State Key Laboratory of Plant Physiology and Biochemistry, College of Life Science, Zhejiang University, Hangzhou 310058, PR China.
Plant Cell Physiol. 2010 Mar;51(3):475-85. doi: 10.1093/pcp/pcq012. Epub 2010 Jan 22.
The stromal processing peptidase (SPP) is a metalloendopeptidase that cleaves a broad range of precursor substrates. In this study, we isolated a rice mutant showing leaf chlorosis at the early seedling stage but inhibition of root growth during the whole growth period. Genetic analysis demonstrates that the phenotypes of the mutant were caused by a recessive single gene mutation. The mutated gene was cloned by map-based cloning, and was shown to encode an SPP. Sequence analysis showed a glutamate deletion in the highly conserved C-terminus of SPP in the mutant. The mutation of SPP in the mutant was verified by transgenic complementation. SPP is constitutively expressed in all tissues. Subcellular localization analysis indicates that SPP is targeted to the chloroplast. The expression of some genes associated with chloroplast development was decreased in young seedlings of the spp mutant, but not in 14-day-old seedlings. Western blot analysis revealed that the Rubisco small subunit is not precisely processed in the spp mutant in 7-day-old seedlings, but the processing activity in the spp mutant is restored in 14-day-old seedlings. Moreover, the expression levels of Cab1R and Cab2R for the light-harvesting chlorophyll a/b-binding protein (LHCP) were highly up-regulated in the transgenic plants with overexpression of SPP. The present results reveal that SPP is essential for chloroplast biogenesis at the early growth stage and for rice root development; this is the first report on the function of SPP in monocot plants.
基质蛋白酶(SPP)是一种金属内肽酶,可切割广泛的前体底物。在这项研究中,我们分离出一个水稻突变体,该突变体在幼苗早期表现出叶片黄化,但整个生长期间根生长受到抑制。遗传分析表明,突变体的表型是由隐性单基因突变引起的。通过基于图谱的克隆克隆了突变基因,并证实其编码 SPP。序列分析显示突变体中 SPP 的高度保守 C 末端缺失了一个谷氨酸。通过转基因互补验证了突变体中 SPP 的突变。SPP 在所有组织中均持续表达。亚细胞定位分析表明 SPP 定位于叶绿体。在 spp 突变体的幼苗中,一些与叶绿体发育相关的基因的表达降低,但在 14 天大的幼苗中没有。Western blot 分析显示,在 7 天大的 spp 突变体中 Rubisco 小亚基不能精确加工,但在 14 天大的 spp 突变体中加工活性得到恢复。此外,过表达 SPP 的转基因植物中光捕获叶绿素 a/b 结合蛋白(LHCP)的 Cab1R 和 Cab2R 的表达水平高度上调。本研究结果表明,SPP 对于早期生长阶段的叶绿体生物发生和水稻根发育是必需的;这是 SPP 在单子叶植物中的功能的首次报道。