FIG 6.

asp1^D333A cells require Mph1 for the prolonged delay into anaphase. (A) Serial dilution patch tests (10⁴ to 10¹ cells) of the viability of the indicated strains on YE5S plates with (+TBZ) or without (−TBZ) 4 μg/ml TBZ. The plates were incubated for 4 days at 25°C. (B) Confocal live-cell images of the indicated strains expressing SV40::GFP-Atb2⁺. Time between images, 5 min. Bars, 2 μm. (C) Diagrammatic illustration of mitotic spindle length over time for the cells shown in panel B. (D) Quantification of time required for transition from prometaphase to anaphase B due to spindle elongation of the indicated strains (asp1⁺ strain, n = 16; asp1^D333A strain, n = 16; asp1^D333A mph1Δ strain, n = 13; ***, P < 0.0005 for the asp1^D333A mph1Δ [two-sample t test] and asp1^D333A [Welch test] strains compared to the asp1⁺ strain and for the asp1^D333A mph1Δ [Welch test] strain compared to the asp1^D333A strain). (E) Phenotypes of aberrant cen1-GFP segregation behavior, i.e., lagging cen1-GFP signal that moved to the correct SPB during the time of measurement (light gray) or cen1-GFP signals both present at one SPB or a cen1-GFP signal that did not move to either SPB (dark gray) (asp1⁺, n = 62; mph1Δ, n = 46; asp1^D333A, n = 34; asp1^D333A mph1Δ, n = 36).