FIG. 3.

Comparison of the steady-state levels of CYC1, ACT1, and CYH2 precursor and matured mRNAs at 25°C (mock-shifted) and 37°C (shifted) in NUP116 (normal), nup116-Δ, nup116-Δ cbc1-Δ, nup116-Δ rrp6-Δ, nup116-Δ cbc1-Δ rrp6-Δ, nup116-Δ rai1-Δ, and nup116-Δ upf1-Δ deletion strains. (A) Northern blots of steady-state levels of CYC1, ACT1, and CYH2 mRNAs and pre-mRNA in different strains are indicated at the top of each lane. All the strains were grown at 25°C until mid-log phase, half of the culture of each strain was then shifted to 37°C. Both cultures of each strain at 25 and 37°C were incubated for 1 h at the respective temperature and harvested. Subsequently, the steady-state levels of the each mRNA and pre-mRNA were determined by Northern blot analysis with the total RNA isolated from each of these strains and probing for respective pre-mRNA and mRNAs isolated from strains grown at 25°C (lanes 1 to 7) and shifted to 37°C (lanes 8 to 14). The signal for each mRNA in each lane was quantified as described in Materials and Methods and normalized against the 18S rRNA signals (shown at the lowest panel of A) for loading errors and the relative steady-state levels of each mRNA are presented in Table 2. (B) Quantification of the CYH2 pre-mRNA and mRNA signals. The intensity of each band of mature and precursor mRNA were determined by scanning the blots with a PhosphorImager and by normalizing for loading differences with respect to 18S rRNA signals. The relative levels of pre-mRNA and mRNA in each strain were expressed with respect to that in the NUP116 (normal) strain at each temperature, which was considered to be 100%. The error bar represents the range of three independent experiments.