title: A polygalacturonase-inhibiting protein from grapevine reduces the symptoms of the endopolygalacturonase BcPG2 from Botrytis cinerea in Nicotiana benthamiana leaves without any evidence for in vitro interaction.
abstract: Six endopolygalacturonases from Botrytis cinerea (BcPG1 to BcPG6) as well as mutated forms of BcPG1 and BcPG2 were expressed transiently in leaves of Nicotiana benthamiana using agroinfiltration. Expression of BcPG1, BcPG2, BcPG4, BcPG5, and mutant BcPG1-D203A caused symptoms, whereas BcPG3, BcPG6, and mutant BcPG2-D192A caused no symptoms. Expression of BcPG2 caused the most severe symptoms, including wilting and necrosis. BcPG2 previously has been shown to be essential for B. cinerea virulence. The in vivo effect of this enzyme and the inhibition by a polygalacturonase-inhibiting protein (PGIP) was examined by coexpressing Bcpg2 and the Vvpgipl gene from Vitis vinifera in N. benthamiana. Coinfiltration resulted in a substantial reduction of the symptoms inflicted by the activity of BcPG2 in planta, as evidenced by quantifying the variable chlorophyll fluorescence yield. In vitro, however, no interaction between pure VvPGIP1 and pure BcPG2 was detected. Specifically, VvPGIP1 neither inhibited BcPG2 activity nor altered the degradation profile of polygalacturonic acid by BcPG2. Furthermore, using surface plasmon resonance technology, no physical interaction between VvPGIP1 and BcPG2 was detected in vitro. The data suggest that the in planta environment provided a context to support the interaction between BcPG2 and VvPGIP1, leading to a reduction in symptom development, whereas neither of the in vitro assays detected any interaction between these proteins.
authors.affiliation: Institute for Wine Biotechnology, Department of Viticulture and Oenology, Stellenbosch University, Stellenbosch, 7600, South Africa.
authors.forename: Dirk A
authors.initials: DA
authors.lastname: Joubert
authors.forename: Ilona
authors.initials: I
authors.lastname: Kars
authors.forename: Lia
authors.initials: L
authors.lastname: Wagemakers
authors.forename: Carl
authors.initials: C
authors.lastname: Bergmann
authors.forename: Gabré
authors.initials: G
authors.lastname: Kemp
authors.forename: Melané A
authors.initials: MA
authors.lastname: Vivier
authors.forename: Jan A L
authors.initials: JA
authors.lastname: van Kan
mesh_terms: D020171:Botrytis / Q000201:enzymology* / Q000235:genetics; D002734:Chlorophyll / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism*; D005453:Fluorescence; D005656:Fungal Proteins / Q000378:metabolism*; D010935:Plant Diseases / Q000382:microbiology*; D018515:Plant Leaves / Q000382:microbiology*; D010940:Plant Proteins / Q000378:metabolism*; D030821:Plants, Genetically Modified; D011096:Polygalacturonase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics / Q000382:microbiology*; D027843:Vitis / Q000737:chemistry* / Q000235:genetics
chemical_list: D004791:Enzyme Inhibitors; D005656:Fungal Proteins; C081479:PGIP protein, plant; D010940:Plant Proteins; D002734:Chlorophyll; D011096:Polygalacturonase