Kinney A J, Jung R, Herman E M
DuPont Nutrition and Health, P.O. Box 80402, Experimental Station, Wilmington, Delaware 19880-0402, USA.
Plant Cell. 2001 May;13(5):1165-78. doi: 10.1105/tpc.13.5.1165.
The expression of the alpha and alpha' subunits of beta-conglycinin was suppressed by sequence-mediated gene silencing in transgenic soybean seed. The resulting seeds had similar total oil and protein content and ratio compared with the parent line. The decrease in beta-conglycinin protein was apparently compensated by an increased accumulation of glycinin. In addition, proglycinin, the precursor of glycinin, was detected as a prominent polypeptide band in the protein profile of the transgenic seed extract. Electron microscopic analysis and immunocytochemistry of maturing transgenic soybean seeds indicated that the process of storage protein accumulation was altered in the transgenic line. In normal soybeans, the storage proteins are deposited in pre-existing vacuoles by Golgi-derived vesicles. In contrast, in transgenic seed with reduced beta-conglycinin levels, endoplasmic reticulum (ER)-derived vesicles were observed that resembled precursor accumulating-vesicles of pumpkin seeds and the protein bodies accumulated by cereal seeds. Their ER-derived membrane of the novel vesicles did not contain the protein storage vacuole tonoplast-specific protein alpha-TIP, and the sequestered polypeptides did not contain complex glycans, indicating a preGolgi and nonvacuolar nature. Glycinin was identified as a major component of these novel protein bodies and its diversion from normal storage protein trafficking appears to be related to the proglycinin buildup in the transgenic seed. The stable accumulation of proteins in a protein body compartment instead of vacuolar accumulation of proteins may provide an alternative intracellular site to sequester proteins when soybeans are used as protein factories.
在转基因大豆种子中,通过序列介导的基因沉默抑制了β-伴大豆球蛋白α和α'亚基的表达。与亲本系相比,所得种子的总油和蛋白质含量及比例相似。β-伴大豆球蛋白蛋白的减少显然被大豆球蛋白积累的增加所补偿。此外,大豆球蛋白的前体前大豆球蛋白在转基因种子提取物的蛋白质谱中被检测为一条突出的多肽带。对成熟转基因大豆种子的电子显微镜分析和免疫细胞化学表明,转基因系中储存蛋白积累的过程发生了改变。在正常大豆中,储存蛋白通过高尔基体衍生的小泡沉积在预先存在的液泡中。相比之下,在β-伴大豆球蛋白水平降低的转基因种子中,观察到内质网(ER)衍生的小泡,类似于南瓜种子的前体积累小泡和谷物种子积累的蛋白体。这些新型小泡的内质网衍生膜不含蛋白储存液泡液泡膜特异性蛋白α-TIP,且隔离的多肽不含复合聚糖,表明其具有高尔基体前和非液泡的性质。大豆球蛋白被鉴定为这些新型蛋白体的主要成分,其从正常储存蛋白运输途径的转移似乎与转基因种子中前大豆球蛋白的积累有关。当大豆用作蛋白质工厂时,蛋白质在蛋白体区室中的稳定积累而非在液泡中的积累可能为隔离蛋白质提供了一个替代的细胞内位点。