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. Author manuscript; available in PMC: 2014 Sep 18.
Published in final edited form as: Curr Biol. 2011 Nov 23;21(23):1968–1978. doi: 10.1016/j.cub.2011.10.051

Figure 2. Csc1p promotes asymmetric localization of SIN-kinases Cdc7p and Sid1p.

Figure 2

A. csc1Δ generates uninucleate cells with a division septum and binucleate cells with more than one septum. csc1Δ cells were grown at 30°C, fixed, and stained with DAPI (for DNA) and aniline blue (for cell wall). B. Loss of Csc1p function can cause inappropriate septation in interphase. Wild type (WT) and csc1Δ cells were grown at 24°C or 30°C, treated with 12 mM HU for 6 hrs to arrest the cells at S-phase. Interphase cells with a division septum were quantitated (n=500×3). Cells with septum deposition were counted for WT as well as csc1Δ. C–D. Cdc7p and Sid1p localize symmetrically in csc1Δ cells. Wild type cells expressing Cdc7p OR Sid1p-GFP and mCherry Atb2p were fixed with methanol and the localization of the fluorescent proteins investigated. The fraction of cells displaying Cdc7p or Sid1p in an asymmetric configuration or the fraction of csc1Δ cells displaying a symmetric distribution of Cdc7p or Sid1p are indicated in the figure. E. Simultaneous compromise of Csc1p and Cdc16p function leads to growth defects. csc1Δ, cdc16-116 and csc1Δ cdc16-116 cultures were serially diluted 5 fold for 4 times and spotted on YES-agar medium and grown at 24°C and 30°C. The two parental strains were capable of colony formation at 24°C and 30°C. The csc1Δ cdc16-116 double mutant was capable of colony formation at 24°C, but not at 30°C. F. Simultaneous compromise of Csc1p and Cdc16p generates cells with multiple septa. csc1Δ, cdc16-116 and csc1Δ cdc16-116 mutants were grown at 24°C till early log phase, shifted to 30°C for 3 hours, fixed and stained with DAPI and aniline blue. G. Graphical representation of the percentage of hyperseptated cells in strains described F (n=500×3). H. Rescue of cdc16-116 growth defect with Csc1p overexpression (v-pREP41-GST and csc1-pREP41-csc1-GST). I. Csc1p-GFP cells do not display synthetic lethality with cdc16-116. csc1-GFP, cdc16-116 and csc1-GFP cdc16-116 cultures were serially diluted 5 fold and spotted on YES-agar plates and incubated at 24°C and 30°C.