Improved Resistance to Cucumber mosaic virus in Petunia Transformed with Non‐Cytotoxic Pokeweed Antiviral Protein Gene

Abstract

Petunia hybrida is an important ornamental plant that can be seriously affected by cucumber mosaic virus (CMV). Pokeweed antiviral protein (PAP), a ribosome‐inactivating protein, has been recognized as a broad spectrum virus inhibitory agent. Mutant PAP efficiently inhibited viral gene expression at both the translational and transcriptional levels without causing host cell toxicity. We have transferred the non‐cytotoxic pokeweed antiviral protein (mutant PAP) gene into petunia cells with Agrobacterium tumefaciens. Forty‐two putative transgenic regenerated lines were obtained from the selected explants. Fifty‐six plants immune to CMV infection were recovered from nine transgenic lines. Another 51 plants tolerant to CMV were obtained from 10 transgenic regenerated lines. Successful integration of the mutant PAP gene into the genome of transgenic petunia was confirmed by PCR and Southern blot analysis. Expression of the PAP gene was further confirmed by RT‐PCR and Western blot analysis. These results were consistent with the assay of resistance to CMV.

Plant-derived ribosome-inactivating proteins involved in defense against plant viruses

“…have been the most studied antiviral proteins in transgenic development, mainly PAP I, PAP II, PhRIP I, Pac-PAP, and PIP. The PAP has been successfully overexpressed in different transgenic plants, which lead to improved resistance against multiple plant viruses such as CMV, PLRV, PVX, PVY, TMV, and TuMV (Di & Tumer, 2005;Fu et al, 2000;Li et al, 2013;Lodge et al, 1993;Moon et al, 1994;Smirnov et al, 1997;Tumer et al, 1997;Zhang et al, 1999;Zhao et al, 2008;Zoubenko et al, 2000). Lodge et al (1993) introduced PAP into tobacco and potato plants which enabled transgenic plants to develop resistance to a broad-spectrum of plant viruses, and the resistance was also effective against both mechanically and aphid-transmitted viruses.…”

“…However, it has been reported that high levels of wild-type or variant PAP (PAPv) expression in transgenic plants was also toxic to plant cells (Lodge et al, 1993). The non-toxic PAP mutants (PAPc, PAPn) have also been efficiently expressed in transgenic plants and induced resistance to virus infections, but not toxic to plants harbouring the transgene (Li et al, 2013;Tumer et al, 1997;Zoubenko et al, 2000). The transgenic expression of PAP II in tobacco also exhibited broadspectrum resistance to PVX and TMV, as well as a fungal pathogen.…”

Agrobacterium-mediated transformation of ornamental species: A review

“…Removing a specific adenine base from the large ribosomal RNA, PAP acts as a ribosome inactivating protein. LI et al (2013) have transferred modified PAP gene into petunia cells with A. tumefaciens and obtained transgenic plants immune or highly resistant to Cucumber mosaic virus (CMV).…”