A novel form of motility in filopodia revealed by imaging myosin-X at the single-molecule level

Curr Biol. 2009 Jun 9;19(11):967-73. doi: 10.1016/j.cub.2009.03.067. Epub 2009 Apr 23.

Abstract

Although many proteins, receptors, and viruses are transported rearward along filopodia by retrograde actin flow, it is less clear how molecules move forward in filopodia. Myosin-X (Myo10) is an actin-based motor hypothesized to use its motor activity to move forward along actin filaments to the tips of filopodia. Here we use a sensitive total internal reflection fluorescence (TIRF) microscopy system to directly visualize the movements of GFP-Myo10. This reveals a novel form of motility at or near the single-molecule level in living cells wherein extremely faint particles of Myo10 move in a rapid and directed fashion toward the filopodial tip. These fast forward movements occur at approximately 600 nm/s over distances of up to approximately 10 microm and require Myo10 motor activity and actin filaments. As expected for imaging at the single-molecule level, the faint particles of GFP-Myo10 are diffraction limited, have an intensity range similar to single GFP molecules, and exhibit stepwise bleaching. Faint particles of GFP-Myo5a can also move toward the filopodial tip, but at a slower characteristic velocity of approximately 250 nm/s. Similar movements were not detected with GFP-Myo1a, indicating that not all myosins are capable of intrafilopodial motility. These data indicate the existence of a novel system of long-range transport based on the rapid movement of myosin molecules along filopodial actin filaments.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / physiology
  • Animals
  • Bridged Bicyclo Compounds, Heterocyclic / pharmacology
  • Cattle
  • Cell Movement / drug effects
  • Cell Movement / physiology*
  • Green Fluorescent Proteins / analysis
  • HeLa Cells
  • Humans
  • Microscopy, Fluorescence
  • Myosins / analysis
  • Myosins / physiology*
  • Myosins / ultrastructure
  • Pseudopodia / metabolism
  • Pseudopodia / physiology*
  • Pseudopodia / ultrastructure
  • Thiazolidines / pharmacology

Substances

  • Actins
  • Bridged Bicyclo Compounds, Heterocyclic
  • Thiazolidines
  • Green Fluorescent Proteins
  • Myosins
  • latrunculin B