Elicitation of hypersensitive cell death by extracellularly targeted HrpZ(Psph) produced in planta

The ability of the Pseudomonas syringae pv. phaseolicola harpin (HrPZ(Psph) ) to elicit hypersensitive response was investigated in three Nicotiana gen otypes. The hrpZ(Psph) gene was placed under chemical regulation (tetracycl ine induction) in TetR(+) Nicotiana tabacum cv. Wisconsin 38 (W38) or was t ransiently expressed in N. benthamiana following infection with a PVX-deriv ed vector and in three Nicotiana genotypes by agroinfiltration. The constru cts were designed to express either the canonical form of harpin (HrpZ(Psph )) or an N-terminally extended version of the protein carrying the signal p eptide portion of the tobacco pathogenesis-related protein PR1a (SP-HrPZ(Ps ph)). Stable transformants of N. tabacum cv. W38 did not develop necrosis u pon induction with tetracycline, probably as a result of insufficient harpi n accumulation. In contrast, N. benthamiana plants infected with the PVX co nstructs produced high concentrations of harpin in biologically active form , but only those expressing the secretable form of harpin developed necroti c symptoms. These symptoms were less severe than those caused by PVX::avrPt o; however, they were accompanied by induction of hsr203J, a hypersensitive response-specific gene transcript. These results suggest that the plant ce llular receptor(s) for harpin is extracellular.