|Artlip, Timothy - Tim|
|Norelli, John (jay) - Jay|
Submitted to: Physiologia Plantarum
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/1/2003
Publication Date: 11/1/2003
Citation: WISNIEWSKI, M.E., BASSETT, C.L., ARTLIP, T.S., WEBB, R.P., JANISIEWICZ, W.J., NORELLI, J.L., GOLDWAY, M., DROBY, S. CHARACTERIZATION OF A DEFENSIN IN BARK & FRUIT TISSUES OF PEACH AND IN VITRO ANTIMICROBIAL ACTIVITY OF A RECOMBINANT DEFENSIN IN THE YEAST PICHIA. PLANT MOLECULAR BIOLOGY. NOVEMBER 2003, Vol. 119, pgs 563-572. Interpretive Summary: To survive, all living things must be able to protect themselves successfully from environmental extremes and from attack by other organisms. In order to accomplish this, many plants and animals have developed suites of proteins to assist in defending themselves against drastic changes in temperature and against diseases. We previously reported the isolation of a seasonally-expressed protein (dehydrin) with antifreeze properties from peach bark. In this paper we report the isolation and characterization of another seasonally-expressed protein, called defensin, from peach bark. Like other members of this protein family, the peach defensin shows antifungal activity; in this case specifically inhibiting the growth of fungi that cause fruit to rot after harvest. Identifying and understanding the regulation of proteins that protect plants from potentially hazardous conditions is critical to the development of strategies aimed at improving the yield and quality of fruits.
Technical Abstract: Considerable research in our laboratory has focused on identifying seasonally-regulated proteins and their genes in peach (Prunus persica [L.] Batsch) that may be related to cold hardiness. In the present study, we describe the cloning and characterization of a defensin gene (PpDfn1) from a cDNA library made from winter bark tissues. A partial clone obtained from the library was extended to full length by 5' RACE. The open reading frame of 237 bp codes for a 79 amino acid peptide related to the defensin family of proteins. Sequence comparison of the encoded protein using BLAST analysis revealed significant homology to defensins from other plant species. RNA gel blot analysis indicated that the gene is seasonally-expressed in bark tissues of one-year-old shoots and is also expressed in early fruit development. Quantitative RT-PCR and protein blot analysis largely paralleled the RNA gel blot analyses for the bark tissues. A recombinant version, rDFN1 was expressed in the yeast, Pichia pastoris. It was found that rDFN1 inhibited germination of the fungal pathogens Penicillium expansum and Botrytis cinerea, but not the Gram-negative bacterium Erwinia amylovora. The potential physiological role of PpDFN1 and its antimicrobial properties are discussed.