PHD锌指蛋白GmPHD6调控大豆耐盐的分子机制研究

Soybean (Glycine max) is an important crop. Abiotic stress is one of the main reasons that cause the drop in soybean production, and it is thus significant to improve the stress tolerance of soybean. Through comparing gene differential expression between stress tolerant and sensitive soybean, we identified a small group of genes encoding PHD zinc-finger proteins. These genes only exist in plant and they respond to different stresses. Most members of this group have DNA binding ability and transcription regulatory activity, except for a special one, GmPHD6. GmPHD6 does not has the transcription regulatory activity. Using a transgenic hairy root system, we demonstrate that over-expression of GmPHD6 improved stress tolerance in soybean, not only in transgenic hairy roots, but also in the whole plants. We screened the possible proteins interacted with GmPHD6 by yeast two-hybrid assay, and two heterochromatin proteins, namely LHPa and LHPb, were identified. It is possible that GmPHD6 may recruit these co-factors, LHPa or LHPb, for regulation of downstream genes and affect stress responses. The present project will further analyze the biochemical characteristics of GmPHD6 and LHP. Molecular mechanism that GmPHD6 cooperates with LHPa/b to regulate salt stress response will be studied. We will further identify the downstream genes of GmPHD6 to establish the integrated regulation model of GmPHD6. Completion of this project would be beneficial for the breeding of stress tolerant soybean in the future.

大豆是我国农业生产中极其重要的一类粮食作物。非生物胁迫是造成大豆减产的重要原因，提高大豆的耐逆能力有重大意义。通过比较耐逆大豆和逆境敏感大豆基因表达差异，我们发现了一类PHD锌指蛋白编码基因。这一类基因仅存在于植物中且响应逆境胁迫。这类PHD蛋白具有DNA结合能力和转录调控能力，然而其中有一个特例GmPHD6，它没有转录调控能力。使用毛状根转化系统，证实过表达GmPHD6基因能提高转基因毛状根的耐盐能力，且提高了整个大豆植株的耐盐能力。利用酵母双杂交体系，筛选到了两个异染色质蛋白LHPa和LHPb。GmPHD6可能需要与LHP形成复合体，通过LHP的激活能力调控下游基因表达。本研究拟解析GmPHD6和LHP的生化特性；解析二者协同调控大豆耐逆的机制；筛选GmPHD6调控的下游基因，建立完整的调控模型，为大豆育种奠定理论基础。

大豆是一种重要的粮食作物和油料作物，然而大豆种植中会遇到各种非生物胁迫，造成大豆的减产，提高大豆的耐逆能力有重大意义。本研究发现了一类在耐盐大豆和盐敏感大豆中差异表达的基因，编码一组仅存在于植物中的PHD锌指蛋白。这个家族中的一个成员，GmPHD6，仅具有DNA结合能力却没有转录调控能力。使用毛状根转化系统发现过量表达GmPHD6基因能提高转基因植株的耐盐能力，而GmPHD6表达量下调的RNAi植株则表现为盐敏感表型。稳定的GmPHD6转基因大豆植株亦表现出耐盐表型，与毛状根转化体系结果相符。利用酵母双杂交体系，筛选到了两个异染色质蛋白LHPa和LHPb。GmPHD6的PHD结构域能与LHP蛋白结合，而其N端结构域则主要负责识别组蛋白H3K4的甲基化修饰密码，结合下游基因的启动子也主要由N端结构域参与。LHP蛋白具有转录激活活性，但是过量表达LHP基因并不能提高转基因毛状根的耐盐能力。使用RNAi系统将两个LHP基因下调后，转基因植株却呈现盐敏感表型。因此推测LHP在GmPHD6的耐盐调控途径中必不可少，但它们并不直接发挥作用。在GmPHD6识别组蛋白密码并定位下游耐盐基因后，通过招募LHP蛋白的转录活性促使这些耐盐基因的表达量上调，从而提高大豆耐盐能力。本研究详细的解析了GmPHD6/LHP复合体调控大豆耐盐的分子机制，为培育耐盐大豆品种提供了重要的理论依据。