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. Author manuscript; available in PMC: 2010 Feb 10.
Published in final edited form as: Curr Biol. 2009 Jan 29;19(3):196–205. doi: 10.1016/j.cub.2008.12.047

Figure 6. Dominant negative effects of motor-3YFP can be rescued by overexpression of Pac1/LIS, but not Bik1/CLIP-170.

Figure 6

(A) The percentage of cells with a misoriented mitotic spindle in a cold (16°C) spindle position assay [24, 29] is plotted for diploid strains carrying DYN1/HC-2mCherry at one DYN1 locus, and either 3GFP, TAIL-3GFP, or MOTOR-3YFP (wild-type or AAA3 mutants) at the other DYN1 locus (n > 570 cells for each strain). The spindle was visualized using CFP-Tub1. A control diploid strain carrying no functional DYN1 is indicated on the right. (B) The percentage of cells that display Dyn1/HC-3mCherry foci at a given location is plotted for diploid strains similar to those in (A), except that the DYN1/HC-2mCherry allele was replaced with DYN1/HC-3mCherry for brighter fluorescence (n > 150 cells for each strain). Movies of Dyn1/HC-3mCherry and CFP-Tub1 were collected to distinguish foci at plus-ends from SPB and the cell cortex. Strains were imaged after growth under the same conditions as in (A). (C) The percentage of cells with a misoriented mitotic spindle in a cold spindle position assay is plotted for diploid strains carrying DYN1/HC-3mCherry and either 3GFP or MOTOR-3YFP, as indicated. The GAL1 promoter was inserted at the 5′ end of one of the chromosomal BIK1 or PAC1 loci. Strains were imaged after growth at 16°C to mid-log in synthetic defined media supplemented with 2% galactose (n > 250 cells for each strain). (D) Percentage of cells that display Dyn1/HC-3mCherry foci at a given location is plotted for the same strains used in (C), and imaged after growth under the same conditions as in (C) (n > 160 cells for each strain). (E) Kymograph depicting Dyn1/HC-3mCherry migrating along cytoplasmic microtubule toward a plus-end in a Pac1/LIS overexpressing cell. The motor-3YFP signal was used as reference point to identify the plus-end. Vertical bar represents 10 s; horizontal bar represents 1 μm. See Movie S10.