The start sites of sequences of 18S and 25S rRNA found to be exonuclease resistant as indicated by oligo attachment

Abstract

 

Background

We have previously reported 18S and 25S ribosomal RNA molecules in Candida albicans resistant to processive 5´→3´ exonuclease, appearing as cells approached stationary growth phase. Initial analysis pointed to extra phosphate(s) at their 5’- end raising the possibility that they were newly transcribed. Here we report on additional experiments exploring this possibility and try to establish which of the RNA polymerases may be transcribing them.

Results

Oligo-ligation and primer extension again showed the presence of extra phosphate at 5’-end at or near the reported processing sites for both 18S and 25S ribosomal RNA components. Inhibition of Pol I with BMH-21 increased the presence of the molecules.  Quantitation with an Agilent Bioanalyzer showed that resistant 18S and 25S molecules are primarily produced in the nucleus.  Utilizing an RNA cap specific antibody, a signal could be detected on these molecules via immunoblotting; such signal could be eli...