Identification of target genes of the bZIP transcription factor OsTGAP1, whose overexpression causes elicitor-induced hyperaccumulation of diterpenoid phytoalexins in rice cells.
Identification of target genes of the bZIP transcription factor OsTGAP1, whose overexpression causes elicitor-induced hyperaccumulation of diterpenoid phytoalexins in rice cells.
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DOI：
10.1371/journal.pone.0105823
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Okada K
中科院分区：
文献类型：
--
作者：
Miyamoto K;Matsumoto T;Okada A;Komiyama K;Chujo T;Yoshikawa H;Nojiri H;Yamane H;Okada K
植物抗菌素是一种特殊的抗微生物代谢产物，由植物对病原体的攻击做出反应。马利内酮和植物卡介烷是水稻中主要的二萜类植物保卫素，由香叶基香叶基二磷酸合成，香叶基香叶基二磷酸是甲基赤藓糖醇磷酸(MEP)途径的产物。在OsTGAP1过表达(OsTGAP1ox)的水稻细胞中，由于相关生物合成基因和MEP途径基因OsDXS3的高诱导表达，棉铃虫和植物内酯的超量积累已被几丁质低聚糖激发子刺激。本研究通过转录组分析和染色质免疫沉淀结合下一代测序的方法，鉴定了122个OsTGAP1靶基因，以阐明激发子诱导的这些植物保卫素生物合成基因在OsTGAP1ox细胞中协同超诱导的机制。转录组分析表明，在激发子刺激的OsTGAP1ox细胞中，几乎所有分别位于第4号和第2号染色体上的丁香内酯和植物内酯生物合成基因以及MEP途径基因都得到了高诱导表达。令人意外的是，这些聚集的基因都不包括在OsTGAP1的靶基因中，这表明OsTGAP1不是通过与每个启动子区域结合来直接调控这些生物合成基因的表达。然而，有趣的是，在簇区之间和附近的基因间隔区中发现了几个OsTGAP1结合区。关于MEP途径基因，只有编码MEP途径关键酶的OsDXS3在其上游区域具有OsTGAP1结合区。随后的转录激活实验进一步证实了OsTGAP1对OsDXS3表达的直接调控，但其他MEP途径基因不包括在OsTGAP1靶基因中。综上所述，这些结果表明，OsTGAP1参与了二萜类植物保卫素的增强积累，主要是通过直接转录调控这些植物保卫素生物合成途径中涉及的基因。
Phytoalexins are specialised antimicrobial metabolites that are produced by plants in response to pathogen attack. Momilactones and phytocassanes are the major diterpenoid phytoalexins in rice and are synthesised from geranylgeranyl diphosphate, which is derived from the methylerythritol phosphate (MEP) pathway. The hyperaccumulation of momilactones and phytocassanes due to the hyperinductive expression of the relevant biosynthetic genes and the MEP pathway gene OsDXS3 in OsTGAP1-overexpressing (OsTGAP1ox) rice cells has previously been shown to be stimulated by the chitin oligosaccharide elicitor. In this study, to clarify the mechanisms of the elicitor-stimulated coordinated hyperinduction of these phytoalexin biosynthetic genes in OsTGAP1ox cells, transcriptome analysis and chromatin immunoprecipitation with next-generation sequencing were performed, resulting in the identification of 122 OsTGAP1 target genes. Transcriptome analysis revealed that nearly all of the momilactone and phytocassane biosynthetic genes, which are clustered on chromosomes 4 and 2, respectively, and the MEP pathway genes were hyperinductively expressed in the elicitor-stimulated OsTGAP1ox cells. Unexpectedly, none of the clustered genes was included among the OsTGAP1 target genes, suggesting that OsTGAP1 did not directly regulate the expression of these biosynthetic genes through binding to each promoter region. Interestingly, however, several OsTGAP1-binding regions were found in the intergenic regions among and near the cluster regions. Concerning the MEP pathway genes, only OsDXS3, which encodes a key enzyme of the MEP pathway, possessed an OsTGAP1-binding region in its upstream region. A subsequent transactivation assay further confirmed the direct regulation of OsDXS3 expression by OsTGAP1, but other MEP pathway genes were not included among the OsTGAP1 target genes. Collectively, these results suggest that OsTGAP1 participates in the enhanced accumulation of diterpenoid phytoalexins, primarily through mechanisms other than the direct transcriptional regulation of the genes involved in the biosynthetic pathway of these phytoalexins.
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