Figure 3. Assembly of microtubule binding subcomplexes Dam1 and Ndc80 is affected by loss of Fin1. (a) The MIND submodule component Dsn1 is not significantly affected by deletion of FIN1 or FIN1 and CNN1. The normalized intensity of GFP-tagged Dsn1 protein was quantified in G1 and anaphase cells from an asynchronous culture. For the box plots in a–c, 200–250 kinetochore clusters were quantified, and for statistical analysis, a two-tailed t test was used. The middle line represents the mean, the box represents SD, and whiskers represent SEM. (b) The addition of the Ndc80 submodule component Nuf2 in anaphase is significantly reduced by deletion of FIN1 or FIN1 and CNN1, while the G1 level is unchanged. The normalized intensity of GFP-tagged Nuf2 protein was quantified in G1 and anaphase cells from an asynchronous culture (*, P < 0.0001). (c) The level of Dam1 submodule component Ask1 does not change between G1 and anaphase in WT cells. However, lower levels overall are observed when FIN1 or FIN1 and CNN1 are deleted (*, P < 0.0001). (d) The percent recovery of Nuf2-GFP in fin1Δ (n = 26 cells) shows significantly less recovery than in WT (n = 16 cells). For statistical analysis, a two-tailed t test was performed. In the plot, the middle line represents the mean, and whiskers represent SD; *, P, < 0.0001. (e) The percent recovery of Ask1-GFP is similarly poor in fin1Δ (n = 30) and WT cells (n = 10). Plotting and statistical analysis were performed as in d. (f) SPA-SIM of kinetochore clusters in metaphase (top, SPB distance <2 µm) and anaphase (bottom, SPB distance > 2 µm) aligned to Spc42 in WT and fin1Δ backgrounds. All images are averages of >24 individual images. Bar, 200 nm.
Figure 3.

Assembly of microtubule binding subcomplexes Dam1 and Ndc80 is affected by loss of Fin1. (a) The MIND submodule component Dsn1 is not significantly affected by deletion of FIN1 or FIN1 and CNN1. The normalized intensity of GFP-tagged Dsn1 protein was quantified in G1 and anaphase cells from an asynchronous culture. For the box plots in a–c, 200–250 kinetochore clusters were quantified, and for statistical analysis, a two-tailed t test was used. The middle line represents the mean, the box represents SD, and whiskers represent SEM. (b) The addition of the Ndc80 submodule component Nuf2 in anaphase is significantly reduced by deletion of FIN1 or FIN1 and CNN1, while the G1 level is unchanged. The normalized intensity of GFP-tagged Nuf2 protein was quantified in G1 and anaphase cells from an asynchronous culture (*, P < 0.0001). (c) The level of Dam1 submodule component Ask1 does not change between G1 and anaphase in WT cells. However, lower levels overall are observed when FIN1 or FIN1 and CNN1 are deleted (*, P < 0.0001). (d) The percent recovery of Nuf2-GFP in fin1Δ (n = 26 cells) shows significantly less recovery than in WT (n = 16 cells). For statistical analysis, a two-tailed t test was performed. In the plot, the middle line represents the mean, and whiskers represent SD; *, P, < 0.0001. (e) The percent recovery of Ask1-GFP is similarly poor in fin1Δ (n = 30) and WT cells (n = 10). Plotting and statistical analysis were performed as in d. (f) SPA-SIM of kinetochore clusters in metaphase (top, SPB distance <2 µm) and anaphase (bottom, SPB distance > 2 µm) aligned to Spc42 in WT and fin1Δ backgrounds. All images are averages of >24 individual images. Bar, 200 nm.

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