Quantifying protein dynamics and stability in a living organism

Nat Commun. 2019 Mar 12;10(1):1179. doi: 10.1038/s41467-019-09088-y.

Abstract

As an integral part of modern cell biology, fluorescence microscopy enables quantification of the stability and dynamics of fluorescence-labeled biomolecules inside cultured cells. However, obtaining time-resolved data from individual cells within a live vertebrate organism remains challenging. Here we demonstrate a customized pipeline that integrates meganuclease-mediated mosaic transformation with fluorescence-detected temperature-jump microscopy to probe dynamics and stability of endogenously expressed proteins in different tissues of living multicellular organisms.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Cell Line, Tumor
  • Embryo, Nonmammalian
  • Endodeoxyribonucleases / metabolism
  • Fluorescence Resonance Energy Transfer / instrumentation
  • Fluorescence Resonance Energy Transfer / methods
  • Fungal Proteins / chemistry
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism*
  • Humans
  • Intravital Microscopy / instrumentation
  • Intravital Microscopy / methods*
  • Kinetics
  • Microscopy, Fluorescence / instrumentation
  • Microscopy, Fluorescence / methods
  • Phosphoglycerate Kinase / chemistry
  • Phosphoglycerate Kinase / genetics
  • Phosphoglycerate Kinase / metabolism*
  • Protein Folding
  • Protein Stability
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Temperature
  • Zebrafish

Substances

  • Fungal Proteins
  • Recombinant Fusion Proteins
  • Phosphoglycerate Kinase
  • Endodeoxyribonucleases