Cerboneschi Matteo, Decorosi Francesca, Biancalani Carola, Ortenzi Maria Vittoria, Macconi Sofia, Giovannetti Luciana, Viti Carlo, Campanella Beatrice, Onor Massimo, Bramanti Emilia, Tegli Stefania
Dipartimento di Scienze delle Produzioni Agroalimentari e dell'Ambiente, Laboratorio di Patologia Vegetale Molecolare, Università degli Studi di Firenze, Via della Lastruccia 10, 50019 Sesto Fiorentino (Firenze), Italy.
Dipartimento di Scienze Produzioni Agroalimentari e dell'Ambiente, Genexpress, Università degli Studi di Firenze, Via della Lastruccia 14, 50019 Sesto Fiorentino (Firenze), Italy.
Res Microbiol. 2016 Nov-Dec;167(9-10):774-787. doi: 10.1016/j.resmic.2016.09.002. Epub 2016 Sep 13.
The plant pathogenic bacterium Pseudomonas savastanoi, the causal agent of olive and oleander knot disease, uses the so-called "indole-3-acetamide pathway" to convert tryptophan to indole-3-acetic acid (IAA) via a two-step pathway catalyzed by enzymes encoded by the genes in the iaaM/iaaH operon. Moreover, pathovar nerii of P. savastanoi is able to conjugate IAA to lysine to generate the less biologically active compound IAA-Lys via the enzyme IAA-lysine synthase encoded by the iaaL gene. Interestingly, iaaL is now known to be widespread in many Pseudomonas syringae pathovars, even in the absence of the iaaM and iaaH genes for IAA biosynthesis. Here, two knockout mutants, ΔiaaL and ΔiaaM, of strain Psn23 of P. savastanoi pv. nerii were produced. Pathogenicity tests using the host plant Nerium oleander showed that ΔiaaL and ΔiaaM were hypervirulent and hypovirulent, respectively and these features appeared to be related to their differential production of free IAA. Using the Phenotype Microarray approach, the chemical sensitivity of these mutants was shown to be comparable to that of wild-type Psn23. The main exception was 8 hydroxyquinoline, a toxic compound that is naturally present in plant exudates and is used as a biocide, which severely impaired the growth of ΔiaaL and ΔiaaM, as well as growth of the non-pathogenic mutant ΔhrpA, which lacks a functional Type Three Secretion System (TTSS). According to bioinformatics analysis of the Psn23 genome, a gene encoding a putative Multidrug and Toxic compound Extrusion (MATE) transporter, was found upstream of iaaL. Similarly to iaaL and iaaM, its expression appeared to be TTSS-dependent. Moreover, auxin-responsive elements were identified for the first time in the modular promoters of both the iaaL gene and the iaaM/iaaH operon of P. savastanoi, suggesting their IAA-inducible transcription. Gene expression analysis of several genes related to TTSS, IAA metabolism and drug resistance confirmed the presence of a concerted regulatory network in this phytopathogen among virulence, fitness and drug efflux.
植物病原菌野油菜黄单胞菌（Pseudomonas savastanoi）是橄榄和夹竹桃结瘤病的病原体，它通过所谓的“吲哚 - 3 - 乙酰胺途径”，经由iaaM/iaaH操纵子中基因编码的酶催化的两步途径，将色氨酸转化为吲哚 - 3 - 乙酸（IAA）。此外，野油菜黄单胞菌的致病变种nerii能够通过iaaL基因编码的IAA - 赖氨酸合酶，将IAA与赖氨酸结合，生成生物活性较低的化合物IAA - Lys。有趣的是，现在已知iaaL在许多丁香假单胞菌（Pseudomonas syringae）致病变种中广泛存在，即使在缺乏用于IAA生物合成的iaaM和iaaH基因的情况下也是如此。在此，构建了野油菜黄单胞菌致病变种nerii的菌株Psn23的两个基因敲除突变体，即ΔiaaL和ΔiaaM。使用寄主植物夹竹桃进行的致病性测试表明，ΔiaaL和ΔiaaM分别表现出超毒力和低毒力，这些特征似乎与它们游离IAA的差异产生有关。使用表型微阵列方法表明，这些突变体的化学敏感性与野生型Psn23相当。主要例外是8 - 羟基喹啉，一种天然存在于植物分泌物中并用作杀菌剂的有毒化合物，它严重损害了ΔiaaL和ΔiaaM以及缺乏功能性III型分泌系统（TTSS）的非致病性突变体ΔhrpA的生长。根据对Psn23基因组的生物信息学分析，在iaaL上游发现了一个编码假定的多药和有毒化合物外排（MATE）转运蛋白的基因。与iaaL和iaaM类似，其表达似乎依赖于TTSS。此外，首次在野油菜黄单胞菌的iaaL基因和iaaM/iaaH操纵子的模块化启动子中鉴定出生长素反应元件，表明它们的IAA诱导型转录。对与TTSS、IAA代谢和耐药性相关的几个基因的基因表达分析证实，在这种植物病原体中，毒力、适应性和药物外排之间存在协同调节网络。
