浙麦冬HMA基因家族鉴定及镉胁迫响应分析

Identification of HMA gene family and response to cadmium stress in Ophiopogon japonicas

摘要:

土壤镉污染是当前全球面临的重要环境问题之一。浙麦冬(Ophiopogon japonicus)作为一种多功能植物，不仅在传统医学中广泛应用，其在环境修复领域中的应用潜力也逐渐受到关注。本文旨在探讨浙麦冬在不同浓度镉胁迫下的镉富集规律，鉴定和分析浙麦冬重金属ATP酶(heavy metal ATPase,HMA)家族成员。研究发现浙麦冬富集系数高达2.75，在土壤镉污染修复中显示出良好的镉富集潜力。基于前期转录组数据鉴定出9个浙麦冬P1B型重金属ATP酶(P1B-ATPases)家族成员，其中OjHMA1–OjHMA6属于锌/钴/镉/铅ATP酶(Zn/Co/Cd/Pb-ATPases)，OjHMA7–OjHMA9则属于铜/银ATP酶(Cu/Ag-ATPases)。在镉胁迫下，OjHMA1、OjHMA2、OjHMA3和OjHMA7等基因表达显著上调，表明其在镉离子的吸收和转运过程中发挥核心作用。拓扑结构分析揭示了HMA蛋白具有该家族典型的跨膜(transmembrane,TM)片段以及调节离子吸收与释放的A、P和N功能域，跨膜片段上存在着金属离子结合位点(M4、M5、M6)。根据跨膜域数目及金属结合位点上氨基酸的不同，可将植物的HMA家族蛋白分为3个亚组：P1B-1 ATPases、P1B-2 ATPases和P1B-4 ATPases。P1B-1 ATPases亚组包含的TM4(CPC)、TM5(YN[X]₄P)和TM6(M[XX]SS)基序，P1B-2 ATPases亚组的TM4(CPC)、TM5(K)以及TM6(DKTGT)基序，P1B-4 ATPases亚组的TM4中的SPC和TM6中的HE[X]GT基序，都是其蛋白发挥功能的关键。在此基础上，利用分子对接对金属离子结合位点进行了详细解析，揭示了CPC/SPC、DKTGT和HE[X]GT等关键保守序列在金属离子配位和稳定中的重要作用。本研究结果不仅提供了通过基因工程提高浙麦冬对镉吸附和耐受性的分子靶点，也为培育具有高镉吸附能力的新品种提供了理论基础。

关键词:浙麦冬;HMA家族;镉胁迫;富集系数;分子对接

Abstract:

Soil cadmium (Cd) pollution is one of the major environmental problems globally. Ophiopogon japonicus, a multifunctional plant extensively used in traditional Chinese medicine, has demonstrated potential in environmental remediation. This study investigated the Cd accumulation pattern of O. japonicus under cadmium stress and identified the heavy metal ATPase (HMA) family members in this plant. Our results demonstrated that O. japonicus exhibited a Cd enrichment factor (EF) of 2.75, demonstrating strong potential for soil Cd pollution remediation. Nine heavy metal ATPase (HMA) members of P1B-ATPases were successfully identified from the transcriptome data of O. japonicus, with OjHMA1–OjHMA6 classified as the Zn/Co/Cd/Pb-ATPases and OjHMA7–OjHMA9 as the Cu/Ag-ATPases. The expression levels of OjHMA1, OjHMA2, OjHMA3, and OjHMA7 were significantly up-regulated under Cd stress, highlighting their crucial roles in cadmium ion absorption and transport. The topological analysis revealed that these proteins possessed characteristic transmembrane (TM) segments of the family, along with functional A, P, and N domains involved in regulating ion absorption and release. Metal ion-binding sites (M4, M5, and M6) existed on the TM segments. Based on the number of transmembrane domains and the residues at metal ion-binding sites, the plant HMA family members were categorized into three subgroups: P1B-1 ATPases, P1B-2 ATPases, and P1B-4 ATPases. Specifically, the P1B-1 ATPase subgroup included the motifs TM4(CPC), TM5(YN[X]₄P), and TM6(M[XX]SS); the P1B-2 ATPase subgroup featured the motifs TM4(CPC), TM5(K), and TM6(DKTGT); the P1B-4 ATPase subgroup contained the motifs TM4(SPC) and TM6(HE[X]GT), all of which were critical for protein functions. Molecular docking results revealed the importance of conserved sequences such as CPC/SPC, DKTGT, and HE[X]GT in metal ion coordination and stabilization. These findings provide potential molecular targets for enhancing Cd uptake and tolerance of O. japonicus by genetic engineering and lay a theoretical foundation for developing new cultivars with high Cd accumulation capacity.

Key words:Ophiopogon japonicas;HMA family;cadmium (Cd) stress;enrichment factor (EF);molecular docking