Expression of a polyphemusin variant in transgenic tobacco confers resistance against plant pathogenic bacteria, fungi and a virus.

Apurva Bhargava; Milan Osusky; Benjamin Forward; Robert Hancock; William Kay; Santosh Misra

Abstract??Antimicrobial cationic peptides provide a promising means of engineering plant resistance to a range of plant pathogens, including viruses. PV5 is a synthetic structural variant of polyphemusin, a cationic peptide derived from the horseshoe crab-Limulus polyphemus. PV5 has been shown to be benign toward eukaryotic membranes but with enhanced antimicrobial activity against animal pathogens. In this work, the cytotoxicity of PV5 toward tobacco protoplasts and leaf discs was assessed using TTC (2,3,5-triphenyltetrazolium chloride) and Evans blue colorimetric assays. PV5 showed no measurable cytotoxic effects even at levels as high as 60??g. As a possible approach to enhancing plant resistance, a gene encoding PV5 was fused to the signal sequence encoding the C-terminus portion of the BiP protein fromPseudotsuga menziesii, under the control of 2???35S CaMV promoter. When introduced intoNicotiana tabacumvar Xanthi gene integration and expression was confirmed by both Southern and northern analyses. When transgenic plants were subsequently challenged with bacterial and fungal phytopathogens enhanced resistance was observed. Moreover, transgenic plants also displayed antiviral properties against Tobacco Mosaic Virus making PV5 an excellent candidate for conferring unusually broad spectrum resistance to plants and the first anti-plant virus antimicrobial peptide.

TRANSGENIC plants; TOBACCO; PHYTOPATHOGENIC bacteria; LIMULUS polyphemus

Plant Cell, Tissue & Organ Culture, 2007, Vol 88, Issue 3, p301

0167-6857

Academic Journal

10.1007/s11240-007-9204-9