A Fluorescent RNA-Based Genetically Encoded Mg2+ Sensor Allows Robust Detection of Mg2+ in Live Cells.
TL;DR
It is demonstrated that mammalian cells maintain Mg2+ homeostasis even under conditions of elevated extracellular Mg2+, revealing the robustness of intracellular regulatory mechanisms governing Mg2+ balance.
Abstract
Magnesium plays crucial roles in many biological processes and the stabilization of biomolecules, including DNA, RNA, and proteins. Despite significant progress, however, our understanding of how cells regulate Mg2+ homeostasis and transport remains incomplete. One of the goals is to develop approaches for detecting Mg2+ dynamics with high spatial and temporal resolution. Herein, we describe the development of MagFR, a ratiometric Mg2+ sensor based on Pepper and Clivia fluorescent RNAs. In this sensor, Pepper fluorescence is highly Mg2+ dependent, and Clivia fluorescence acts as the normalizer. MagFR has favourable properties, including a large dynamic range, pH insensitivity, and high selectivity, allowing robust detection of Mg2+ in both live bacterial and mammalian cells. By targeting MagFR to distinct subcellular compartments in mammalian cells, we observed that the free cytoplasmic Mg2+ concentration is slightly higher than that in the nucleoplasm. We demonstrated that mammalian cells maintain Mg2+ homeostasis even under conditions of elevated extracellular Mg2+, revealing the robustness of intracellular regulatory mechanisms governing Mg2+ balance. MagFR also enabled real-time detection of cellular Mg2+ dynamics following ATP depletion. Overall, this study offers a robust and versatile tool for imaging Mg2+ dynamics in cells, which will be useful for elucidating the functionality and mechanism of Mg2+ homeostasis underlying diverse cellular processes.