Brassica napus polygalacturonase inhibitor proteins inhibit Sclerotinia sclerotiorum polygalacturonase enzymatic and necrotizing activities and delay symptoms in transgenic plants

Sclerotinia sclerotiorum releases a battery of polygalacturonases (PGs) during infection, which the host plant may cope with through production of polygalacturonase inhibitor proteins (PGIPs). To study the interaction between S. sclerotiorum PGs and Brassica napus PGIPs, 5 S. sclerotiorum PGs and 4 B. napus PGIPs were expressed in Pichia pastoris. SsPG3, SsPG6, and BnPGIP1 were successfully produced in the yeast system, and BnPGIP1 inhibited SsPG6 enzymatic activity in vitro. SsPG3 and SsPG6 both induced light-dependent necrosis when infiltrated into leaves, which was reduced in an Arabidopsis thaliana line expressing BnPGIP2 and to a lesser extent in a line expressing BnPGIP1. The line expressing BnPGIP2 also exhibited a delay in the onset of symptoms upon S. sclerotiorum inoculation, but no long-term effect on S. sclerotiorum disease progression was observed. The P. pastoris system was found to be suitable for expressing high levels of some S. sclerotiorum PGs, but PGIP interaction studies were best performed in planta. Arabidopsis thaliana forms necrotic lesions upon infiltration of PGs, is susceptible to S. sclerotiorum, and is easily transformed, and thus, is well-suited for the qualitative study of PG-PGIP interactions.