Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan.
Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan.
Appl Environ Microbiol. 2020 Nov 10;86(23). doi: 10.1128/AEM.02080-20.
Extracellular DNA (eDNA) is a biofilm component that contributes to the formation and structural stability of biofilms. , a major cariogenic bacterium, induces eDNA-dependent biofilm formation under specific conditions. Since cell death can result in the release and accumulation of DNA, the dead cells in biofilms are a source of eDNA. However, it remains unknown how eDNA is released from dead cells and is localized within biofilms. We focused on cell death induced by the extracellular signaling peptide called competence-stimulating peptide (CSP). We demonstrate that nucleic acid release into the extracellular environment occurs in a subpopulation of dead cells. eDNA production induced by CSP was highly dependent on the gene, which encodes an autolysin. Although expression was induced bimodally by CSP, -expressing cells further divided into surviving cells and eDNA-producing dead cells. Moreover, we found that -expressing cells were abundant near the bottom of the biofilm, even when all cells in the biofilm received the CSP signal. Dead cells and eDNA were also abundantly present near the bottom of the biofilm. The number of -expressing cells in biofilms was significantly higher than that in planktonic cultures, which suggests that adhesion to the substratum surface is important for the induction of expression. The deletion of resulted in reduced adherence to a polystyrene surface. These results suggest that expression and eDNA production induced near the bottom of the biofilm contribute to a firmly attached and structurally stable biofilm. Bacterial communities encased by self-produced extracellular polymeric substances (EPSs), known as biofilms, have a wide influence on human health and environmental problems. The importance of biofilm research has increased, as biofilms are the preferred bacterial lifestyle in nature. Furthermore, in recent years it has been noted that the contribution of phenotypic heterogeneity within biofilms requires analysis at the single-cell or subpopulation level to understand bacterial life strategies. In , a cariogenic bacterium, extracellular DNA (eDNA) contributes to biofilm formation. However, it remains unclear how and where the cells produce eDNA within the biofilm. We focused on LytF, an autolysin that is induced by extracellular peptide signals. We used single-cell level imaging techniques to analyze expression in the biofilm population. Here, we show that generates eDNA by inducing expression near the bottom of the biofilm, thereby enhancing biofilm adhesion and structural stability.
细胞外 DNA (eDNA) 是生物膜的组成部分，有助于生物膜的形成和结构稳定性。作为一种主要的致龋菌，在特定条件下诱导依赖于 eDNA 的生物膜形成。由于细胞死亡会导致 DNA 的释放和积累，生物膜中的死细胞是 eDNA 的来源。然而，eDNA 如何从死细胞中释放并在生物膜内定位仍然未知。我们专注于细胞死亡，这种死亡是由一种叫做感应肽（CSP）的细胞外信号肽诱导的。我们证明，核酸释放到细胞外环境发生在死细胞的一个亚群中。CSP 诱导的 eDNA 产生高度依赖于 基因，该基因编码一种自溶素。尽管 CSP 诱导 表达呈双峰模式，但 -表达细胞进一步分为存活细胞和产生 eDNA 的死细胞。此外，我们发现 -表达细胞在生物膜底部附近大量存在，即使生物膜中的所有细胞都接收到 CSP 信号。死细胞和 eDNA 也在生物膜底部附近大量存在。生物膜中 -表达细胞的数量明显高于浮游培养物中的数量，这表明粘附到基质表面对于 表达的诱导很重要。 的缺失导致对聚苯乙烯表面的粘附减少。这些结果表明，生物膜底部诱导的 表达和 eDNA 产生有助于形成牢固附着和结构稳定的生物膜。被称为生物膜的自我产生的细胞外聚合物物质（EPS）包裹的细菌群落对人类健康和环境问题有广泛的影响。生物膜研究的重要性增加了，因为生物膜是自然界中细菌首选的生活方式。此外，近年来，人们注意到生物膜内表型异质性的贡献需要在单细胞或亚群水平上进行分析，以了解细菌的生活策略。在 中，一种致龋菌，细胞外 DNA (eDNA) 有助于生物膜的形成。然而，仍然不清楚细胞在生物膜内如何以及在哪里产生 eDNA。我们专注于 LytF，一种由细胞外肽信号诱导的自溶素。我们使用单细胞水平成像技术来分析生物膜群体中的 表达。在这里，我们表明 通过在生物膜底部附近诱导 表达来产生 eDNA，从而增强生物膜的粘附和结构稳定性。
