A G protein-coupled receptor (GPCR) in red: live cell imaging of the kappa opioid receptor-tdTomato fusion protein (KOPR-tdT) in neuronal cells.

ABSTRACT:

Introduction

In contrast to green fluorescent protein and variants (GFPs), red fluorescent proteins (RFPs) have rarely been employed for the generation of GPCR fusion proteins, likely because of formation of aggregates and cell toxicity of some RFPs. Among all the RFPs, tdTomato (tdT), one of the non-aggregating RFP, has the highest brightness score (about 3 times that of eGFP) and unsurpassed photostability.

Methods

We fused tdT to the KOPR C-terminus. The KOPR-tdT cDNA construct was transfected into a Neuro2A mouse neuroblastoma cell line (Neuro2A cells) and rat cortical primary neurons for characterization of pharmacological properties and imaging studies on KOPR trafficking.

Results

KOPR-tdT retained KOPR properties (cell surface expression, ligand binding, agonis

SUBMITTER: Huang P 

PROVIDER: S-EPMC3954113 | biostudies-literature | 2013 Nov-Dec

REPOSITORIES: biostudies-literature