Pds1 depletion causes immediate spindle elongation in a phospho-mimetic Esp1 mutant.
(A) Pds1 depletion causes synthetic sickness in ESP1-3D cells. Eight-fold serial dilutions of the indicated strains were spotted onto the indicated plates and grown at 25°C. (B) The ESP1-3D allele is semi-dominant in diploids. Ten-fold serial dilutions of the indicated diploids were grown in YEP + dextrose media, spotted onto the indicated plates and grown at 25°C. (C) The ESP1-3D allele is semi-dominant when complemented by a plasmid borne ESP1 (ESP1-CEN-HIS3). Ten-fold serial dilutions of the indicated strains were grown in media lacking histidine, spotted onto the indicated plates and grown at 25°C. (D) Mitotic spindle morphology of individual ESP1-3D cells depleted of Pds1. PDS1-AID SPC42-eGFP and ESP1-3D PDS1-AID SPC42-eGFP cells were grown at 25°C to log phase and arrested in G1 with α-factor. 30 min before α-factor release +/- auxin was added. Cells were released at t = 0 and at t = 25 min cells were plated onto YPD live microscopy pads +/- auxin and Spc42-eGFP was imaged every minute. Each strain was imaged at least two times in each condition. Cells that undergo normal metaphase spindle formation are shown in black. Cells that undergo immediate spindle elongation upon spindle formation are shown in green. Cells that exhibit failed or no anaphase spindle elongation are shown in red. See S3 Fig for cell traces of experiments done in the absence of auxin and Fig 3 for tabulation of all imaging data. The scoring metric is described in the text and in the material and methods.