Rocheleau Hélène, Al-Harthi Reem, Ouellet Thérèse
Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, 960 Carling Ave, Ottawa, ON K1A 0C6, Canada.
Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, 960 Carling Ave, Ottawa, ON K1A 0C6, Canada; Department of Biology, University of Ottawa, 30 Marie Currie, Ottawa, ON K1N 6N5, Canada.
Fungal Biol. 2019 Jan;123(1):77-86. doi: 10.1016/j.funbio.2018.11.002. Epub 2018 Nov 17.
Fusarium head blight (FHB) is a major cereal crop disease, caused most frequently by the fungus Fusarium graminearum. We have previously demonstrated that F. graminearum can utilize SA as sole source of carbon to grow. In this current study, we further characterized selected four fungal SA-responsive genes that are predicted to encode salicylic acid (SA)-degrading enzymes and we used a gene replacement approach to characterize them further. These included two genes predicted to encode a salicylate 1-monooxygenase, FGSG_03657 and FGSG_09063, a catechol 1, 2-dioxygenase gene, FGSG_03667, and a 2, 3-dihydroxybenzoic acid decarboxylase gene, FGSG_09061. For each gene, three independent gene replacement strains were assayed for their ability to degrade salicylic acid in liquid culture. Salicylate 1-monooxygenase FGSG_03657 and catechol 1, 2-dioxygenase FGSG_03667 were shown to be essential for SA degradation, while a loss of 2, 3-dihydroxybenzoic acid decarboxylase FGSG_09061 caused only a partial reduction of SA degradation and a loss of salicylate 1-monooxygenase FGSG_09063 had no effect when compared to wild type culture. Salicylate 1-monooxygenase FGSG_03657 and catechol 1, 2-dioxygenase FGSG_03667 were identified as the first two key enzyme steps of SA degradation via catechol in the β-ketoadipate pathway. Expression profiles for all four genes were also determined in liquid culture and in planta. Salicylate 1-monooxygenase FGSG_03657 and catechol 1, 2-dioxygenase FGSG_03667 were co-expressed and their expression was substrate dependent in liquid culture; however their expression was uncoupled in planta. Disruption of the gene for catechol 1, 2-dioxygenase FGSG_03667 was shown to have no effect on fungal virulence on wheat. Our results with 2, 3-dihydroxybenzoic acid decarboxylase FGSG_09061 raise the possibility of an alternate non-oxidative decarboxylation pathway for the conversion of SA to catechol via 2, 3-dihydrozybenzoic acid and for a connection between the oxidative and the non-oxidative decarboxylation pathways for SA conversion.
小麦赤霉病（FHB）是一种主要的谷物作物病害，最常见的致病真菌是禾谷镰刀菌。我们之前已经证明，禾谷镰刀菌可以利用水杨酸（SA）作为唯一碳源进行生长。在本研究中，我们进一步对选定的四个预测编码水杨酸（SA）降解酶的真菌SA反应基因进行了表征，并使用基因替换方法对其进行了进一步表征。其中包括两个预测编码水杨酸1-单加氧酶的基因FGSG_03657和FGSG_09063、一个儿茶酚1,2-双加氧酶基因FGSG_03667和一个2,3-二羟基苯甲酸脱羧酶基因FGSG_09061。对于每个基因，检测了三个独立的基因替换菌株在液体培养中降解水杨酸的能力。结果表明，水杨酸1-单加氧酶FGSG_03657和儿茶酚1,2-双加氧酶FGSG_03667对SA降解至关重要，而2,3-二羟基苯甲酸脱羧酶FGSG_09061的缺失仅导致SA降解部分减少，与野生型培养相比，水杨酸1-单加氧酶FGSG_09063的缺失没有影响。水杨酸1-单加氧酶FGSG_03657和儿茶酚1,2-双加氧酶FGSG_03667被确定为β-酮己二酸途径中通过儿茶酚进行SA降解的前两个关键酶步骤。还测定了所有四个基因在液体培养和植物中的表达谱。水杨酸1-单加氧酶FGSG_03657和儿茶酚1,2-双加氧酶FGSG_03667在液体培养中共同表达，且它们的表达依赖于底物；然而，它们在植物中的表达是解偶联的。结果表明，儿茶酚1,2-双加氧酶FGSG_03667基因的破坏对小麦上的真菌毒力没有影响。我们对2,3-二羟基苯甲酸脱羧酶FGSG_09061的研究结果提出了一种通过2,3-二羟基苯甲酸将SA转化为儿茶酚的替代非氧化脱羧途径以及SA转化的氧化和非氧化脱羧途径之间存在联系的可能性。
