基于 RNA 的荧光传感器对鸟苷四和五磷酸（p）ppGpp 的活细胞成像*。
Live-Cell Imaging of Guanosine Tetra- and Pentaphosphate (p)ppGpp with RNA-based Fluorescent Sensors*.
机构信息
Department of Chemistry, University of Massachusetts, Amherst, USA.
Department of Biochemistry and Molecular Biology, University of Massachusetts, Amherst, USA.
出版信息
Angew Chem Int Ed Engl. 2021 Nov 2;60(45):24070-24074. doi: 10.1002/anie.202111170. Epub 2021 Oct 5.
Abstract
Guanosine tetra- and pentaphosphate, (p)ppGpp, are important alarmone nucleotides that regulate bacterial survival in stressful environment. A direct detection of (p)ppGpp in living cells is critical for our understanding of the mechanism of bacterial stringent response. However, it is still challenging to image cellular (p)ppGpp. Here, we report RNA-based fluorescent sensors for the live-cell imaging of (p)ppGpp. Our sensors are engineered by conjugating a recently identified (p)ppGpp-specific riboswitch with a fluorogenic RNA aptamer, Broccoli. These sensors can be genetically encoded and enable direct monitoring of cellular (p)ppGpp accumulation. Unprecedented information on cell-to-cell variation and cellular dynamics of (p)ppGpp levels is now obtained under different nutritional conditions. These RNA-based sensors can be broadly adapted to study bacterial stringent response.
摘要
鸟苷四磷酸和五磷酸，（p）ppGpp，是调节细菌在应激环境中生存的重要警报核苷酸。直接检测活细胞中的（p）ppGpp 对于我们理解细菌严格反应的机制至关重要。然而，对细胞内（p）ppGpp 的成像仍然具有挑战性。在这里，我们报告了基于 RNA 的荧光传感器，用于活细胞中（p）ppGpp 的成像。我们的传感器是通过将最近鉴定的（p）ppGpp 特异性核糖开关与荧光 RNA 适体 Broccoli 偶联而成的。这些传感器可以进行基因编码，并能够直接监测细胞内（p）ppGpp 的积累。在不同的营养条件下，现在可以获得关于（p）ppGpp 水平的细胞间变异性和细胞动力学的前所未有的信息。这些基于 RNA 的传感器可以广泛适用于研究细菌严格反应。
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