In order to understand why the first
tRNA (
tRNAGln) in the T4
tRNA gene cluster is not produced when T4 infects an
RNase III- mutant of Escherichia coli,
RNA metabolism was analyzed in
RNase III-
RNase P- (rnc, rnp) cells infected with bacteriophage T4. After such an
infection a new dimeric
precursor RNA molecule of
tRNAGln and
tRNALeu has been identified and analyzed. This molecule is structurally very similar to K band
RNA that accumulates in rnc+ rnp strains. It is four
nucleotides shorter than K
RNA at the 5' end. This molecule like K
RNA contains two
RNase P processing sites at the 5' ends of each
tRNA. Both sites are accessible to
RNase P. However, while in the K
RNA the site at the 5' end of
tRNALeu (the site in the middle of the substrate) is more efficiently cleaved than the other site, this differential is even increased in the Ks (K like) molecule. This difference is sufficiently large that in vivo in the
RNase III- strain the smaller precursor of
tRNAGln is degraded rather than being matured to
tRNAGln by
RNase P. This information contributes to the elucidation of the key role of
RNase III in the processing of T4
tRNA. It shows the dependence of
RNase P activity at the 5' end of
tRNAGln on a correct and specific cleavage by
RNase III at a position six
nucleotides proximal to the
RNase P site, and it explains why in the absence of
RNase III the first
tRNA in the T4
tRNA cluster,
tRNAGln, does not accumulate.