H+- and Na+- elicited rapid changes of the microtubule cytoskeleton in the biflagellated green alga Chlamydomonas

  1. Yi Liu
  2. Mike Visetsouk
  3. Michelle Mynlieff
  4. Hongmin Qin
  5. Karl F Lechtreck
  6. Pinfen Yang  Is a corresponding author
  1. Marquette University, United States
  2. Texas A&M University, United States
  3. University of Georgia, United States
7 figures, 6 videos, 1 table and 1 additional file

Figures

EB1 in Chlamydomonas.

(a) A schematic picture depicting flagella and the MT network in the cell body (top panel). Black dots, basal bodies (BB). Thick lines, four stable rootlet microtubule bundles. Thin lines in the …

https://doi.org/10.7554/eLife.26002.002
An HA pulse elicited swift sequential changes in the MT system.

(a) Schematics depicting an open-ended perfusion chamber (left panels) and a diffusion chamber (right panels) for capturing the HA-induced rapid changes. (b) A 10 μl aliquot of cells resuspended in …

https://doi.org/10.7554/eLife.26002.007
Abatements of HA-induced changes.

(a) Concentration-dependent HA effects on MTs in WT cells. WT cells resuspended in 5 mM HA exhibited a bird cage pattern with dynamic comets (top and middle panels). Shrinking MTs were visible in …

https://doi.org/10.7554/eLife.26002.008
HA bath and a subsequent wash induced long-lived yet reversible changes to the MT system.

(a) Static EB1 signals remained at BBs after cells were resuspended in 10 mM pH3 HA for 5 min (left panel). After replacing HA with the HEPES buffer, EB1 signal at the BB area intensified within 1 …

https://doi.org/10.7554/eLife.26002.010
Figure 5 with 1 supplement
The MT pattern in tub2 cells recovered from the HA bath.

(a) The frozen bird cage pattern of fine MTs remains after 5 min HA bath as shown in two representative cells (left panel). After wash, dynamics resumes within 60 s. Notably, comets (panel I and II, …

https://doi.org/10.7554/eLife.26002.012
Figure 5—figure supplement 1
(for Figure 5a) tub2 cells in 5 min HA bath.
https://doi.org/10.7554/eLife.26002.013
Na+-dependent changes of the MT system.

(a) MTs in cells were largely frozen after 5 min 10 mM pH3 HA bath and 3 min in the wash solution, such as 5 mM pH7.4 Na+/HEPES buffer or 5 mM NaCl solution (black arrowheads). In contrast, growing …

https://doi.org/10.7554/eLife.26002.015
Na+ concentration-dependent inhibition of cell division.

(a) Reduced abundance and delayed appearance of sporangia with newly divided daughter cells as a function of NaCl concentration in TAP media. Triplicate cell pellets from a typical log-phase culture …

https://doi.org/10.7554/eLife.26002.016

Videos

Video 1
(for Figure 1b) EB1-NG comets in WT cells.
https://doi.org/10.7554/eLife.26002.003
Video 2
(for Figure 1f1) Transient bird-cage pattern in WT cells that occurred sporadically during imaging.
https://doi.org/10.7554/eLife.26002.004
Video 3
(for Figure 1f2) Disappearance and return of comets in compressed cells following alternate periods of illumination and darkness.
https://doi.org/10.7554/eLife.26002.005
Video 4
(for Figure 3a) WT cells treated with 5 mM HA.
https://doi.org/10.7554/eLife.26002.009
Video 5
(for Figure 4a) WT cells in Na+/HEPES after HA bath.
https://doi.org/10.7554/eLife.26002.011
Video 6
(for Figure 5a) tub2 cells in Na+/HEPES after HA bath.
https://doi.org/10.7554/eLife.26002.014

Tables

Table 1
Summary of the treatments and corresponding patterns of microtubules (MTs) and EB1-NeonGreen (NG) in wild type and tub2 mutant cells.

Identical treatments were highlighted with a same gray shade for easy comparison. HA, acetic acid; TAP, Tris-Acetate-Phosphate culture medium.

https://doi.org/10.7554/eLife.26002.006
TreatmentsMT (revealed by EB1-NG)EB1-NG
WT
TAP or 5 mM Na+/HEPES (Control)Dynamic; cold labileComets
20 mM pH4.5 HA/TAP pulse (perfusion as pH shock)InvisibleNo comets
100 mM HA pulse (diffusion)ShrinkFibers; long comets
10 mM pH3.0 HAInvisible
7.5 mM pH3.4 HAShrinkFibers; no comets
5.0 mM pH3.5 HADynamicFibers; comets
10 mM pH3.0 HA bath; 5 mM [Na+]ex washFreeze; bundle/branch; cold resistantFibers; no comets
10 mM pH3.0 HA bath; 5 mM [K+]ex washDynamicComets
21 mM Na+/EGTAFreeze; bundle/branch; cold resistantFibers; no comets
21 mM K+/EGTADynamicComets
55 mM [Na+]exDynamicLong comets
150 mM [Na+]exAbsence or a few low dynamic MTsFibers; no comets
TAP + 55 mM [Na+]exDynamicFibers; long comets
TAP + 100 mM [Na+]exDynamicFibers; long comets
TAP + 200 mM [Na+]exAbsence or a few low dynamic MTsFibers; no comets
tub2 (colchicine-resistant)
TAP or 5 mM Na+/HEPESDynamicComets
10 mM pH3.0 HAFreezeFibers; no comets
10 mM pH3.0 HA bath; 5 mM [Na]ex washFreeze; bundle/branchFibers; no comets

Additional files

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