Figure 5.

Rat1 is necessary for normal ACT1 poly(A) site selection. (A) ACT1 poly(A) site choice is shifted in favor of more 3′ sites when Rat1 is inactivated. Total RNA from wild-type (WT), rat1-1, or pcf11-2 strains (DBY548, DBY745, and DBY593) grown at 25°C or 37°C for 90 min was analyzed by RPA with 3′ ACT1 and 5S rRNA probes. (Un) Uncleaved RNA. The diagram shows poly(A) cleavage sites 1–5, the ACT1 anti-sense riboprobe, and RNase protection products. (B) Quantification of ACT1 mRNA cleavage at sites 1–5 in wild-type (WT), rat1-1, and pcf11-2 cells at 37°C. Cleavage relative to site 1 is shown after normalization for the ³²P-U content of the protection products 1–5. Values are means and standard deviations from two experiments.