School of Life Science and Engineering, Lanzhou University of Technologygrid.411291.e, Lanzhou, China.
School of Petrochemical Engineering, Lanzhou University of Technologygrid.411291.e, Lanzhou, China.
Appl Environ Microbiol. 2022 Nov 8;88(21):e0058722. doi: 10.1128/aem.00587-22. Epub 2022 Oct 18.
The molecular mechanism of the Ca-mediated formation of competent cells in Escherichia coli remains unclear. In this study, transcriptome and proteomics techniques were used to screen genes in response to Ca treatment. A total of 333 differentially expressed genes (317 upregulated and 16 downregulated) and 145 differentially expressed proteins (54 upregulated and 91 downregulated) were obtained. These genes and proteins are mainly enriched in cell membrane components, transmembrane transport, and stress response-related functional terms. Fifteen genes with these functions, including , , and , are speculated to play a key role in the cellular response to Ca. Three single-gene deletion strains were constructed with the Red homologous recombination method to verify its function in genetic transformation. The transformation efficiencies of , , and deletion strains for different-size plasmids were significantly increased. None of the three gene deletion strains changed in size, which is one of the main elements of microscopic morphology, but they exhibited different membrane permeabilities and transformation efficiencies. This study demonstrates that Ca-mediated competence formation in E. coli is not a simple physicochemical process and may involve the regulation of genes in response to Ca. This study lays the foundation for further in-depth analyses of the molecular mechanism of Ca-mediated transformation. Using transcriptome and proteome techniques and association analysis, we identified several key genes involved in the formation of Ca-mediated E. coli DH5α competent cells. We used Red homologous recombination technology to construct three single-gene deletion strains and found that the transformation efficiencies of , , and deletion strains for different-size plasmids were significantly increased. These results proved that the genetic transformation process is not only a physicochemical process but also a reaction process involving multiple genes. These results suggest ways to improve the horizontal gene transfer mechanism of foodborne microorganisms and provide new ideas for ensuring the safety of food preservation and processing.
Ca 介导的大肠杆菌感受态细胞形成的分子机制尚不清楚。本研究采用转录组学和蛋白质组学技术筛选 Ca 处理响应的基因。共获得 333 个差异表达基因（317 个上调和 16 个下调）和 145 个差异表达蛋白（54 个上调和 91 个下调）。这些基因和蛋白主要富集在细胞膜成分、跨膜转运和应激反应相关功能术语中。推测具有这些功能的 15 个基因，包括 、 、和 ，在细胞对 Ca 的响应中发挥关键作用。采用 Red 同源重组方法构建了三个单基因缺失菌株，以验证其在遗传转化中的功能。 、 、和 缺失菌株对不同大小质粒的转化效率均显著提高。三个基因缺失菌株的大小均未发生变化，这是微观形态的主要元素之一，但它们表现出不同的膜通透性和转化效率。本研究表明，Ca 介导的大肠杆菌感受态形成不是一个简单的物理化学过程，可能涉及 Ca 响应基因的调控。本研究为进一步深入分析 Ca 介导转化的分子机制奠定了基础。
使用转录组学和蛋白质组学技术和关联分析，我们鉴定了参与 Ca 介导的大肠杆菌 DH5α 感受态细胞形成的几个关键基因。我们使用 Red 同源重组技术构建了三个单基因缺失菌株，发现 、 、和 缺失菌株对不同大小质粒的转化效率均显著提高。这些结果证明遗传转化过程不仅是一个物理化学过程，也是一个涉及多个基因的反应过程。这些结果为改善食源性微生物的水平基因转移机制提供了思路，为保证食品保鲜和加工安全提供了新的思路。
