Multiple vacuolar sorting determinants exist in soybean 11S globulin
Multiple vacuolar sorting determinants exist in soybean 11S globulin
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DOI：
10.1105/tpc.105.036376
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发表时间：
2006-05-01
期刊：
影响因子：
11.6
通讯作者：
Utsumi, S
中科院分区：
文献类型：
--
作者：
Maruyama, N;Mun, LC;Utsumi, S
通过瞬时表达测定法在成熟大豆子叶中研究了大豆球蛋白（大豆11 S球蛋白）的分选决定因素，该球蛋白介导蛋白质靶向蛋白质储存液泡（PSV）。A1 aB 1b（一种大豆球蛋白I组亚基）的10个氨基酸的C-末端延伸足以将绿色荧光蛋白（GFP）导向PSV。该肽可以对应于C-末端空泡分选决定簇（ctVSD）。由于A1 aB 1b的这种推定ctVSD的功能抑制并不阻止A1 aB 1b的PSV分选，我们使用A1 aB 1b的三维结构来鉴定序列特异性决定簇（ssVSD）的候选者。我们发现，无序区4的下游序列可以将GFP引导至PSV，并且Ile-297对于分选至关重要。然而，ctVSD的功能抑制，结合Ile 297 Gly突变，并没有取消A1 aB 1b的空泡分选，这表明A1 aB 1b除了ctVSD和ssVSD外还有第三个分选决定簇。大豆球蛋白组II亚基，A3 B4，缺乏ctVSD，但含有VSD想起ssVSD和一个额外的排序决定子。我们还表明，通过表达小GTP酶和药物治疗实验的显性阴性突变体，A1 aB 1b的贩运是COPII囊泡依赖和渥曼青霉素和布雷菲德菌素A敏感。
The sorting determinants of glycinin, a soybean (Glycine max) 11S globulin, which mediates protein targeting to the protein storage vacuole (PSV), were investigated in maturing soybean cotyledons by transient expression assays. A C-terminal stretch of 10 amino acids of A1aB1b, a glycinin group I subunit, was sufficient to direct green fluorescent protein (GFP) to the PSV. This peptide may correspond to a C-terminal vacuolar sorting determinant (ctVSD). Because functional inhibition of this putative ctVSD of A1aB1b did not block PSV sorting of A1aB1b, we used the three-dimensional structure of A1aB1b to identify candidates for a sequence-specific determinant (ssVSD). We found that the sequence downstream of disordered region 4 could direct GFP to the PSV and that Ile-297 is critical for sorting. However, functional inhibition of the ctVSD, combined with the Ile297Gly mutation, did not abolish the vacuolar sorting of A1aB1b, suggesting that A1aB1b has a third sorting determinant in addition to ctVSD and ssVSD. A glycinin group II subunit, A3B4, lacked a ctVSD but contained a VSD reminiscent of an ssVSD and an additional sorting determinant. We also demonstrate, by expression of dominant negative mutants of small GTPases and drug treatment experiments, that the trafficking of A1aB1b is COPII vesicle-dependent and wortmannin- and brefeldin A-sensitive.