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Rac1 activation can generate untemplated, lamellar membrane ruffles.

F Leyden1, S Uthishtran2, U K Moorthi2,3

  • 1Single Molecule Science, University of New South Wales, Sydney, Australia.

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|April 14, 2021
PubMed
Summary
This summary is machine-generated.

Optogenetics and lattice light-sheet microscopy control cell membrane protrusions. This study reveals small GTPase Rac1 activation drives actin assembly for sheet-like protrusion formation and macropinosome generation.

Keywords:
Actin polymerizationLamellipodiaLattice light-sheet microscopyOptogeneticsRac1

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Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biophysics

Background:

  • Dorsal membrane protrusions serve as models for studying cellular actin machinery.
  • Small GTPase Rac1 regulates membrane protrusions for cellular uptake processes like pinocytosis and phagocytosis.

Purpose of the Study:

  • To develop optogenetic methods for controlled generation of membrane ruffles using photoactivable Rac1 (PA-Rac1).
  • To investigate the dynamics of actin polymerization and protrusion formation driven by Rac1 activation.
  • To explore the role of Rac1 in macropinosome formation and identify proteins enriched in cellular ruffles.

Main Methods:

  • Utilized lattice light-sheet microscopy (LLSM) for rapid volumetric imaging.
  • Employed optogenetic control of photoactivable Rac1 (PA-Rac1) for precise spatial and temporal activation.
  • Performed multicolour imaging to track protein localization during protrusion formation and deactivation.

Main Results:

  • Demonstrated that continuous PA-Rac1 activation is necessary for sustained actin polymerization and ruffle generation, indicating a local concentration threshold.
  • Showed that Rac1 activation induces actin assembly at the cell membrane, leading to untemplated, sheet-like protrusion formation.
  • Observed that deactivation of PA-Rac1 triggers contractile processes resulting in macropinosome formation.
  • Identified Myo1e as a protein specifically enriched within these cellular ruffles using multicolour imaging.

Conclusions:

  • The combination of LLSM and optogenetics offers advanced control for studying dynamic cellular mechanisms.
  • Rac1 activation can generate lamellar protrusions independently of external templates, highlighting its role in actin-driven cell morphology.
  • The study provides insights into the molecular interplay governing dynamic actin machinery and cellular protrusion formation.