Suppression of PGAL1-NDJ1 lethality in vegetative cells. (A and B) Ndj1 colocalization with Mps3 in vegetative yeast cells. Yeast strains PGAL1-GFP-NDJ1 MPS3-RFP (HY4179) and PGAL1-GFP-NDJ1 RAP1-RFP (HY4217) were grown in dextrose medium, then transferred to galactose medium 2 h before microscopy. Arrows point to the Ndj1 (green) focus that overlaps with Mps3-RFP (red) but not with Rap1-RFP (red). Quantification of Ndj1-GFP focus formation is shown in B. Bar, 2 µm. (C) Localization of Ndj1 to SPB depends on the N terminus of Mps3 in vegetative cells. Yeast strains 3HA-MPS3(Δ1-64) TUB4-RFP (HY4149) and 3HA-MPS3(Δ1-64) PGAL1-GFP-NDJ1 TUB4-RFP (HY4150) were prepared for microscopy as in A. Cells from the above strains were mixed before microscopy. The two cells shown were from the same microscopy field. Note that in the absence of the N terminus of Mps3, GFP-Ndj1 (green) no longer formed a focus at the SPB marked by Tub4-RFP (red). (D and E) Ectopic expression of NDJ1 is lethal in vegetative cells. Yeast cells (HY3799, HY4128, HY4149, HY4150, HY4917, and HY4933) were grown in dextrose medium, serially diluted, and spotted on dextrose and galactose plates. Note that both removal of the N terminus of Mps3 and pom152Δ suppressed the lethality caused by ectopic expression of Ndj1. (F) Quantification of Ndj1 localization to the SPB in selected yeast strains. The SPB was marked by Tub4-RFP. Strains HY4128, HY4150, and HY4947 were used.
