Nucleic Acids Research, Vol 25, Issue 21 4201-4208, Copyright © 1997 by Oxford University Press
J Klovins, J van Duin and RCL Olsthoorn
The secondary structure of the RNA from the single-stranded RNA
bacteriophages, like MS2 and Qb, has evolved to serve a variety of
functions such as controlling gene expression, exposing binding sites for
the replicase and capsid proteins, allowing strand separation and so forth.
On the other hand, all of these foldings have to perform in bacterial cells
in which various RNA splitting enzymes are present. We therefore examined
whether phage RNA structure is under selective pressure by host RNases.
Here we show this to be true for RNase III. A fully double-stranded hairpin
of 17 bp, which is an RNase III target, was inserted into a non-coding
region of the MS2 RNA genome. In an RNase III-host these phages survived
but in wild-type bacteria they did not. Here the stem underwent Darwinian
evolution to a structure that was no longer a substrate for RNase III. This
was achieved in three different ways: (i) the perfect stem was maintained
but shortened by removing all or most of the insert; (ii) the stem acquired
suppressor mutations that replaced Watson-Crick base pairs by mismatches;
(iii) the stem acquired small deletions or insertions that created bulges.
These insertions consist of short stretches of non-templated A or U
residues. Their origin is ascribed to polyadenylation at the site of the
RNase III cut (in the + or - strand) either by Escherichia coli poly(A)
polymerase or by idling MS2 replicase.
ARTICLES
Rescue of the RNA phage genome from RNase III cleavage
Department of Biochemistry, Leiden Institute of Chemistry, Leiden University, 2300 RA Leiden, The Netherlands.
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