Figure 3. Ypt1 knockdown effect on HAC1 RNA transcription and decay.

(A) Knocking down YPT1 does not affect HAC1 promoter activity as evaluated by assaying RNA levels of a transcriptional reporter gene containing the HAC1 promoter sequence fused to a GFP ORF sequence and the ACT1 3′UTR. Normalized expression, reported as a fraction of actin RNA levels, was determined using quantitative RT-PCR. Data represent the average of three replicate experiments. (B-C) Quantitative RT-PCR measurements of unspliced (B) and spliced (C) HAC1 RNA expression 30 min post addition of 3 ug/ml thiolutin (“Treated”) relative to HAC1 RNA levels in untreated cells. Values are normalized to GAPDH levels and data are averages of 3–4 replicates. Asterisks indicate significant differences. * = p<0.05 by t-test.