Large-scale cell-type-specific imaging of protein synthesis in a vertebrate brain
| Autor: | Erin M. Schuman, Or David Shahar |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
Nervous system
translation control protein synthesis QH301-705.5 Science translation Endogeny Nerve Tissue Proteins neuron specific General Biochemistry Genetics and Molecular Biology Animals Genetically Modified 03 medical and health sciences 0302 clinical medicine In vivo medicine Protein biosynthesis Animals Biology (General) Zebrafish 030304 developmental biology 0303 health sciences General Immunology and Microbiology biology Chemistry General Neuroscience Brain Translation (biology) General Medicine Cell Biology Zebrafish Proteins cell type specific biology.organism_classification zebrafish Cell biology Tools and Resources medicine.anatomical_structure Glycine Click chemistry Medicine 030217 neurology & neurosurgery Neuroscience |
| Zdroj: | eLife, Vol 9 (2020) eLife Elife |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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