Fe(2+)调控水稻HAC砷酸还原酶还原As(V)的分子机理

Inorganic As soil contamination is common problem in many areas in the world. However, rice grown under upland or water-saving cultivation methods experiences long periods of aerobic soil conditions with arsenate being the predominant form of As taken up by rice. Recently, HAC arsenate reductase genes have been cloned and identified to be involved in arsenate reduction and arsenic accumulation in rice. Previous studies showed that the expression level of HAC genes and As(V) reduction ability could be improved by Fe(II). However, the mechanism of the work remains unknown. In this research, the expression level of HAC genes influenced by Fe(II) will be analyzed by the observation of GFP labeled HAC transgenic plants and qRT-PCR methods. The influence on As(V) reduction ability by Fe(II) will be identified by vitro enzyme activity analysis and heterologous expression system. Hydroponic and pot experiments with different Fe(II) conditions was conducted to investigate the phenotype and As accumulation in rice among different HAC genes transgenic plants. Results from this work will provide useful data for clarifying the molecular mechanisms of the As(V) reduction process involved by HAC arsenate reductase which was influenced by Fe(II) as well as controlling accumulation of arsenic in rice for food security.

土壤砷(As)污染主要以无机砷污染为主。好氧环境中，土壤中的无机砷主要以五价砷As(V)被水稻吸收。水稻HAC砷酸还原酶家族介导As(V)向As(III)的还原过程，是As(V)解毒的重要步骤。申请者前期研究发现，外源Fe(II)能显著提高水稻HAC砷酸还原酶基因的转录水平，促进As(V)的还原能力，但其分子机理尚不清楚。本项目拟通过观察绿色荧光蛋白(GFP)标记HAC转基因材料以及利用qRT-PCR技术，考察外源Fe(II)对HAC砷酸还原酶基因表达丰度与部位的调控作用；利用体外酶活分析以及异源表达体系，考察Fe(II)对HAC还原能力的调控作用；通过水培与盆栽实验，分析外源Fe(II)对水稻体内As(V)还原能力以及砷吸收富集能力的影响；揭示外源Fe(II)调控水稻HAC砷酸还原酶介导As(V)还原的分子机理，为调控水稻对As(V)的吸收富集，保障稻米质量安全提供理论依据。

水稻籽粒砷的积累与超标可以通过食物链传递到人体，严重威胁人类健康。降低和阻控水稻籽粒砷的积累需要对水稻砷的积累机制有明确了解。申请人前期研究表明外源铁可以显著提高砷酸盐还原酶的表达水平，促进五价砷的还原，但是具体机制仍不清楚。本项目旨在解析外源铁调控水稻对砷的吸收转运机制，主要通过明确外源铁添加对水稻根系砷的吸收和转运的影响，铁影响下水稻对不同浓度砷的响应和砷的吸收动力学分析，结合转录组测序技术和酵母高通量水稻砷耐性基因筛选技术，阐述铁影响水稻对砷的吸收的分子机制。试验获的证据表明，铁可以显著增加水稻根系对砷的吸附，降低砷向地上部分的转运，降低水稻根系对砷的吸收。同时，不同形态的铁对水稻根系对砷的吸附明显不同。试验也获得了铁和砷影响下水稻根系差异化表达基因以及水稻中具有砷耐性的基因，为进一步解析水稻对砷的吸收以及耐性奠定了研究基础。本项目的研究结果也有助于指导水稻安全生产，阻控水稻籽粒砷的积累。