Ammonia activates pacC and patulin accumulation in an acidic environment during apple colonization by Penicillium expansum
Ammonia activates pacC and patulin accumulation in an acidic environment during apple colonization by Penicillium expansum
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DOI：
10.1111/mpp.12327
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发表时间：
2016-06-01
影响因子：
4.9
通讯作者：
Prusky, Dov
中科院分区：
文献类型：
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作者：
Barad, Shiri;Espeso, Eduardo A.;Prusky, Dov
扩展青霉（Penicillium expansum）是蓝霉腐病的致病因子，会导致采后果实严重浸渍，同时在定植组织中分泌 D-葡萄糖酸 (GLA) 和霉菌毒素展青霉素。在宿主酸性环境定植期间诱导棒曲霉素生物合成的因素尚不清楚。在苹果果实体内定植和培养物生长过程中，P. expansum 会分泌调节 pH 值的 GLA 和氨。尽管棒曲霉素及其可能的机会性前体 GLA 在真菌发育过程中一起积累，但在定植组织的前缘和扩展培养后检测到氨，接近棒曲霉素积累。在这里，我们通过以下方法证明了氨诱导的全局 pH 调节剂 PacC 的转录激活和在 GLA 存在下棒曲霉素的积累：(i) 对固体培养基上生长的 P. expansum 进行直接外源处理； (ii) 对定植的苹果组织进行直接外源处理； (iii) 在有限碳的自产氨条件下生长； (iv) 展青霉素生物合成簇对氨的转录反应的分析。氨诱导棒曲霉素积累，同时激活 pacC 和棒曲霉素生物合成簇基因的转录本，表明氨对酸性条件下 pacC 转录本表达的调节作用。使用 P. expansum PacC 和针对不同裂解蛋白的抗体进行的电泳迁移率变动测定表明，相对于 pH 7.0，PacC 在 pH 4.5 时不受蛋白水解信号传导的保护，但添加 NH4 并没有进一步增强其蛋白水解裂解。在酸性条件下，氨增强了 Pal 途径中 palF 转录本的激活。在酸性 pH 条件下，病原体在宿主环境中积累氨可能是 pacC 激活的调节信号，从而导致次生代谢产物（如棒曲霉素）的积累。
Penicillium expansum, the causal agent of blue mould rot, causes severe post-harvest fruit maceration simultaneously with the secretion of D-gluconic acid (GLA) and the mycotoxin patulin in colonized tissue. The factor(s) inducing patulin biosynthesis during colonization of the host acidic environment is unclear. During the colonization of apple fruit in vivo and growth in culture, P. expansum secretes pH-modulating GLA and ammonia. Although patulin and its possible opportunistic precursor GLA accumulate together during fungal development, ammonia is detected on the colonized tissue's leading edge and after extended culture, close to patulin accumulation. Here, we demonstrate ammonia-induced transcript activation of the global pH modulator PacC and patulin accumulation in the presence of GLA by: (i) direct exogenous treatment of P. expansum growing on solid medium; (ii) direct exogenous treatment on colonized apple tissue; (iii) growth under self-ammonia production conditions with limited carbon; and (iv) analysis of the transcriptional response to ammonia of the patulin biosynthesis cluster. Ammonia induced patulin accumulation concurrently with the transcript activation of pacC and patulin biosynthesis cluster genes, indicating the regulatory effect of ammonia on pacC transcript expression under acidic conditions. Electrophoretic mobility shift assays using P. expansum PacC and antibodies to the different cleaved proteins showed that PacC is not protected against proteolytic signalling at pH 4.5 relative to pH 7.0, but NH4 addition did not further enhance its proteolytic cleavage. Ammonia enhanced the activation of palF transcript in the Pal pathway under acidic conditions. Ammonia accumulation in the host environment by the pathogen under acidic pH may be a regulatory cue for pacC activation, towards the accumulation of secondary metabolites, such as patulin.