Pseudomonas syringae HrpP Is a Type III Secretion Substrate Specificity Switch Domain Protein That Is Translocated into Plant Cells but Functions Atypically for a Substrate-Switching Protein

Pseudomonas syringae delivers virulence effector proteins into plant cells via an Hrp1 type III secretion system (T3SS).P. syringae pv. tomato DC3000 HrpP has a C-terminal, putative T3SS substrate specificity switch domain, likeYersinia YscP. A ΔhrpP DC3000 mutant could not cause disease in tomato or elicit a hypersensitive response (HR) in tobacco, but the HR could be restored by expression of HrpP intrans . Though HrpP is a relatively divergent protein in the T3SS of differentP. syringae pathovars,hrpP fromP. syringae pv. syringae 61 andP. syringae pv. phaseolicola 1448A restored HR elicitation and pathogenicity to DC3000 ΔhrpP . HrpP was translocated intoNicotiana benthamiana cells via the DC3000 T3SS when expressed from its native promoter, but it was not secreted in culture. N- and C-terminal truncations of HrpP were tested for their ability to be translocated and to restore HR elicitation activity to the ΔhrpP mutant. No N-terminal truncation completely abolished translocation, implying that HrpP has an atypical T3SS translocation signal. Deleting more than 20 amino acids from the C terminus abolished the ability to restore HR elicitation. HrpP fused to green fluorescent protein was no longer translocated but could restore HR elicitation activity to the ΔhrpP mutant, suggesting that translocation is not essential for the function of HrpP. No T3SS substrates were detectably secreted by DC3000 ΔhrpP except the pilin subunit HrpA, which unexpectedly was secreted poorly. HrpP may function somewhat differently than YscP because theP. syringae T3SS pilus likely varies in length due to differing plant cell walls.