MHZ3调控水稻乙烯信号转导的分子机制研究

The ethylene signal transduction pathway has been established in model dicotyledonous plant Arabidopsis. However, the signaling mechanisms in monocotyledonous plants such as rice remain largely unknown. Recently, we have isolated a set of ethylene-response mutants of rice (mhz). Functional studies on some of the genes revealed that ethylene signaling in rice has both conserved and diverged mechanisms. In this study, we will characterize the mhz3 mutant, which shows ethylene insensitivity in both coleoptiles and roots of etiolated seedlings. Map-based cloning revealed that MHZ3 code for a function-unknown membrane protein. MHZ3 expression is induced by ethylene at both transcriptional and protein levels. Genetic interactions between MHZ3 and ethylene signaling components indicated that MHZ3 may acts at the level of OsEIN2. The purpose of this study is to identify and characterize the molecular mechanism of MHZ3 in regulating ethylene signaling on the base of the following researches: 1) determine the target site of MHZ3 in ethylene signaling pathway by further analysis of genetic interactions; 2) investigate the potential roles of MHZ3 in regulating OsEIN2 function by cell biology and biochemical analysis; 3) identify MHZ3 interacting proteins and its regulatory mechanisms using biochemical and genetic approaches. This study should provide clues for better understanding of ethylene signaling mechanisms by characterization of a novel regulatory component of the signaling pathway.

目前有关乙烯信号转导的研究主要集中在拟南芥等双子叶植物，人们对单子叶植物中乙烯作用机制的了解还十分有限。近年来我们分离鉴定了一系列水稻乙烯反应突变体(mhz)。对其中一些基因的功能研究发现，水稻乙烯信号转导既有保守的机制又有不同于拟南芥的新的机制。本项目将对mhz3进行深入研究。该突变体表现为胚芽鞘和根对乙烯完全不敏感。MHZ3编码一个功能未知的膜蛋白，在转录和蛋白水平受乙烯诱导。与已知信号组分遗传互作初步研究显示，MHZ3可能作用在中心组分OsEIN2水平。本项目将通过进一步的遗传互作分析确认MHZ3在乙烯信号通路中的作用位置；运用细胞生物学和生物化学等手段探究MHZ3对OsEIN2可能的调控作用；通过生化和遗传分析鉴定MHZ3互作蛋白及其调控机制。综合上述研究解析MHZ3调控乙烯信号转导的分子机制。本研究有望鉴定一个新的乙烯信号调控组分，为进一步完善水稻乙烯信号转导通路提供线索

本项目鉴定了乙烯信号通路的一个新组分MHZ3。该基因编码一个功能未知的内质网膜蛋白，其表达在转录和蛋白水平都受到乙烯诱导。遗传分析发现，MHZ3在乙烯信号通路中特异地作用于OsEIN2。其功能缺失导致OsEIN2蛋白稳定性降低，并阻碍乙烯诱导的OsEIN2蛋白积累；而MHZ3过量表达则显著提高OsEIN2蛋白丰度。进一步研究发现，MHZ3与OsEIN2蛋白直接互作，其N端和C端都结合在OsEIN2 Nramp-like结构域上。二者互作能够有效抑制OsEIN2泛素化，是稳定OsEIN2蛋白所必须的。综上，乙烯诱导的MHZ3蛋白可能通过结合OsEIN2 Nramp-like结构域来抑制其泛素化从而稳定OsEIN2蛋白以诱发乙烯反应。这一研究揭示了EIN2通过其N端Nramp-like结构域感知上游乙烯信号的潜在机制，进一步完善了乙烯信号转导通路。