Fig 5. Effect of mutations of AvrPto1_Psy acetylation sites on PsyB728a growth in tomato.

(A) AvrPto1_Psy_T91A and H125A/H130A mutants did not reduce PsyB728a growth in PTO-containing tomato in the absence of HopZ3. Plants were spray-inoculated with indicated strains at an OD₆₀₀ = 0.01. Epiphytic bacterial populations were quantified in leaf disc washes from eight different plants per strain four days after inoculation. Different letters indicate significant differences in growth as assessed by ANOVA with Tukey’s test (P<0.05). Similar results were found in at least two other experiments (AvrPto1_Psy_H125A/H130A did not reduce the growth of PsyB728a ΔHopZ3 in three out of five experiments). Bars indicate standard errors. V, vector control; A1, AvrPto1_Psy; Z3, HopZ3; T91, AvrPto1_Psy_T91A; H125/H130 and HH, AvrPto1_Psy_H125A/H130A. (B) H125A/H130A mutation of AvrPto1_Psy did not affect its binding to PTO. AvrPto1_Psy-GST, AvrPto1_Psy_H125A/H130A-GST and PTO-MBP were expressed in E. coli. Purified soluble AvrPto1_Psy or H125A/H130A mutant was pulled down with immobilized PTO-MBP or, alternatively, soluble PTO was pulled down with immobilized AvrPto1_Psy-GST or AvrPto1_Psy_H125A/H130A-GST. Band intensities were quantified from seven experiments. (T-test, P = 0.1). (C) AvrPto1 mutant variants were expressed to similar levels as AvrPto1_Psy in ΔAvrPto1 and ΔhopZ3ΔAvrPto1 PsyB728a grown in type III secretion-inducing conditions.