Otegui Marisa S, Noh Yoo-Sun, Martínez Dana E, Vila Petroff Martin G, Staehelin L Andrew, Amasino Richard M, Guiamet Juan J
Instituto de Fisiología Vegetal (INFIVE), Universidad Nacional de La Plata, c.c. 327, 1900, La Plata, Argentina.
Plant J. 2005 Mar;41(6):831-44. doi: 10.1111/j.1365-313X.2005.02346.x.
Vacuolar compartments associated with leaf senescence and the subcellular localization of the senescence-specific cysteine-protease SAG12 (senescence-associated gene 12) were studied using specific fluorescent markers, the expression of reporter genes, and the analysis of high-pressure frozen/freeze-substituted samples. Senescence-associated vacuoles (SAVs) with intense proteolytic activity develop in the peripheral cytoplasm of mesophyll and guard cells in Arabidopsis and soybean. The vacuolar identity of these compartments was confirmed by immunolabeling with specific antibody markers. SAVs and the central vacuole differ in their acidity and tonoplast composition: SAVs are more acidic than the central vacuole and, whereas the tonoplast of central vacuoles is highly enriched in gamma-TIP (tonoplast intrinsic protein), the tonoplast of SAVs lacks this aquaporin. The expression of a SAG12-GFP fusion protein in transgenic Arabidopsis plants shows that SAG12 localizes to SAVs. The analysis of Pro(SAG12):GUS transgenic plants indicates that SAG12 expression in senescing leaves is restricted to SAV-containing cells, for example, mesophyll and guard cells. A homozygous sag12 Arabidopsis mutant develops SAVs and does not show any visually detectable phenotypical alteration during senescence, indicating that SAG12 is not required either for SAV formation or for progression of visual symptoms of senescence. The presence of two types of vacuoles in senescing leaves could provide different lytic compartments for the dismantling of specific cellular components. The possible origin and functions of SAVs during leaf senescence are discussed.
利用特异性荧光标记、报告基因表达以及高压冷冻/冷冻置换样品分析，研究了与叶片衰老相关的液泡区室以及衰老特异性半胱氨酸蛋白酶SAG12（衰老相关基因12）的亚细胞定位。在拟南芥和大豆的叶肉细胞和保卫细胞的外周细胞质中，形成了具有强烈蛋白水解活性的衰老相关液泡（SAVs）。通过用特异性抗体标记进行免疫标记，证实了这些区室的液泡特性。SAVs和中央液泡在酸度和液泡膜组成上存在差异：SAVs比中央液泡酸性更强，而且，中央液泡的液泡膜高度富集γ-TIP（液泡膜内在蛋白），而SAVs的液泡膜缺乏这种水通道蛋白。SAG12-GFP融合蛋白在转基因拟南芥植物中的表达表明，SAG12定位于SAVs。对Pro（SAG12）：GUS转基因植物的分析表明，衰老叶片中SAG12的表达仅限于含有SAVs的细胞，例如叶肉细胞和保卫细胞。纯合的拟南芥sag12突变体形成SAVs，并且在衰老过程中没有表现出任何视觉上可检测到的表型改变，这表明SAVs的形成或衰老视觉症状的进展都不需要SAG12。衰老叶片中两种类型液泡的存在可能为特定细胞成分的拆解提供不同的溶酶区室。本文讨论了叶片衰老过程中SAVs可能的起源和功能。
