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. Author manuscript; available in PMC: 2018 Sep 12.
Published in final edited form as: Nat Cell Biol. 2018 Mar 12;20(4):432–442. doi: 10.1038/s41556-018-0056-9

Figure 7. Modification of nucleoporins by Hos3 regulates Whi5 transport and CLN2 tethering.

Figure 7

(A) Acetylation of Nup60 in wild type cells released from a metaphase arrest. GAL1pr:CDC20 cells were arrested in glucose media for 3 h. Nup60 protein levels were assessed at the indicated times after Cdc20 induction (galactose addition) with anti-myc antibodies and acetylation levels were assayed with an antibody against acetylated lysines. (B) Acetylation of nucleoporins in Hos3 mutants. Endogenously tagged proteins were immunoprecipitated from HOS3-NLS and hos3Δ extracts and assayed as in panel (A). Experiments in A-B were repeated two times with similar results. (C) Distribution of Whi5 and its transport receptors in wild-type and acetyl-mimic (KN) mutants of the indicated nucleoporins. Nuclear fluorescence was measured in mother and daughter cells at the time of birth as in Fig. 5C. Boxes include 50% of data points, the line represents the median and whiskers extend to maximum and minimum values. The number of cells (n values) analyzed is in brackets; p < 0.05 (*), 0.01 (**) and 0.001 (***), two-sided Mann-Whitney test relative to WT. Exact p values: (Whi5) nup60 = 2.01x10-2, nup49 = 1.09x10-4, nup60 nup49 = 1.22 x10-5; hos3EN = 3.49 x10-10; (Kap95) nup60 nup49 = 1.48 x10-4; nup60 nup49 nup53 = 1.76 x10-5, nup60 nup49 nup57 = 1.58 x10-2; (Msn5) nup60 nup49 nup57 = 2.79 x10-5. (D) (Top) Subnuclear position of CLN2 (as in Fig. 6D) during the first 24 minutes following cytokinesis, determined by time-lapse imaging in daughter (D) and mother cells (M) of the indicated strains. Images were acquired at 12 min intervals (the number of cells analyzed is in brackets; cells were pooled from three independent experiments). (Bottom) GFP, mCherry and phase contrast composite images of CLN2::lacO LacI-GFP NUP49-mCherry mother (M) and daughter (D) cells. Images were acquired at 12 min intervals. Time is indicated in minutes; time 0 corresponds to cytokinesis. Maximal projections are shown. Position of CLN2 relative to the nuclear periphery (zone I or II) was determined in single confocal planes and is indicated in frames after cytokinesis.