Fig 1. The fission yeast SCYL homologue Ppk32 regulates TOR signalling.

(A) Tor1 immuno-precipitations. Soluble proteins were extracted from exponentially grown strains: ppk32.PK strain (upper) and wild-type, ppk32.Δ and tor1 Δ (lower). Tor1 immuno-precipites and Ppk32 co-immuno-precipites were assessed from the same gels. The relative levels of specific Ppk32 immuno-precipitation are shown. (B) Live cell imaging: GFP and transmitted light images of GFP-Ppk32 and wild type control strains. (1) shows a projection of 4 single planes covering 0.5 nm (2) shows a single plane image, → indicates cytoplasmic foci. For wild type cells full Z-stack projection is shown (C, D) Growth assay. Exponentially YES-grown cells of indicated strains were spotted in 10-fold serial dilution onto rich medium plates (YES) or YES supplemented with Torin1, BFA or vehicle (DMSO) alone. (E,F) Early exponential cells grown in indicated media. Samples from un-perturbed cultures were subjected to Western blot analysis to monitor Ppk32 levels (E), * indicates a background band, or Maf1.PK (upper part) or Ser546 Gad8 phosphorylation (Lower part) (F). Ponceau S staining was used as a loading control.