Cytoplasmic transport in keratocytes: Direct visualization of particle translocation along microtubules

John H. Hayden*, Robert D. Allen, Robert Goldman

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

49 Scopus citations

Abstract

We report the first direct demonstration that the cytoplasmic transport of organelles and vesicles (collectively called particles) takes place along microtubules. Living keratocytes from the corneal stroma of the frog, Rana pipiens, were observed with Allen video‐enhanced constrast, differential interference constrast (AVEC‐DIC) microscopy [Allen et al, 1981]. In sufficiently thin regions of these cells a network of linear elements was visible. When particles were observed in motion, they always moved along these linear elements. The linear elements remained intact and in focus on the microscope when lysed in a cell lysis solution that stabilized microtubules. Preparations were then fixed in formaldehyde, washed with phosphate‐buffered saline (PBS), incubated with rabbit antitubulin, washed with PBS, stained with rhodamine‐conjugated goat antirabbit, and washed with PBS. The extracted cells continued to remain in place and in focus on the microscope throughout these procedures. The same cells were then observed using epifluorescence optics and a silicon‐intensified target (SIT) video camera. A network of fluorescent linear elements was seen to correspond in number, form, and position to the linear elements seen in the live AVEC‐DIC image. Taken together, the AVEC‐DIC and fluorescence microscopy observations prove that the linear elements along which particles move are microtubules (MTLEs). The observed particle speeds, pause times, and distances moved varied widely, even for the same particle on the same microtubule. Particles were also observed to switch from one microtubule to another as they were transported. The polarity of the microtubules did not seem to affect the particle direction, since particles were observed to move in both directions on the same MTLE. When not in motion these particles behaved as if anchored to the microtubules since they showed negligible Brownian motion. Finally, it was observed that an elongate particle could move onto two intersecting linear elements such that it was deformed into an inverted “Y” shape. This indicates that there may be more than a single site of attachment between the force generator and the particle.

Original languageEnglish (US)
Pages (from-to)1-19
Number of pages19
JournalCell Motility
Volume3
Issue number1
DOIs
StatePublished - Jan 1 1983

Keywords

  • Saltation
  • cytoplasmic transport
  • keratocytes
  • microtubules

ASJC Scopus subject areas

  • Cell Biology

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