Supplemental Material

This article contains the following supporting material:

• Supplemental Figure 1 - Yeast depleted of Pwp2 accumulate unbudded cells and pre-rRNAs prior to an increase in 25S/18S rRNA levels. GAL::PWP2 tagged yeast and the untagged parental strain (W303) were grown to early log phase in YPG media and shifted into YPD media for up to 24 hours. Cell growth and the percentage of unbudded cells were monitored after 0, 3, 6, 9, 12, and 24 hours of growth in dextrose. Yeast depleted (GAL::PWP2) or not (W303) of Pwp2 were grown to early log phase in YPG and shifted into YPD media for up to 24 hours. RNA was collected at either 0, 4, 8, 12, 16, 20, or 24 hours in dextrose. Twenty μg of RNA was run on a 1.25% formaldehyde agarose gel, transferred to Hybond N+ membrane, and probed by Northern blot for pre-rRNAs and the mature 25S and 18S rRNAs. Ratio of the 25S/18S, 20S/25S, and 35S/25S ratios were analyzed and plotted by time in YPD (Hours DEX) for GAL::PWP2 and W303. GAL::PWP2 and W303 yeast were grown in YPG and shifted into YPD. Cells were labeled with ³⁵S translabel and protein was TCA precipitated. New protein synthesis rates are also plotted by time in YPD (Hour DEX). Five μl of TCA precipitated protein was run on 10% SDS-PAGE gel and coomassie stained and exposed to film.
• Supplemental Table 1 - Summary of Elutriation Results. All yeast strains were compared at a reference point of 50% budded for cell size (fl), protein content (arbitrary units), and RNA content (arbitrary units). Due to variation between elutriations, comparison between samples can only be made between the paired strains that were analyzed at the same time under the same conditions (YPH499 and GAL::PWP2; whi5· and GAL-PWP2 whi5·).