水稻ABC转运蛋白OsARAPT1响应砷胁迫的分子机理研究

Arsenic contamination of rice and is a worldwide concern for human health and world food security. Now, the molecular mechanism of plant response to the arsenic was still limited understanding. Using previous Illumina high-throughput sequencing, we obtained an ABC transporter OsARAPT1 (Arsenate-responsive ABC PDR-type transporter 1), significantly elevated in higher or lower arsenic treatments both in roots and shoots, whose mutant was more sensitive than wild type plant exposed to the arsenic stress, suggesting that it might act as a positive regulator in arsenic stress response of rice. In this proposed study, we used Promoter-GUS method to get the fine expression pattern of OsARAPT1; and used GFP fusion protein approach to analysis the variation of OsARAPT1 protein subcellular localization, under the arsenic stress. Stable transgenic rice lines with overexpression (OE) for OsARAPT1 will be generated and analyzed for their phenotypes against arsenic. The content of arsenic of the rice expose to the arsenic stress will be carried out and compared among them. Results from this proposed project will be helpful in clarifying the biological functions and transport mechanism of OsARAPT1 in arsenic resistance as well as in elucidating the molecular mechanism of rice response against arsenic, which could be used as useful canadidate genes in generation of novel germplasm resources with improved arsenic resistance character through genetic engineering approach.

水稻砷污染是影响人类健康和威胁世界食品安全的重要因素之一，但目前对水稻响应砷胁迫的分子机理了解有限。我们前期通过高通量测序研究发现，水稻OsARAPT1基因在高浓度和低浓度的砷胁迫下表达量都明显上升，而且其突变体在砷胁迫下比野生型具有更加敏感的表型，推测该基因在水稻响应砷胁迫中起正向调控功能。本项目拟利用Promoter-GUS分析砷胁迫下的精细表达谱；构建GFP融合蛋白，解析OsARAPT1在砷胁迫下的亚细胞定位变化规律；构建基因过表达，比较转基因株系在砷胁迫下体内砷含量的动态变化及耐性差异。研究结果不仅有助于解析 OsARAPT1响应砷胁迫的生物学功能及转运砷的分子机理，而且能为创制高耐性、低富集性的转基因新品种提供基因资源。

水稻砷污染是影响人类健康和威胁世界食品安全的重要因素之一，但目前对水稻响应砷胁迫的分子机理了解有限。前期通过高通量测序研究发现，水稻OsARAPT1基因在高浓度和低浓度的砷胁迫下表达量都明显上升，推测该基因对水稻响应砷胁迫起着正向调控功能。OsARAPT1是ABC转运蛋白（ATP-binding cassette transporters，腺苷三磷酸结合盒转运蛋白）中PDR亚家族全分子转运蛋白中的一员，其NBD 位于TMD的N端，与其他亚家族具有相反的拓扑构型，是植物和真菌特有的一种转运蛋白。植物PDR 蛋白具有转运肽、糖、脂、离子、重金属螯合物、生物碱和类固醇等底物的重要功能。本研究通过PCR技术和GUS组织化学染色，获得了OsARAPT1基因响应砷胁迫的精细表达谱，发现该基因在高浓度砷处理下，6h就被诱导；而在低浓度砷处理下，12h被诱导。亚细胞定位分析显示OsARAPT1-GFP融合蛋白定位于细胞膜上。对OsARAPT1不同基因型植株进行砷处理的耐受性分析，结果显示OsARAPT1基因过表达植株可以提高植物对砷胁迫的耐受性。因此，课题研究成果不仅有助于解析转运蛋白响应砷胁迫的生物学功能和分子机理，而且能为后续培育和创制高耐性、低富集砷的转基因新品种提供基因资源和新的思路。