 was approximately 6.9% as compared to that of parental strain B301D, while the syrB1 gene expression by mutant strain B301D-HK7 was nearly 14.6%. Mutant strain B301D-HK7 was less virulent by approximately 67% than parental strain B301D. Expression of the pseF gene was induced approximately six times by strain B301D grown on SRMAF, as compared to that of strain B301D grown on SRM. During infection of bean plants by P. syringae pv. syringae strain B728a, expression of the pseF gene increased 3 days after inoculation. Thus, the PseF protein, an ABC transporter, responsible for secretion of syringomycin and syringopeptin is required for full virulence.