Deng Zhiping, Zhang Xin, Tang Wenqiang, Oses-Prieto Juan A, Suzuki Nagi, Gendron Joshua M, Chen Huanjing, Guan Shenheng, Chalkley Robert J, Peterman T Kaye, Burlingame Alma L, Wang Zhi-Yong
Department of Plant Biology, Carnegie Institution of Washington, Stanford, California 94305, USA.
Mol Cell Proteomics. 2007 Dec;6(12):2058-71. doi: 10.1074/mcp.M700123-MCP200. Epub 2007 Sep 11.
The plant steroid hormones brassinosteroids (BRs) play an important role in a wide range of developmental and physiological processes. How BR signaling regulates diverse processes remains unclear. To understand the molecular details of BR responses, we performed a proteomics study of BR-regulated proteins in Arabidopsis using two-dimensional DIGE coupled with LC-MS/MS. We identified 42 BR-regulated proteins, which are predicted to play potential roles in BR regulation of specific cellular processes, such as signaling, cytoskeleton rearrangement, vesicle trafficking, and biosynthesis of hormones and vitamins. Analyses of the BR-insensitive mutant bri1-116 and BR-hypersensitive mutant bzr1-1D identified five proteins (PATL1, PATL2, THI1, AtMDAR3, and NADP-ME2) affected both by BR treatment and in the mutants, suggesting their importance in BR action. Selected proteins were further studied using insertion knock-out mutants or immunoblotting. Interestingly about 80% of the BR-responsive proteins were not identified in previous microarray studies, and direct comparison between protein and RNA changes in BR mutants revealed a very weak correlation. RT-PCR analysis of selected genes revealed gene-specific kinetic relationships between RNA and protein responses. Furthermore BR-regulated posttranslational modification of BiP2 protein was detected as spot shifts in two-dimensional DIGE. This study provides novel insights into the molecular networks that link BR signaling to specific cellular and physiological responses.
植物甾体激素油菜素甾醇（BRs）在广泛的发育和生理过程中发挥着重要作用。BR信号如何调节多种过程仍不清楚。为了了解BR反应的分子细节，我们利用二维差异凝胶电泳（DIGE）结合液相色谱-串联质谱（LC-MS/MS）对拟南芥中受BR调控的蛋白质进行了蛋白质组学研究。我们鉴定出42种受BR调控的蛋白质，预计它们在BR对特定细胞过程的调控中发挥潜在作用，如信号传导、细胞骨架重排、囊泡运输以及激素和维生素的生物合成。对BR不敏感突变体bri1-116和BR超敏感突变体bzr1-1D的分析鉴定出5种蛋白质（PATL1、PATL2、THI1、AtMDAR3和NADP-ME2），它们在BR处理和突变体中均受到影响，表明它们在BR作用中的重要性。使用插入敲除突变体或免疫印迹对选定的蛋白质进行了进一步研究。有趣的是，约80%的BR反应蛋白在先前的微阵列研究中未被鉴定出来，并且BR突变体中蛋白质和RNA变化之间的直接比较显示出非常弱的相关性。对选定基因的RT-PCR分析揭示了RNA和蛋白质反应之间基因特异性的动力学关系。此外，在二维DIGE中检测到BiP2蛋白的BR调控的翻译后修饰表现为斑点迁移。这项研究为将BR信号与特定细胞和生理反应联系起来的分子网络提供了新的见解。
