The Hippo kinase cascade regulates a contractile cell behavior and cell density in a close unicellular relative of animals.
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The coWts kinase domain was deleted from both alleles.
"For each gene, homologous recombination was used to replace the kinase domain of both WT alleles with selectable markers"
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coWts loss increases nuclear coYki, more strongly than coHpo loss.
"Interestingly, we found that coWts-/- cells were significantly more likely to show nuclear mScarlet-coYki localization than coHpo-/- cells (Figure 1D)"
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coWts is not a negative regulator of proliferation in Capsaspora.
"this result suggests that coWts is not a negative regulator of cell proliferation as in Drosophila and mammals, but instead is required for normal cell viability under some growth conditions"
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coWts loss produces elongated contractile cells, rescued by a coWts transgene.
"The presence of elongated cells was rescued by transgenic expression of coHpo or coWts in the respective mutant (Figure 3A and B), demonstrating that the elongated cell phenotype was specifically due to the absence of these kinases."
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coWts mutant aggregates are less circular and more densely packed.
"Whereas the number of cells per unit area in coYki -/- aggregates was not significantly different from WT, the number of cells per area in coHpo -/- and coWts -/- was significantly increased relative to WT (Figure 5E)."
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coWts mutant cells have shorter filopodia within aggregates.
"Whereas filopodia from coHpo -/- cells resembled those of WT and often extended more than a cell diameter distance within the aggregate, filopodia from coWts -/- cells were much shorter"
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The phenotypes depend on coYki transcriptional activity; epistasis could not be tested.
"Since techniques are currently unavailable to create double mutants in Capsaspora, we could not test this possibility through classic genetic epistasis."