Álvarez-Morales Ariel, Hernández-Morales Alejandro, Arvizu-Gómez Jackeline L
Centro de Investigación y de Estudios Avanzados del IPN, Irapuato, Gto., Mexico.
Unidad Académica Multidisciplinaria Zona Huasteca, Universidad Autónoma de San Luis Potosí, San Luis Potosí, Mexico.
Rev Argent Microbiol. 2018 Apr-Jun;50(2):115-125. doi: 10.1016/j.ram.2017.07.003. Epub 2017 Nov 10.
Pseudomonas syringae pv. phaseolicola is a phytopathogenic bacterium in beans that produces a phytotoxin called phaseolotoxin, in whose synthesis a group of genes that belong to the "Pht cluster" are involved. This cluster comprises 23 genes arranged in 5 transcriptional units, two monocistronic (argK, phtL) and three polycistronic (phtA, phtD, phtM) operons, whose expression is increased at 18°C, correlating with the production of phaseolotoxin by the bacterium. So far, the regulatory mechanisms involved in phaseolotoxin synthesis are poorly understood and only the requirement of low temperatures for its synthesis has been demonstrated. Therefore, in this study we searched for regulatory proteins that could be involved in the phaseolotoxin synthesis, focusing on the regulation of the phtM operon. Gel shift assays showed that the promoter region of the phtM operon contains binding sites for putative regulatory proteins, which are encoded outside the Pht cluster and are independent of the GacS-GacA two-component system. Deletion assays with the promoter region of the phtM operon show that the binding site for a putative transcription factor is located within a 58bp region. The putative transcription factor of the phtM operon has an apparent molecular mass in the 14-20kDa range. Furthermore, the results demonstrate that the transcription factor recognizes and binds the upstream phtM region as monomer o multimer of a single polypeptide. Our findings provide new insights into the regulatory mechanisms involved in phaseolotoxin production, and suggest that the Pht cluster was integrated into the global regulatory mechanism of P. syringae pv. phaseolicola.
菜豆丁香假单胞菌菜豆致病变种是豆类中的一种植物致病细菌，它能产生一种名为菜豆毒素的植物毒素，在其合成过程中涉及一组属于“Pht簇”的基因。该簇由23个基因组成，排列在5个转录单元中，两个单顺反子（argK、phtL）和三个多顺反子（phtA、phtD、phtM）操纵子，其表达在18°C时增加，这与该细菌产生菜豆毒素相关。到目前为止，菜豆毒素合成所涉及的调控机制了解甚少，仅证明了其合成需要低温。因此，在本研究中，我们寻找可能参与菜豆毒素合成的调控蛋白，重点关注phtM操纵子的调控。凝胶迁移试验表明，phtM操纵子的启动子区域含有假定调控蛋白的结合位点，这些蛋白在Pht簇外编码，且独立于GacS - GacA双组分系统。对phtM操纵子启动子区域的缺失试验表明，一种假定转录因子的结合位点位于一个58bp的区域内。phtM操纵子的假定转录因子的表观分子量在14 - 20kDa范围内。此外，结果表明该转录因子以单个多肽的单体或多聚体形式识别并结合phtM上游区域。我们的研究结果为菜豆毒素产生所涉及的调控机制提供了新的见解，并表明Pht簇已整合到菜豆丁香假单胞菌菜豆致病变种的全局调控机制中。
