Nutrient depletion and TOR inhibition induce 18S and 25S ribosomal RNAs resistant to a 5′-phosphate-dependent exonuclease in Candida albicans and other yeasts

Background: Messenger RNA (mRNA) represents a small percentage of RNAs in a cell, with ribosomal RNA (rRNA) making up the bulk of it. To isolate mRNA from eukaryotes, typically poly-A selection is carried out. Recently, a 5'-phosphate-dependent, 5' -> 3' processive exonuclease called Terminator has become available. It will digest only RNA that has a 5'-monophosphate end and therefore it is very useful to eliminate most of rRNAs in cell. Results: We have found that in the pathogenic yeast Candida albicans, while 18S and 25S components isolated from yeast in robust growth phase are easily eliminated by Terminator, those isolated from cells in the nutritionally diminished stationary phase, become resistant to digestion by this enzyme. Additional digestions with alkaline phosphatase, tobacco pyrophosphatase combined with Terminator point toward the 5'-prime end of 18S and 25S as the source of this resistance. Inhibition of TOR by rapamycin also induces resistance by these molecules. We also find that these molecules are incorporated into the ribosome and are not just produced incidentally. Finally, we show that three other yeasts show the same behavior. Conclusions: Digestion of RNA by Terminator has revealed 18S and 25S rRNA molecules different from the accepted processed ones seen in ribosome generation. The reason for these molecules and the underlying mechanism for their formation is unknown. The preservation of this behavior across these yeasts suggests a useful biological role for it, worthy of further inquiry.