Figure 5.

Spc7 is required for the integrity of the spindle. (A) Photomicrographs of spindle defects observed in spc7-23 cells incubated at 36°C for 6 h: thin spindle midzones (a and b), two half-spindles per cell (c and d), bent spindle (e), and unequally stained spindle (f). Fixed cells were stained with anti-tubulin antibody. (B) Serial dilution patch tests (10⁴ to 10¹ cells) of wild-type, spc7-30, and spc7-24 cells carrying the integrated nmt81-GFP-atb2. Strains were grown at the indicated temperatures for 5 d under derepressed conditions. (C) Chromosome segregation and spindle phenotypes observed in spc7-23 and spc7-23 nmt81-GFP-atb2 (spc7-23 gfp-atb2) strains grown at 34°C. Right diagram (from left to right), wild-type anaphase spindle, anaphase spindles with thinly staining midzone, disintegrating/broken anaphase spindles, bent spindles and unequally stained spindles. N/strain = 100. (D) Immunoprecipitations of Spc7-HA and GFP-Atb2 proteins. Protein extracts prepared from strains expressing Spc7-HA, GFP-Atb2, or both were used for immunoprecipitations (IP) with an anti-GFP or anti-HA antibody, followed by Western blot analysis using anti-GFP and anti-HA antibodies.