不同耐铝型杉木无性系响应铝毒胁迫的细胞壁调控机制

Aluminum (Al) toxicity is the main factor limiting the productivity of Chinese fir (Cunninghamia Lanceolata) in acid soils of southern China. Therefore, elucidating the adaptive strategy and regulatory mechanism of Chinese fir under Al stress is of great important in enhancing the Al resistance in Chinese fir. It has been reported that cell wall mediated Al exclusion is the key Al resistance mechanism in plants, and our previous studies showed that there existed significant differences responses in root cell wall of Chinese fir clones differing in Al resistance under Al stress, however, little is known about how the Al resistance of Chinese fir is regulated by cell wall. In the present proposal, two Chinese fir clones differing in Al resistance were employed to investigate the different responses of several key physiological indexes relating to Al resistance and cell wall polysaccharides under Al stress, revealing the changes in cell wall polysaccharides and its relationship with Chinese fir Al resistance. Then the differences in adsorption ability of different cell wall components to Al ion as well as their main functional groups that interact with Al ion were determined, which revealed the underlying mechanism of cell wall adsorbed and accumulated Al. Moreover, Al-induced hydrogen peroxide formation and the regulatory pathway and mechanism of hydrogen peroxide mediated cell wall modification were examined by using hydrogen peroxide and its scavenger, in addition, gene co-expression networks were also constructed with functional genes associations with cell wall modification after the samples were subjected to high-throughput sequencing. Through implementation of this project, it will increase our understanding of the cell wall regulatory mechanism of Chinese fir clones differing in Al resistance in response to Al stress, and provide a theoretical basis for genetic improvement of Al resistance in Chinese fir.

铝毒是制约南方酸性土壤中杉木人工林产量的重要因素，解析杉木适应铝毒的策略和调控机制对提高杉木铝耐性具有重要意义。研究表明细胞壁介导的铝外排是植物关键的耐铝机制，申请者在前期研究中发现不同耐铝型杉木无性系根细胞壁对铝胁迫响应存在显著差异，但关于细胞壁如何介导杉木耐铝性调控的机制仍不清楚。本项目以不同耐铝型杉木无性系为材料，通过比较铝胁迫下杉木各抗铝关键生理指标和细胞壁多糖对铝毒响应的差异，揭示细胞壁多糖的变化及其与杉木耐铝性的关系；同时分析细胞壁各组分对铝吸附能力的差异及其与铝发生作用的功能基团，揭示细胞壁吸收积累铝的内在机制；在此基础上利用外源过氧化氢及其清除剂，探讨铝胁迫诱导产生的过氧化氢形成机制及其在细胞壁修饰中的调控途径和作用机制，结合高通量测序，构建过氧化氢介导的细胞壁修饰基因共表达网络，阐明不同耐铝型杉木无性系响应铝胁迫的细胞壁调控机制，从而为杉木耐铝性状的遗传改良提供科学依据。

本项目研究结果表明：1）铝胁迫下，不同耐铝杉木基因型根系伸长受到显著抑制，其根中铝、H2O2、丙二醛和胼胝质含量显著增加，且铝敏感基因型YX02显著高于耐铝基因型YX01。进一步研究发现铝胁迫引起敏感型杉木根中氧化损伤加剧与铝在其细胞壁、果胶、半纤维素中显著增加有关。铝胁迫下，敏感型杉木细胞壁及其各组分中铝含量显著高于耐性基因型与铝胁迫显著增加其细胞壁果胶和半纤维素1含量以及降低细胞壁果胶甲酯化程度有关。细胞壁吸附试验表明，铝胁迫诱导细胞壁组成和性质的改变能显著增加细胞壁对铝的吸附能力，细胞壁不同组分和细胞壁改性吸附结果表明果胶和半纤维素1是杉木根中吸附铝的主要细胞壁组分，而羧基则是吸附铝的主要基团。2）为进一步明确H2O2在铝胁迫介导的细胞壁修饰中的功能和地位，进一步通过外源添加/清除H2O2进行研究。结果表明铝胁迫下外源添加H2O2进一步增加杉木根中H2O2、丙二醛和铝含量。进一步分析发现铝胁迫添加H2O2下铝含量的增加与其根中果胶和半纤维素1含量以及降低细胞壁果胶甲酯化程度有关，而铝胁迫添加H2O2清除剂则能在一定程度上逆转上述过程，缓解氧化损伤，暗示铝胁迫诱导的H2O2作为一个重要信号分子在杉木耐铝调控中扮演关键角色。3）转录组结果表明不同处理下苯丙氨酸生物合成途径、谷胱甘肽代谢、植物激素和信号转导、淀粉和蔗糖代谢、单萜化合物生物合成途径、氨基糖和核苷酸糖代谢和氮代谢是差异基因显著富集的通路。基因共表达分析结果表明筛选出参与铝胁迫下杉木耐铝调控的基因主要是抗氧化酶类、细胞壁修饰类以及乙烯和WRKY类转录因子。蛋白质互作网络结果也表明ERF、WRKY转录因子、苯丙氨酸代谢途径以及细胞壁合成修饰基因在不同处理中都是关键节点基因，暗示其在杉木耐铝调控中扮演重要角色，研究结果为后续进一步深入研究杉木耐铝的分子生物学机制奠定良好的基础。