Activity-Based Sensing with a Metal-Directed Acyl Imidazole Strategy Reveals Cell Type-Dependent Pools of Labile Brain Copper

Abstract

Copper is a required nutrient for life and particularly important to the brain and central nervous system. Indeed, copper redox activity is essential to maintaining normal physiological responses spanning neural signaling to metabolism, but at the same time copper misregulation is associated with inflammation and neurodegeneration. As such, chemical probes that can track dynamic changes in copper with spatial resolution, especially in loosely-bound, labile forms, are valuable tools to identify and characterize its contributions to healthy and disease states. In this report, we present an activity-based sensing (ABS) strategy for copper detection in live cells that preserves spatial information by a copper-dependent bioconjugation reaction. Specifically, we designed copper-directed acyl imidazole (CD) dyes that operate through copper-mediated activation of acyl imidazole electrophiles for subsequent labeling of proximal proteins at sites of elevated labile copper to provide a permanent stain that resists washing and fixation. To showcase the utility of this new ABS platform, we sought to characterize labile copper pools in the three main cell types in the brain: neurons, astrocytes, and microglia. Exposure of each of these cell types to physiologically relevant stimuli shows distinct changes in labile copper pools. Neurons display translocation of labile copper from somatic cell bodies to peripheral processes upon activation, whereas astrocytes and microglia exhibit global decreases and increases in intracellular labile copper pools, respectively, after exposure to inflam-matory stimuli. This work provides foundational information on cell type-dependent homeostasis of copper, an essential metal in the brain, as well as a starting point for the design of new activity-based probes for metals and other dynamic signaling and stress analytes in biology. File list (2) download file view on ChemRxiv CJC Copper DAI Main Text ChemRxiv.pdf (837.74 KiB) download file view on ChemRxiv CJC Copper DAI SI Rev Clean Final.pdf (760.44 KiB)

Neurodegenerative Disease Diagnosis via Ion‐Level Detection in the Brain

“…Recently, a copper‐directed acyl imidazole probe has been constructed and applied to monitor labile copper pools in neurons, astrocytes, and microglia, thereby contributing to the understanding of metal metabolism in cells. [ 43 ] For Zn 2+ detection, there is a two‐photon fluorescent probe Chromis‐1, the emission peak of which changes from 440 to 510 nm under the influence of Zn 2+ in NIH3T3 fibroblasts. [ 44 ] Moreover, a two‐photon fluorescence probe, P‐Zn, has been developed for monitoring Zn 2+ in live cell, hippocampal tissue, and zebrafish (Figure 2d).…”

Selective Detection of Cu+ Ions in Live Cells via Fluorescence Lifetime Imaging Microscopy

“…Over the last two decades several such fluorescent probes for copper(I) have been reported in the literature. [ 16 , 17 , 18 , 19 , 20 , 21 ] Most of these probes turn‐on their fluorescence intensity upon binding to copper(I) via a photoinduced electron transfer (PET) process or a related charge transfer (CT) pathway. [22] However, intensity‐based probes have the disadvantage that they are concentration dependent, that is, their signal is strongly dependent on probe uptake to a particular intracellular location.…”

Selective Detection of Cu+ Ions in Live Cells via Fluorescence Lifetime Imaging Microscopy

“…Over the last two decades several such fluorescent probes for copper(I) have been reported in the literature. [16][17][18][19][20][21] Most of these probes turn-on their fluorescence intensity upon binding to copper(I) via a photoinduced electron transfer (PET) process or a related charge transfer (CT) pathway. [22] However, intensity-based probes have the disadvantage that they are concentration dependent, that is, their signal is strongly dependent on probe uptake to a particular intracellular location.…”