Figure 5.

Sen1 High Copy Expression Suppresses Termination Defects in APT Mutants

(A) Serial 5-fold dilutions of wild-type, ref2Δ, or pta1-1 cells transformed with vector, pGSen1Myc, or two catalytically inactive point mutants of Sen1 in the Walker A and B motifs (pGSM-K1363A and pGSM-D1590A).

(B) Immunoblot analysis of whole-cell extracts from cells induced with 2% galactose to express the indicated constructs. Because of different growth rates, the induction time was varied for the wild-type-ref2Δ (6 hr) and the wild-type-pta1-1 (13 hr) pair.

(C) RNA blot analysis of SNR33 in REF2 or ref2Δ cells. 20 μg RNA was separated on a 1% agarose gel and normalized to the SNR33::YCR015c/SNR33 signal in the ref2Δ vector samples.

(D) RNA blot analysis of SNR33 in PTA1 or pta1-1 cells, performed as in (C).