Surface-associated motility of Acinetobacter baumannii and its epigenetic control via AamA-specific DNA methylation

表面相关的运动性在鲍曼不动杆菌的临床分离株中广泛存在，并被认为有助于在医院环境中的毒性和持久性。在之前的资助期间，我们在mellonella感染模型中描述了一组超过30个动力缺陷突变体的运动性，生长，生物膜形成，抗生素耐药性和毒力。总的来说，我们的研究强调了运动性、生物膜和毒力特性的相互关系。特别有趣的是，我们发现亚硫酸盐还原酶活性是生物膜形成、毒力和抗生素耐药性的关键因素，这表明H2S是一种信号分子。此外，我们鉴定了一种新的DNA腺嘌呤甲基转移酶，我们对其进行了功能表征，并将其命名为AamA。我们首次使用单分子实时测序技术研究了鲍曼不动杆菌菌株的DNA甲基组，结果表明aama依赖的调控参与了运动性和对不同环境条件的响应。我们的模型是通过AamA介导的瞬时DNA甲基化对基因表达的表观遗传控制。为了验证我们的假设，我们将继续定义aama依赖性甲基组在不同环境条件下的反应，包括运动性、生物膜形成、渗透和抗生素应激。RT-PCR和RNA测序将用于定义反应并将其与特定的甲基化模式相关联。为了补充我们对AamA调控的理解，我们打算利用蛋白质组学方法来确定AamA相互作用伙伴并研究它们对DNA甲基化的影响。通过这种方式，我们希望有助于了解这种病原体的适应过程。

Surface-associated motility is widespread among clinical isolates of Acinetobacter baumannii and considered to contribute to virulence and persistence in the hospital environment. During the previous funding period we characterized a set of more than 30 motility-deficient mutants with respect to motility, growth, biofilm formation, antibiotic resistance and virulence in the Galleria mellonella infection model. Overall, our studies highlight the interrelatedness of motility, biofilm and virulence properties. Of particular interest, we identified sulfite reductase activity as a crucial factor of biofilm formation, virulence and antibiotic resistance suggesting a role of H2S as a signaling molecule. Moreover, we identified a novel DNA adenine methyltransferase which we functionally characterized and which we termed AamA. For the first time, we investigated the DNA methylome of an A. baumannii strain using single-molecule real time sequencing and our results suggest an AamA-dependent regulon involved in motility and responding to different environmental conditions. Our model is that of an epigenetic control of gene expression via transient DNA methylation mediated by AamA. To scrutinize our hypotheses, we would like to continue to define the AamA-dependent methylome in response to different environmental conditions including motility, biofilm formation, osmotic and antibiotic stress. RT-PCR and RNA sequencing will be applied to define the responses and correlate them with specific methylation patterns. To complement our understanding of the AamA regulon we intend to exploit proteomics approaches to identify AamA interaction partners and to study their influence on DNA methylation. In this way, we expect to contribute to understanding of adaptation processes of this pathogen.