Heterogeneous gene expression, metabolic variability and differentiation in Staphylococcus epidermidis biofilms

表皮葡萄球菌是一种形成生物膜的原型细菌，其特征是生物膜群落中存在明显的表型变异。本研究旨在了解表皮葡萄球菌生物膜异质性的生物学意义以及触发和控制异质性的因素。在之前的资助时期，我们发现，在表皮葡萄球菌生物被膜种群中，表型变异共存。它们在生物膜基质的产生和代谢特性方面有所不同，有不同的细菌亚群经历DNA的裂解和释放。此外，生物膜阴性变异体在生物膜内出现的频率很高。激光共聚焦扫描显微镜(CLSM)分析揭示了表皮葡萄球菌生物膜的有序空间结构，具有显著程度的异质性基因表达模式，并在生物膜的明确区域集中了各种亚群。最引人注目的是，一个小的非编码RNA(即RsaE)被认为是表皮葡萄球菌生物被膜群落产生异质性的触发因素。因此，RsaE被认为介导了蛋白质和多糖生物膜基质产生之间的(可逆)转换，很可能是通过重新编程葡萄球菌的新陈代谢。同时，该分子也可能能够限制导致生物膜异质性的遗传事件的频率。由于RSAE本身是异质性表达的，我们建议这种核转录调节因子作为产生和控制生物膜多样性的重要驱动因素发挥作用。为了证实这一假设，我们将在即将到来的资金阶段，特别关注调节表皮葡萄球菌生物膜多样性的因素和分子机制。因此，我们将研究RsaE在这一过程中的分子功能，并研究其与其他重要调控电路的可能联系，如群体感应以及程序性细菌裂解和细胞死亡。此外，我们将继续利用共聚焦激光扫描显微镜分析生物膜发育的时空动力学，并将这些研究扩展到流室模型。最后，在与我们联盟内的合作者的共同努力下，我们将建立合适的模型来模拟和预测表皮葡萄球菌生物膜的发育和多样化。

Staphylococcus epidermidis is a prototype biofilm-forming bacterium characterised by a pronounced phenotypic variability within biofilm communities. The proposed research aims at an understanding of the biological significance and the factors that trigger and control heterogeneity in S. epidermidis biofilms. In the previous funding period, we established that, within S. epidermidis biofilm populations, phenotypic variants coexist. They vary in terms of biofilm matrix production and metabolic properties, with distinct subpopulations of bacteria undergoing lysis and release of DNA. In addition, biofilm-negative variants arise in high frequencies within the biofilm. Confocal laser scanning microscopy (CLSM) analyses revealed an organised spatial structure of the S. epidermidis biofilms with a significant degree of heterogeneus gene expression patterns and the concentration of various subpopulations in well-defined regions of the biofilm. Most strikingly, a small non-coding RNA (i.e., RsaE) was identified to act as a putative trigger for generating heterogeneity in S. epidermidis biofilm communities. Thus, RsaE is supposed to mediate the (reversible) switch between protein- and polysaccharide-biofilm matrix production, most likely through re-programing of the staphylococcal metabolism. At the same time, the molecule is possibly capable of limiting the frequency of genetic events contributing to biofilm heterogeneity as well. As RsaE itself is heterogeneously expressed, we suggest this riboregulator to function as an important driver for generating and controlling biofilm diversity. To substantiate this, hypothesis we will, in the upcoming funding period, specifically focus on the factors and molecular mechanisms that mediate diversity in S. epidermidis biofilms. Thus, we will investigate the molecular function of RsaE in the process, and study its putative link to other important regulatory circuits such as quorum sensing as well as programmed bacterial lysis and cell death. Moreover, we will continue to analyse the spatiotemporal dynamics of biofilm development by confocal laser scanning microscopy and extend these studies to flow chamber models. Finally, in an joint effort with our collaborators within the consortium, we will establish suitable models to simulate and predict S. epidermidis biofilm development and diversification.