Investigating the prevalence of queuine in Escherichia coli RNA via incorporation of the tritium-labeled precursor, preQ1

There are over 100 modified bases that occur in RNA with the majority found in transfer RNA. It has been widely believed that the queuine modification is limited to four transfer RNA species in vivo. However, given the vast amount of the human genome (60–70%) that is transcribed into non-coding RNA (Mattick [10]), probing the presence of modified bases in these RNAs is of fundamental importance. The mechanism of incorporation of queuine, via transglycosylation, makes this uniquely poised to probe base modification in RNA. Results of incubations of Escherichia coli cell cultures with [3H] preQ1 (a queuine precursor in eubacteria) clearly demonstrate preQ1 incorporation into a number of RNA species of various sizes larger than transfer RNA. Specifically, significant levels of preQ1 incorporation into ribosomal RNA are observed. The modification of other large RNAs was also observed. These results confirm that non-coding RNAs contain modified bases and lead to the supposition that these modifications are necessary to control non-coding RNA structure and function as has been shown for transfer RNA.

► Escherichia coli cells deficient in preQ1 biosynthesis can utilize exogenous preQ1 from growth media.
► Exogenous radiolabeled preQ1 is incorporated into cellular RNA in both wild-type and preQ1 deficient cells.
► Radiolabeled preQ1 is incorporated into RNAs of differing sizes larger than tRNA.
► Analysis of isolated ribosomes shows that radiolabeled preQ1 is incorporated into ribosomal RNA.
► The conclusion of this study is that queuine is incorporated into many other RNAs in addition to the known tRNAs.