Cortical actin networks induce spatio-temporal confinement of phospholipids in the plasma membrane--a minimally invasive investigation by STED-FCS

Sci Rep. 2015 Jun 29:5:11454. doi: 10.1038/srep11454.

Abstract

Important discoveries in the last decades have changed our view of the plasma membrane organisation. Specifically, the cortical cytoskeleton has emerged as a key modulator of the lateral diffusion of membrane proteins. Cytoskeleton-dependent compartmentalised lipid diffusion has been proposed, but this concept remains controversial because this phenomenon has thus far only been observed with artefact-prone probes in combination with a single technique: single particle tracking. In this paper, we report the first direct observation of compartmentalised phospholipid diffusion in the plasma membrane of living cells using a minimally invasive, fluorescent dye labelled lipid analogue. These observations were made using optical STED nanoscopy in combination with fluorescence correlation spectroscopy (STED-FCS), a technique which allows the study of membrane dynamics on a sub-millisecond time-scale and with a spatial resolution of down to 40 nm. Specifically, we find that compartmentalised phospholipid diffusion depends on the cortical actin cytoskeleton, and that this constrained diffusion is directly dependent on the F-actin branching nucleator Arp2/3. These findings provide solid evidence that the Arp2/3-dependent cortical actin cytoskeleton plays a pivotal role in the dynamic organisation of the plasma membrane, potentially regulating fundamental cellular processes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actin-Related Protein 2-3 Complex / metabolism
  • Actins / metabolism*
  • Animals
  • Cell Line
  • Cell Membrane / metabolism*
  • Computer Simulation
  • Diffusion
  • Embryo, Mammalian / cytology
  • Fibroblasts / metabolism
  • Gene Knockdown Techniques
  • Mice
  • Phospholipids / metabolism*
  • Rats
  • Reproducibility of Results
  • Spatio-Temporal Analysis*
  • Spectrometry, Fluorescence / methods*

Substances

  • Actin-Related Protein 2-3 Complex
  • Actins
  • Phospholipids