Figure 2.

SPLIFF analysis of the Net1p/Sir2p interaction. (A) Cartoon of the RENT complex. (B) Interaction between Net1p and Sir2p as measured by the Split-Ub growth assay. Cells expressing an interacting N_ub fusion grow on medium containing 5-FOA. (C) Interaction between Net1p and Sir2p as measured by SPLIFF. Selected frames of the time-lapse analysis of the mating of two yeast cells expressing Net1CCG and N_ub-Sir2, respectively, (white border) are shown. Nuclei of unfarmed cells belong to haploid a-cells. Note the selective loss of green fluorescence from the a-cell-originated nucleus in the diploid at 10 and 20 min. Time 0 indicates the time point shortly before cell fusion. Scale bar, 5 μm. (D) Quantitative analysis of the experiment shown in (C) (upper panel) or the averages of 6 independent experiments (lower panel). The relative fluorescence of GFP (), Cherry () and the calculated conversion of Net1CCG to Net1CC () are plotted against the time (error bars, standard error (s.e.)). See also Supplementary Figure 1 and Supplementary Movie 1. Source data for this figure is available on the online supplementary information page.

Source data for Figure 2D ^{(25.5KB, xls)}