油菜金属耐受蛋白基因BnMTP10参与锰元素积累调控

BnMTP10 regulates manganese accumulation in Brassica napus

摘要:

环境中锰(Mn)等微量矿质元素缺乏或过量胁迫是油菜(Brassica napus L.)等作物生产中的常见限制因素。为了鉴定油菜中介导Mn转运的关键基因，并解析其作用机理，选取前期利用Mn胁迫条件筛选油菜cDNA文库得到的MTP家族成员基因为研究对象。依据序列信息和系统发育树分析结果，将其命名为BnMTP10，其属于Mn-CDF亚家族成员。BnMTP10在酿酒酵母中表达后能够显著增强转化子对过量的Mn和铁(Fe)胁迫的耐受性，转化子中Fe和Mn的含量显著降低，但是对过量镉(Cd)、硼(B)、铝(Al)、锌(Zn)和铜(Cu)胁迫的作用不明显。实时荧光定量PCR (quantitative real-time PCR,qRT-PCR)结果表明，BnMTP10在油菜的花中表达较高，其次为根和叶；亚细胞定位结果表明其定位于内质网。与野生型相比，过表达BnMTP10的转基因拟南芥对Mn的过量胁迫的耐受性更强，但过量Fe胁迫后的表型与野生型没有显著差异。相应地，在过量Mn胁迫条件下，转基因拟南芥根中的Mn含量显著增加；而在过量Fe胁迫下，转基因拟南芥中的Fe含量没有显著变化。推测BnMTP10在植物中可能通过介导Mn向内质网的转运，在植物抵御Mn过量胁迫的过程中发挥作用。本研究在揭示油菜矿质营养高效机理的同时，为相关油菜种质创新提供了理论依据和基因资源。

Abstract:

Stresses induced by the deficiency or excess of trace mineral elements, such as manganese (Mn), represent a common limiting factor for the production of crops like Brassica napus. To identify key genes involved in Mn allocation in B.napus and elucidate the underlying mechanisms, a member of the metal tolerance protein (MTP) family obtained in the previous screening of cDNA library of B.napus under Mn stress was selected as the research subject. Based on the sequence information and phylogenetic analysis, it was named as BnMTP10. It belongs to the Mn-cation diffusion facilitator (CDF) subfamily. Expression of BnMTP10 in yeast significantly improved the tolerance of transformants to excessive Mn and iron (Fe) and reduced the accumulation of Mn and Fe. However, the yeast transformants exhibited no significant changes in tolerance to excess cadmium, boron, aluminum, zinc, or copper. The qRT-PCR results demonstrated that the flowers of B.napus had the highest expression of BnMTP10, followed by roots and leaves. Subcellular localization studies revealed that BnMTP10 was localized in the endoplasmic reticulum (ER). Compared with wild-type plants, transgenic Arabidopsis overexpressing BnMTP10 exhibited enhanced tolerance to excessive Mn stress but showed no significant difference under Fe stress. Correspondingly, under excessive Mn stress, the Mn content in the roots of transgenic Arabidopsis increased significantly. However, under excessive Fe stress, the Fe content in transgenic Arabidopsis did not alter significantly. According to the results, we hypothesize that BnMTP10 may alleviate excessive Mn stress in plants by mediating Mn transport to the ER. This study facilitated our understanding of efficient mineral nutrients, and provided theoretical foundations and gene resources for breeding B.napus.