Influence of αβ-tubulin on γ-tubulin binding to the SPB. (A) Localization of β-tubulin at SPBs in TUB2 wild-type and tub2-403 cells with TUB4-yeGFP. Both cell types were synchronized with α-factor, subsequently incubated with nocodazole, fixed, and stained with anti-Tub2 antibodies. (top) Images of fixed cells. Signals of anti-Tub2, Tub4-GFP, and stained DNA (blue) are shown. (bottom) Quantification of absolute intensities of anti-Tub2 at the SPB in tub2-403 and TUB2 cells. n, number of analyzed SPBs; fl. int., fluorescence intensity; a.u., arbitrary unit. (B) FRAP experiments of Tub4-yeGFP of TUB2 and tub2-403 cells treated with nocodazole for 1 h after synchronization with α-factor. (top) Relative fluorescence intensity (rel. fl. int.) of nonbleached cells. (middle) Mean FRAP recovery curve with SD error bars. (bottom) Images of photobleached cells and mean of t1/2 and Ymax ± SEM are shown. The t test (**, P = 0.0024) is for the t1/2 of the experiment. Bars: (A and B) 5 µm; (B, magnified SPB images) 2.7 µm2.
