Function And Regulation Analyses Of The Hrp Cluster Of Xanthomonas Campestris Pv.Campestris

Xanthomonas campestris pv. campestris (Xcc) is the causal agent of black rot disease of crucifers worldwide. In both plant and mammalian Gram-negative pathogenic bacteria, type III secretion systems (TTSS), encoded by hrp cluster, play a crucial role in interactions with the host. The proteins assembled TTSS were highly conserved. Nevertheless, each bacterium also possesses variable Type III proteins unique, inhomogeneous or conserved only in limited systems. In order to investigate the function of hrp genes of Xcc, the mutants of Xcc 8004 were generated by transposon Tn5gusA5. A series of hrp genes were mutated. The phenotypes of the hrp mutants, such as pathogenicity, production of exopolysaccharides and extracellular enzymes, were determined. The hrp or hrc mutants lose or reduce the virulence, while the pathogenicity of hpa mutants was nearly not affected. Furthermore, the hrp cluster deletion mutant of Xcc 8004 was generated through marker exchange, namely Xcc 8004â–³27. The hrpG integration mutants of Xcc 8004 â–³27 was generated with pT18mob using as homologous nonreplicating plasmid. Strangely, Xcc 8004A27, which contains no hrp cluster, can still invade the host plants. The hrp cluster and hrpG double mutants lose the pathogenicity completely. We speculate that there were somenegative regulators in the hrp cluster of Xcc 8004, which collaborate with other pathogenicity factors. To study the regulation of hrp gene cluster of Xcc, the nonpolar hrpG integration mutants upon those hrp mutant strains, which Tn5 was inserted in ORF transcription direction, were generated. The GUS activities were mensurated in the plant-induced medium XVM2 before and after the hrpG mutagenesis. As a result, hrp genes of Xcc 8004, including hrpX, hpa2, hrpB1, hrcV, hrpD6, conserved hypothetical genes and hrpF, were all in the hrpG regulon.