Figure 1.
The β-CTT differentially regulates budding yeast kinesin-5 motors. (A) Top: Cartoon model depicting the force contribution of spindle midzone components and methods used to quantify spindle localization. Kar3 and Ase1 are thought to contribute inward forces, while Kip3, Cin8, and Kip1 to outward forces. A box was drawn around the spindle as defined by spindle pole body fluorescence (magenta) and the spindle fluorescence intensity on the spindle was quantified (green). A second box in the cytoplasm was used for background subtraction. Bottom: The amino acid sequence of the Tub2 CTT, which starts at E431, and the deletion in the tub2-Δ430 allele. (B) Quantification (top) and example image (bottom) of Kar3-mNeonGreen background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 102 cells, tub2-Δ430 n = 99 cells, P = 0.67. (C) Quantification (top) and example image (bottom) of Ase1-GFP background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 172 cells, tub2-Δ430 n = 162 cells, P = 0.48. (D) Quantification (top) and example image (bottom) of Kip3-mNeonGreen background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 90 cells, tub2-Δ430 n = 96 cells, P = 0.01. (E) Quantification (top) and example image (bottom) of Cin8-3GFP background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 104 cells, tub2-Δ430 n = 117 cells, P < 0.0001. (F) Quantification (top) and example image (bottom) of Kip1-mNeonGreen background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 136 cells, tub2-Δ430 n = 116 cells, P = 0.0049. The spindle poles in all cells are labeled with Spc110-tdTomato. For each graph in B–F, values are normalized to the median wild-type value for each technical replicate. Bolded, outlined points represent the median for each replicate. Error bars are the mean ± 95% CI for the replicate medians. Statistics are Student’s t test between the replicate medians for each protein compared to wild-type. * indicates P < 0.05; ** indicates P < 0.01. Scale bars = 1 µm.

The β-CTT differentially regulates budding yeast kinesin-5 motors. (A) Top: Cartoon model depicting the force contribution of spindle midzone components and methods used to quantify spindle localization. Kar3 and Ase1 are thought to contribute inward forces, while Kip3, Cin8, and Kip1 to outward forces. A box was drawn around the spindle as defined by spindle pole body fluorescence (magenta) and the spindle fluorescence intensity on the spindle was quantified (green). A second box in the cytoplasm was used for background subtraction. Bottom: The amino acid sequence of the Tub2 CTT, which starts at E431, and the deletion in the tub2-Δ430 allele. (B) Quantification (top) and example image (bottom) of Kar3-mNeonGreen background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 102 cells, tub2-Δ430 n = 99 cells, P = 0.67. (C) Quantification (top) and example image (bottom) of Ase1-GFP background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 172 cells, tub2-Δ430 n = 162 cells, P = 0.48. (D) Quantification (top) and example image (bottom) of Kip3-mNeonGreen background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 90 cells, tub2-Δ430 n = 96 cells, P = 0.01. (E) Quantification (top) and example image (bottom) of Cin8-3GFP background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 104 cells, tub2-Δ430 n = 117 cells, P < 0.0001. (F) Quantification (top) and example image (bottom) of Kip1-mNeonGreen background-subtracted spindle fluorescence in the presence or absence of the β-CTT. TUB2 n = 136 cells, tub2-Δ430 n = 116 cells, P = 0.0049. The spindle poles in all cells are labeled with Spc110-tdTomato. For each graph in B–F, values are normalized to the median wild-type value for each technical replicate. Bolded, outlined points represent the median for each replicate. Error bars are the mean ± 95% CI for the replicate medians. Statistics are Student’s t test between the replicate medians for each protein compared to wild-type. * indicates P < 0.05; ** indicates P < 0.01. Scale bars = 1 µm.

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