Microtubule Nucleation Promoters Mto1 and Mto2 Regulate Cytokinesis in Fission Yeast

Abstract

Dundon and Pollard show that compromising the Mto1 or Mto2 regulators of the fission yeast γtubulin complex reduces or eliminates astral microtubules, exaggerates the effects of a non-lethal D277N substitution in β-glucan synthase 1 (Cps1/Bgs1) on the rate of cytokinetic furrow formation, and increases Rho-GTP at the cleavage site. AbstractMicrotubules of the mitotic spindle direct cytokinesis in metazoans but this has not been documented in fungi. We report evidence that astral microtubules help coordinate cytokinetic furrow formation in fission yeast. The temperature-sensitive cps1-191 strain (Liu et al., 1999) with a D277N substitution in β-glucan synthase 1 (Cps1/Bgs1) was reported to arrest with an unconstricted contractile ring. We discovered that rings in cps1-191 cells do constrict slowly and that an S338N mutation in the mto2 gene is required with the bgs1 D277N mutation to reproduce the cps1-191 phenotype. Complexes of Mto2 and Mto1 with γ-tubulin regulate microtubule assembly. Deletion of Mto1 along with the bgs1 D277N mutation also gives the cps1-191 phenotype, which is not observed in mto2 S338N or mto1Δ cells expressing bgs1 + . Both mto2 S338N and mto1Δ cells nucleate fewer astral microtubules than normal and have higher levels of Rho1-GTP at the division site than wild-type cells. We report multiple conditions that sensitize mto1Δ and mto2 S338N cells to furrow ingression phenotypes. (159 words)

Inhibition of cell membrane ingression at the division site by cell walls in fission yeast

“…To test if the turgor pressure inhibited ring contraction in cps1-191 mutants, we cultured cps1-191 cells in EMMA medium containing 0.8 M sorbitol to decrease the turgor pressure to that inside the cells at the restrictive temperature, and we added 2-deoxyglucose (2-DG) to this culture to prevent further glucan synthesis at the division site and elsewhere in the cells (Megnet, 1965;Svoboda and Smith, 1972;Osumi et al, 1998). A recent study showed that rings in cps1-191 mutant cells constricted slowly after shifting to the restrictive temperature for ∼2 h before microscopy at the restrictive temperature (Dundon and Pollard, 2020). To ensure a highly penetrant phenotype for cps1-191, we shifted the cps1-191 cells to the restrictive temperature for ∼6 h before microscopy, which was performed at the restrictive temperature.…”

“…The miscoordination of cytofission and nuclear division spatially could arise from the variable dumbbell-shaped morphology of cps1-191 spheroplasts (Mishra et al, 2012). The functions of Mto1 and Mto2, which are involved in the assembly of postanaphase microtubule arrays, may also contribute to these miscoordination defects (Dundon and Pollard, 2020). Because the cps1-191 mutant spheroplast division was morphologically different from normal fission yeast cell division and was mimicking the morphological changes of some animal cells during division, we have called this type of division cytofission.…”