High-level expression of apple PGIP1 is not sufficient to protect transgenic potato against Verticillium dahliae

Abstract Polygalacturonase-inhibiting proteins (PGIPs) are plant proteins believed to play a role in the defence against pathogenic fungi. Purified apple PGIP1 inhibited polygalacturonases (PGs) secreted by Verticillium dahliae grown on potato root cell walls and pectin. We therefore hypothesised that apple PGIP1 could be used to confer resistance against Verticillium -wilt, a major disease of potato caused by the fungus V. dahliae . Transgenic lines containing the apple pgip1 gene under control of the enhanced CaMV 35S (e35S) promoter were generated. Stable integration of the apple pgip1 transgene into the potato genome was shown by the polymerase chain reaction (PCR) and Southern blot. Cross hybridisation with potato pgip(s) was not observed. High-level expression of the apple PGIP1 in several independent transgenic potato events was verified by silver staining of SDS-PAGE separated proteins and Western blot. All but one of the PGIP1 extracts prepared from the transgenic potato lines were successful in inhibiting V. dahliae PGs. Active PGIP1 was expressed in the leaves as well as the roots of the transgenic plants. The apoplastic localisation of PGIP activity in the pgip -transgenic potato plants was demonstrated by a vacuum infiltration–extraction experiment. A glasshouse trial indicated that six transgenic lines (A10, B10, B13, A3, A14 and B16) had significantly reduced disease symptoms compared to the untransformed control and other lines when grown in the inoculated soil, but five of them also showed significantly slower senescence symptoms when grown in the control soil. It is proposed that an extended juvenile phase in the transgenic lines resulted in the apparent increased disease resistance, and this could not be attributed to inhibition of V. dahliae PGs, despite high-level expression of the apple PGIP1.