Vaccine Development Laboratory, Department of Microbiology, Institute of Biomedical Sciences, University of São Paulo, Av. Prof. Lineu Prestes, 1374, Cidade Universitária, São Paulo, SP, 05508-900, Brazil.
Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo, Brazil.
Braz J Microbiol. 2020 Jun;51(2):455-465. doi: 10.1007/s42770-020-00231-2. Epub 2020 Feb 3.
Enterotoxigenic Escherichia coli (ETEC) represents one of the most important etiological agents of diarrhea in developing countries and characteristically produces at least one of two enterotoxins: heat-labile toxin (LT) and heat-stable toxin (ST). It has been previously shown that the production and release of LT by human-derived ETEC strains are variable. Although the natural genetic polymorphisms of regulatory sequences of LT-encoding (eltAB) genes may explain the variable production of LT, the knowledge of the transcriptional and posttranscriptional aspects affecting LT expression among ETEC strains is not clear. To further understand the factors affecting LT expression, we evaluated the impact of the natural polymorphism in noncoding regulatory sequences of eltAB among clinically derived ETEC strains. Sequence analyses of seven clinically derived strains and the reference strain H10407 revealed polymorphic sites at both the promoter and upstream regions of the eltAB operon. Operon fusion assays with GFP revealed that specific nucleotide changes in the Pribnow box reduce eltAB transcription. Nonetheless, the total amounts of LT produced by the tested ETEC strains did not strictly correspond to the detected LT-specific mRNA levels. Indeed, the stability of LT varied according to the tested strain, indicating the presence of posttranscriptional mechanisms affecting LT expression. Taken together, our results indicate that the production of LT is a strain-specific process and involves transcriptional and posttranscriptional mechanisms that regulate the final amount of toxin produced and released by specific strains.
产肠毒素性大肠杆菌（ETEC）是发展中国家腹泻最重要的病因之一，其特征是至少产生两种肠毒素之一：不耐热毒素（LT）和耐热毒素（ST）。先前已经表明，人源 ETEC 菌株产生和释放 LT 的情况存在差异。尽管 LT 编码（eltAB）基因的调控序列的自然遗传多态性可能解释了 LT 的可变产生，但影响 ETEC 菌株中 LT 表达的转录和转录后方面的知识尚不清楚。为了进一步了解影响 LT 表达的因素，我们评估了临床衍生的 ETEC 菌株中 eltAB 非编码调控序列的自然多态性对 LT 表达的影响。对 7 株临床分离株和参考株 H10407 的序列分析显示，eltAB 操纵子的启动子和上游区域均存在多态性位点。带有 GFP 的操纵子融合测定表明，Pribnow 盒中的特定核苷酸变化会降低 eltAB 的转录。尽管如此，测试的 ETEC 菌株产生的 LT 总量并不严格对应检测到的 LT 特异性 mRNA 水平。事实上，LT 的稳定性因测试菌株而异，表明存在影响 LT 表达的转录后机制。总之，我们的结果表明，LT 的产生是一种菌株特异性的过程，涉及调节特定菌株产生和释放的毒素最终量的转录和转录后机制。
