Ligand-directed two-step labeling to quantify neuronal glutamate receptor trafficking
Kento Ojima, Kazuki Shiraiwa, Tomohiro Doura, Mikiko Takato, Kazuhiro Komatsu, Michisuke Yuzaki, Itaru Hamachi, Shigeki Kiyonaka
Received Date: 19th May 20
The regulation of glutamate receptor localization is critical for development and synaptic plasticity in the central nervous system. Conventional biochemical and molecular biological approaches have been widely used to analyze glutamate receptor trafficking, especially for AMPA-type glutamate receptors (AMPARs). However, conflicting findings have been reported because of a lack of useful tools for analyzing endogenous AMPARs. Here, we develop a new method for the rapid and selective labeling of chemical probes to AMPARs by combining affinity-based protein labeling and bioorthogonal click chemistry under physiological conditions. This method allowed us to quantify AMPAR distribution and trafficking, which revealed some unique features of AMPARs, such as a long lifetime and a rapid recycling in neurons. This method was also successfully expanded to selectively label NMDA-type glutamate receptors. Thus, bioorthogonal two-step labeling may be a versatile tool for investigating the physiological and pathophysiological roles of glutamate receptors in neurons.
Read in full at bioRxiv.
This is an abstract of a preprint hosted on an independent third party site. It has not been peer reviewed but is currently under consideration at Nature Communications.