The regulation of c-di-AMP homeostasis and signaling in bacteria

项目总结 小核苷酸c-diamp是一种普遍存在的第二信使，由数以千计的细菌和 原始族物种。许多细菌，特别是Firmicuts门中的细菌，需要c-diAMP才能在 标准实验室介质和c-diAMP重要性的机理已被广泛研究。 然而，在大量产生c-diamp的细菌中，包括对c-diamp完全 可有可无的、不受监管的c-diAMP积累是有毒的，会削弱细菌的生长和应激反应。 因此，c-di-AMP的动态平衡对产生c-diAMP的细菌至关重要。然而，监管的机制 细菌细胞中c-di-AMP水平及c-di-AMP调节不同细胞途径的机制 要实现最佳的生理学，两者都鲜为人知。我的实验室发现c-di-AMP水平在 C-di-AMP水解酶是调节c-di-AMP水平的关键机制。此外，我们还拥有 产生了缺乏c-diamp磷酸二酯酶的细菌突变体，称为pde突变体，并进行了广泛的研究 他们的表型缺陷。在接下来的五年中，我们将研究c-di-AMP信令的两个主要方面：i) C-diAMP磷酸二酯酶的信号检测和调控机制，以及II)通过 C-di-AMP协调其分子靶标以实现最佳的生理和应激反应。对于 第二个主题，我们将采取两种互补的方式。首先，我们将阐明c-蛋白的生物学功能。 基于目标表型、遗传和生化分析的DI-AMP分子靶标。第二，我们将确定 通过遗传适合度分析，研究了pde突变体毒性表型的分子靶标。我们的 这些发现将揭示c-di-amp信号的基本机制，这些机制将提供一种 了解c-diamp在不同细菌物种的生理中的全球调节作用的框架。

PROJECT SUMMARY The small nucleotide c-di-AMP is a ubiquitous second messenger produced by thousands of bacterial and archeal species. Many bacteria, especially those in the Firmicutes phylum, require c-di-AMP for growth in standard laboratory media, and the mechanisms for c-di-AMP essentiality have been extensively studied. However, in a wide range of c-di-AMP-producing bacteria, including those for which c-di-AMP is completely dispensable, unregulated c-di-AMP accumulation is toxic, diminishing bacterial growth and stress response. Thus, c-di-AMP homeostasis is critical for bacteria that produce it. Nevertheless, the mechanisms that regulate c-di-AMP levels in the bacterial cell, and the mechanisms by which c-di-AMP regulates different cellular pathways to achieve optimal physiology, are both poorly understood. My lab found that c-di-AMP levels are dynamic in the bacterial cell, and c-di-AMP hydrolysis is a key mechanism to modulate c-di-AMP levels. Furthermore, we have generated bacterial mutants lacking c-di-AMP phosphodiesterases, called pde mutants, and extensively studied their phenotypic defects. In the next five years, we will investigate two major aspects of c-di-AMP signaling: i) the signal detection and regulatory mechanisms of c-di-AMP phosphodiesterase, and ii) the mechanisms by which c-di-AMP coordinates its molecular targets to achieve optimal physiology and stress response. For the second theme, we will take two complementary approaches. One, we will elucidate the biological functions of c- di-AMP molecular targets based on targeted phenotypic, genetic, and biochemical analyses. Two, we will identify the molecular targets underlying the toxicity phenotypes of pde mutants, through genetic fitness assays. Our findings will reveal fundamental mechanisms of c-di-AMP signaling, and these mechanisms will provide a framework to understand the global regulatory roles of c-di-AMP in the physiology of different bacterial species.