Photoactivatable-GFP-α-tubulin as a tool to study microtubule plus-end turnover in living human cells

Methods Mol Biol. 2011:777:223-33. doi: 10.1007/978-1-61779-252-6_16.

Abstract

The development of photactivatable (PA) variants of Green fluorescent protein (GFP) has allowed the dynamics of spatially restricted protein pools within living cells to be determined. Over the last 5 years, experiments utilizing PA-GFP fused to α-tubulin have provided important insights into the mechanisms that control microtubule dynamics in living cells. In this chapter, we describe the methodology required to generate stable cell lines expressing photoactivatable-GFP-α-tubulin and to derive quantitative measurements of tubulin turnover at microtubules plus-ends in living cells.

MeSH terms

  • Cell Line
  • Green Fluorescent Proteins / chemistry
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism*
  • Humans
  • Microtubules / chemistry
  • Microtubules / genetics
  • Microtubules / metabolism*
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism*
  • Tubulin / chemistry
  • Tubulin / genetics
  • Tubulin / metabolism*

Substances

  • Recombinant Fusion Proteins
  • Tubulin
  • Green Fluorescent Proteins