Large-scale cell-type-specific imaging of protein synthesis in a vertebrate brain

• Main Authors: Or David Shahar, Erin Margaret Schuman
• Format: Article
• Language: English
• Published: eLife Sciences Publications Ltd 2020-02-01
• Series: eLife
• Subjects: translation; cell type specific; zebrafish; protein synthesis; translation control; neuron specific
• Online Access: https://elifesciences.org/articles/50564

Despite advances in methods to detect protein synthesis, it has not been possible to measure endogenous protein synthesis levels in vivo in an entire vertebrate brain. We developed a transgenic zebrafish line that allows for cell-type-specific labeling and imaging of nascent proteins in the entire animal. By replacing leucine with glycine in the zebrafish MetRS-binding pocket (MetRS-L270G), we enabled the cell-type-specific incorporation of the azide-bearing non-canonical-amino-acid azidonorleucine (ANL) during protein synthesis. Newly synthesized proteins were then labeled via 'click chemistry'. Using a Gal4-UAS-ELAV3 line to express MetRS-L270G in neurons, we measured protein synthesis intensities across the entire nervous system. We visualized endogenous protein synthesis and demonstrated that seizure-induced neural activity results in enhanced translation levels in neurons. This method allows for robust analysis of endogenous protein synthesis in a cell-type-specific manner, in vivo at single-cell resolution.