Figure 7. Characterization of NDC80 and HSK3 overexpression.
(A) CUP-NDC80-3V5 (A30342) and CUP-HSK3-3HA (A32060) cells also carrying the GAL4-ER and GAL-NDT80 fusions were induced to sporulate. After 2 hr 30 min CuSO4 (50 μM) was added and cells were subsequently released from NDT80 block 4 hr 30 min after transfer into sporulation medium. The levels of Ndc80-3V5, Hsk3-3HA and Pgk1 were monitored by Western blot. (B) CUP-NDC80-3V5 CUP-CLB3 (A31949), CUP-NDC80-3V5 CUP-HSK3 (A31951) and CUP-NDC80-3V5 CUP-HSK3 CUP-CLB3 (A31953) cells were induced to sporulate. 4 hr after transfer into sporulation medium CuSO4 (50 μM) was added, and localization of Ndc80-3V5 (green) relative to Ndc10-6HA (red) was determined by nuclear spreads 5 hr after transfer into sporulation medium.