Molecular Characterization of Foodborne Pathogens Research Unit, Eastern Regional Research Center, Agricultural Research Service, US Department of Agriculture, 600 East Mermaid Lane, Wyndmoor, PA, USA.
Department of Food Science, Penn State University, University Park, PA, USA.
Microbiology (Reading). 2013 Aug;159(Pt 8):1586-1596. doi: 10.1099/mic.0.066118-0. Epub 2013 Jun 6.
Biofilm formation in Escherichia coli is a tightly controlled process requiring the expression of adhesive curli fibres and certain polysaccharides such as cellulose. The transcriptional regulator CsgD is central to biofilm formation, controlling the expression of the curli structural and export proteins and the diguanylate cyclase adrA, which indirectly activates cellulose production. CsgD itself is highly regulated by two sigma factors (RpoS and RpoD), multiple DNA-binding proteins, small regulatory RNAs and several GGDEF/EAL proteins acting through c-di-GMP. One such transcription factor MlrA binds the csgD promoter to enhance the RpoS-dependent transcription of csgD. Bacteriophage, often carrying the stx1 gene, utilize an insertion site in the proximal mlrA coding region of E. coli serotype O157 : H7 strains, and the loss of mlrA function would be expected to be the major factor contributing to poor curli and biofilm expression in that serotype. Using a bank of 55 strains of serotype O157 : H7, we investigated the consequences of bacteriophage insertion. Although curli/biofilm expression was restored in many of the prophage-bearing strains by a wild-type copy of mlrA on a multi-copy plasmid, more than half of the strains showed only partial or no complementation. Moreover, the two strains carrying an intact mlrA were found to be deficient in biofilm formation. However, RpoS mutations that attenuated or inactivated RpoS-dependent functions such as biofilm formation were found in >70 % of the strains, including the two strains with an intact mlrA. We conclude that bacteriophage interruption of mlrA and RpoS mutations provide major obstacles limiting curli expression and biofilm formation in most serotype O157 : H7 strains.
大肠杆菌生物膜的形成是一个受到严格控制的过程，需要表达粘性卷曲菌纤维和某些多糖，如纤维素。转录调节因子 CsgD 是生物膜形成的核心，控制着卷曲结构和输出蛋白以及双鸟苷酸环化酶 adrA 的表达，后者间接激活纤维素的产生。CsgD 本身受到两个 sigma 因子（RpoS 和 RpoD）、多种 DNA 结合蛋白、小调控 RNA 和几种通过 c-di-GMP 作用的 GGDEF/EAL 蛋白的高度调控。转录因子 MlrA 就是其中之一，它结合 csgD 启动子，增强 csgD 的 RpoS 依赖性转录。噬菌体通常携带 stx1 基因，利用大肠杆菌 O157:H7 血清型近端 mlrA 编码区的插入位点，mlrA 功能的丧失预计是该血清型卷曲和生物膜表达不良的主要因素。使用 55 株 O157:H7 血清型菌株的文库，我们研究了噬菌体插入的后果。尽管许多带有噬菌体的菌株通过在多拷贝质粒上携带野生型 mlrA 恢复了卷曲/生物膜的表达，但超过一半的菌株只表现出部分或没有互补。此外，携带完整 mlrA 的两个菌株被发现生物膜形成能力不足。然而，我们发现，削弱或失活 RpoS 依赖性功能（如生物膜形成）的 RpoS 突变存在于超过 70%的菌株中，包括两个带有完整 mlrA 的菌株。我们得出结论，噬菌体中断 mlrA 和 RpoS 突变是限制大多数 O157:H7 血清型菌株卷曲表达和生物膜形成的主要障碍。
