Tor1 and CK2 kinases control a switch between alternative ribosome biogenesis pathways in a growth-dependent manner
Fig 2
In vivo pulse-chase and proteomic analyses of the pre-rRNA processing in exponential and postdiauxic shift cultures.
(A) Low density (OD600 = 2) YMK118 was pulse labeled in vivo with 3H-uracil for 6 min and then chased with nonradioactive uracil for 10 min. (B) Yeast culture in early postdiauxic phase (OD600 = 10) was pulse labeled for 6 min. The culture was split, and one-half was chased for 10 min, as in (A) (on the left). To the other half of the culture, fresh media and no-radioactive uracil was added to the same final concentration and chased for 10 min (right side, “exponential chase”). (C) Comparison of intensity based absolute quantification (iBAQ) values normalized to bait of 324 proteins detected in the affinity purified preribosomes from pre- and postdiauxic cultures. Proteins changed more than 2-fold are in black. (D) Histogram of SILAC Heavy/Light (H/L) ratios (normalized to bait) for proteins with more than 2-fold change in the preribosomes purified from postdiauxic shift versus prediauxic shift cultures (for high resolution see S7 Fig). An experiment with a higher coverage is shown. Underlying numerical data are in S1 Data. (E) Northern blot analysis of RNA isolated form wild-type yeast or strains deleted for different exosome/TRAMP complex factors or Xrn1. All strains were grown until their growth stopped after the diauxic shift. The xrn1Δ strain stopped growing at the indicated lower OD600 than the other strains.