Genome-wide analysis of RNA and protein localization and local translation in mESC-derived neurons

Methods. 2019 Jun 1;162-163:31-41. doi: 10.1016/j.ymeth.2019.02.002. Epub 2019 Feb 8.

Abstract

The subcellular localization and translation of mRNAs are fundamental biological processes. In neurons, they underlie cell growth and synaptic plasticity, which serves as a foundation of learning and memory. Multiple approaches have been developed to separate neurons on subcellular compartments - cell bodies (soma) and cell extensions (axons and dendrites) - for further biochemical analyses. Here we describe neurite/soma separation approach in combination with RNA sequencing and proteomic analyses to identify localized and locally translated RNAs and proteins. This approach allows quantification of around 7000 of local proteins and the entire local transcriptome. It provides a powerful tool for investigation of the mechanisms underlying RNA localization and local translation in neurons.

Keywords: Local translation; MRNA localization; Neuron; Non-coding RNAs; RNA-binding proteins (RBPs).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Basic Helix-Loop-Helix Transcription Factors / genetics
  • Basic Helix-Loop-Helix Transcription Factors / metabolism
  • Cell Culture Techniques / methods
  • Cell Differentiation
  • Cell Line
  • High-Throughput Nucleotide Sequencing
  • Mice
  • Mouse Embryonic Stem Cells
  • Neurons / metabolism*
  • Protein Biosynthesis
  • Proteomics / methods*
  • RNA, Messenger / analysis*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • RNA-Seq / methods*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism

Substances

  • Ascl1 protein, mouse
  • Basic Helix-Loop-Helix Transcription Factors
  • RNA, Messenger
  • Recombinant Proteins