A DNA-Based FLIM Reporter for Simultaneous Quantification of Lysosomal pH and Ca2+ during Autophagy Regulation

Zhonghui Zhang; Zhichao Liu; Yang Tian

doi:10.1016/j.isci.2020.101344

A DNA-Based FLIM Reporter for Simultaneous Quantification of Lysosomal pH and Ca²⁺ during Autophagy Regulation

iScience. 2020 Jul 24;23(7):101344. doi: 10.1016/j.isci.2020.101344. Epub 2020 Jul 4.

Authors

Zhonghui Zhang¹, Zhichao Liu², Yang Tian³

Affiliations

¹ State Key Laboratory of Precision Spectroscopy, East China Normal University, Dongchuan Road 500, Shanghai 200241, China; Shanghai Key Laboratory of Green Chemistry and Chemical Processes, School of Chemistry and Molecular Engineering, East China Normal University, Dongchuan Road 500, Shanghai 200241, China.
² Shanghai Key Laboratory of Green Chemistry and Chemical Processes, School of Chemistry and Molecular Engineering, East China Normal University, Dongchuan Road 500, Shanghai 200241, China. Electronic address: zcliu@chem.ecnu.edu.cn.
³ State Key Laboratory of Precision Spectroscopy, East China Normal University, Dongchuan Road 500, Shanghai 200241, China; Shanghai Key Laboratory of Green Chemistry and Chemical Processes, School of Chemistry and Molecular Engineering, East China Normal University, Dongchuan Road 500, Shanghai 200241, China. Electronic address: ytian@chem.ecnu.edu.cn.

Abstract

pH and Ca²⁺ play important roles in regulating lysosomal activity and lysosome-mediated physiological and pathological processes. However, effective methods for simultaneous determination of pH and Ca²⁺ is the bottleneck. Herein, a single DNA-based FLIM reporter was developed for real-time imaging and simultaneous quantification of pH and Ca²⁺ in lysosomes with high affinity, in which a specific probe for recognition of Ca²⁺ was assembled onto a DNA nanostructure together with pH-responsive and lysosome-targeted molecules. The developed DNA reporter showed excellent biocompatibility and long-term stability up to ∼56 h in lysosomes. Using this powerful tool, it was discovered that pH was closely related to Ca²⁺ concentration in lysosome, whereas autophagy can be regulated by lysosomal pH and Ca²⁺. Furthermore, Aβ-induced neuronal death resulted from autophagy abnormal through lysosomal pH and Ca²⁺ changes. In addition, lysosomal pH and Ca²⁺ were found to regulate the transformation of NSCs, resulting in Rapamycin-induced antiaging.

Keywords: Cellular Neuroscience; Optical Imaging; Technical Aspects of Cell Biology.