Cic1p/Nsa3p inactivation impairs pre-rRNA processing. (A) Structure and processing sites of the 35S pre-rRNA. This precursor contains the sequences for the mature 18S, 5.8S, and 25S, which are separated by the two internal transcribed spacers ITS1 and ITS2 and flanked by the two external transcribed spacers 5′ETS and 3′ETS. The positions of the oligonucleotides probes utilized in Northern hybridizations and primer extension analysis are indicated. (B, C) Northern analysis of pre-rRNA processing. Strains cic1–2 and wild type were growth at 25°C in YPD then shifted to 37°C. Cells were harvested at the indicated times and total RNAs were extracted. Equal amounts of RNA (5 μg) were resolved on 1.2% agarose/formaldehyde gel (B) and 6% acrylamide/urea gel (C) and transferred to a nylon membrane. The membranes were consecutively hybridized with the probes indicated in panel A. The position of the mature rRNAs and pre-rRNAs are indicated. (D) Primer extension analysis of the level of the 27SA2 and 27SB pre-rRNAs, which were detected using primer 006.
