Segregation of Molecules at Cell Division Reveals Native Protein Localization

Nat Methods. 2012 Apr 8;9(5):480-2. doi: 10.1038/nmeth.1955.

Abstract

We introduce a nonintrusive method exploiting single-cell variability after cell division to validate protein localization. We found that Clp proteases, widely reported to form biologically relevant foci, were uniformly distributed in Escherichia coli cells, and that many commonly used fluorescent proteins caused severe mislocalization when fused to homo-oligomers. Retagging five other reportedly foci-forming proteins with the most monomeric fluorescent protein tested suggests that the foci were caused by the fluorescent tags.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Cell Division / physiology
  • Endopeptidase Clp / genetics
  • Endopeptidase Clp / metabolism*
  • Escherichia coli / cytology
  • Escherichia coli / enzymology
  • Escherichia coli / metabolism*
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism*
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism*
  • Microscopy, Fluorescence
  • Microscopy, Phase-Contrast
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism*

Substances

  • Bacterial Proteins
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • yellow fluorescent protein, Bacteria
  • Green Fluorescent Proteins
  • Endopeptidase Clp