1A6/DRIM, a Novel t-UTP, Activates RNA Polymerase I Transcription and Promotes Cell Proliferation
Figure 1
Knockdown of 1A6/DRIM results in a decrease in 47S rRNA.
A. Cartoon illustrating the location of the probe on the 47S rRNA used for Northern blotting to detect 47S rRNA transcript. B. A 1A6/DRIM specific siRNA (si2-8) was transfected into U2OS cells. RNA was extracted 72 hours posttransfection, resolved on a 1% glyoxal-agarose gel and transferred onto a positively charged nylon membrane. Blots were probed with a Biotin-labeled DNA oligonucleotide (as shown in A) to detect 47S rRNA (left upper panel). Ethidium Bromide (EB) staining of the 28S rRNA on the agarose gel was used as a loading control. Cell lysates were prepared 72 hours after siRNA transfection and proteins from the lysates were separated on SDS-PAGE and transferred onto a PVDF membrane. Blots were probed with anti-1A6/DRIM antibody. Topoisomerase I (Topo I) was used as a loading control (left lower). The experiment was repeated three times and the difference in the densitometry scanning of the 47S rRNA bands is displayed in the right panel. C. A 1A6/DRIM specific siRNA (si2-8) was transfected into U2OS cells. RNA was extracted 72 hours posttransfection and reverse transcription was carried out. Real time PCR was performed with primers amplifying the 5′-extreme end fragment of the 47S rRNA. The experiment was repeated three times in duplicate and the statistical significance of the difference is shown. Statistical analyses were performed with two-tailed unpaired t test.