Cytoplasmic tagging of the 3’ end of mRNA with non-canonical U and C nucleotides has emerged as a novel mechanism that appears to play an important role in determining the fate of mRNA. In Aspergillus nidulans, expression of genes involved in utilisation of poor nitrogen sources are regulated at different levels, including at the level of regulated transcript stability. Here changes occurring on the 3’ ends of genes involved in nitrate utilization, niaD and niiA, were analysed by Sanger sequencing of clones obtained after the 5’ adenylated (5rApp-) adaptor was ligated to the 3’ end of transcripts followed by RT, PCR and cloning into the TA vector system. It was found that partially degraded 3’ ends of the niiA and niaD transcripts contained sequences of adenosine nucleotides, suggesting that readenylation may be required for 3’-5’ mRNA decay. Furthermore, the addition of poly/oligo (A) tails to the 3’ end of mRNA was found to be diminished in strains depleted for either cutA or cutB and abolished in the absence of both enzymes. These findings strongly suggested that the two nucleotidyltransferases, CutA and CutB, which are responsible for the cytoplasmic C/U tagging in A. nidulans, may have an additional role in mRNA turnover. These findings argued that readenylation of eukaryotic transcripts represents a mechanism which resembles both the degradation of ρ-independent mRNAs in prokaryotes and re-uridylation dependent 3’-5’ degradation of the human cell-cycle regulated histone transcripts.
|Date of Award
|Igor Morozov (Supervisor)