Jayasankar Subramanian, Li Zhijian, Gray Dennis J
Mid-Florida Research and Education Center, Institute of Food Agricultural Sciences University of Florida, 2725Binion Road, Apopka, FL 32703, USA. Current address: Department of Plant Agriculture - Vineland Campus, University of Guelph, 4890VictoriaAvenue N, PO Box 7000, Vineland Station, Ontario, Canada L0R 2E0.
Mid-Florida Research and Education Center, Institute of Food Agricultural Sciences University of Florida, 2725Binion Road, Apopka, FL 32703, USA.
Funct Plant Biol. 2003 Jan;30(11):1105-1115. doi: 10.1071/FP03066.
Anthracnose-resistant grapevine (Vitis vinifera L. cv. Chardonnay) plants were regenerated from embryogenic cultures that had been subjected to in vitro selection with culture filtrate of Elsinoe ampelina (de Bary) Shear. Three secreted proteins differentially expressed by in vitro-selected embryogenic cultures and regenerated plants were identified. An 8-kDa protein was identified as a lipid-transfer protein (LTP) by N-terminal amino acid sequence comparison. Two other differentially expressed proteins, with estimated molecular weights of 21.6 and 22kDa, immuno-reacted with antiserum raised against a thaumatin-like protein (TLP) protein from pinto bean (Phaseolus vulgaris L.). N-terminal amino acid sequencing of the 21.6-kDa protein showed a high degree of homology to V. vinifera thaumatin-like protein 2 (VVTL-2 = grapevine osmotin; Acc no. CAA71883), and that of the 22-kDa protein was homologous to V. vinifera thaumatin-like protein 1 (VVTL-1; AAB61590). Interestingly, both VVTL-1 and VVTL-2 are pathogenesis-related (PR) proteins, belonging to the PR-5 group. Protein produced from the cloned grapevine VVTL-1 gene significantly inhibited E. ampelina spore germination and hyphal growth in vitro. Plants regenerated from in vitro-selected cultures similarly inhibited fungal growth in vivo. Enhanced expression of antifungal VVTL-1 in anthracnose resistant grapevine strongly suggests that it plays an important role, either alone or in conjunction with other PR proteins, by suppressing pathogen growth.
通过用葡萄痂圆孢（Elsinoe ampelina (de Bary) Shear）的培养滤液进行体外筛选的胚性培养物，再生出了抗炭疽病的葡萄（Vitis vinifera L. cv. Chardonnay）植株。鉴定了三种在体外筛选的胚性培养物和再生植株中差异表达的分泌蛋白。通过N端氨基酸序列比较，一种8 kDa的蛋白被鉴定为脂质转移蛋白（LTP）。另外两种差异表达的蛋白，估计分子量分别为21.6 kDa和22 kDa，与针对菜豆（Phaseolus vulgaris L.）的类甜蛋白（TLP）产生的抗血清发生免疫反应。21.6 kDa蛋白的N端氨基酸测序显示与葡萄类甜蛋白2（VVTL-2 = 葡萄渗透素；登录号CAA71883）高度同源，22 kDa蛋白的N端氨基酸测序与葡萄类甜蛋白1（VVTL-1；AAB61590）同源。有趣的是，VVTL-1和VVTL-2都是病程相关（PR）蛋白，属于PR-5组。从克隆的葡萄VVTL-1基因产生的蛋白在体外显著抑制葡萄痂圆孢孢子萌发和菌丝生长。从体外筛选培养物再生的植株在体内同样抑制真菌生长。抗炭疽病葡萄中抗真菌VVTL-1的表达增强强烈表明，它通过抑制病原体生长单独或与其他PR蛋白一起发挥重要作用。
