Stable tRNA precursors in HeLa cells

Abstract

32 Two tRNA precursors were isolated from P-labeled or unlabeled HeLa cells by two dimensional polyacrylamide gel i esctrophoresis and were sequenced. These were the precursors of tRNA e and tRNA e , and both contained four extra nucleotides including 5'-triphosphates at their 5'-end and nine extra nucleotides including oligo U at their 3'-end. These RNAs are the first naturally occurring tRNA precursors from higher eukaryotes whose sequences have I en etermined4 In these molecules, several modified nucleosides such as m G, t A and ac C in mature tRNAs were undermodified. Two additional hydrogen bonds were formed in the clover leaf structures of these tRNA precursors. These extra hydrogen bonds may be responsible for the stabilities of these tRNA precursors.

Precursors of tRNAs are stabilized by methylguanosine cap structures

“…We detected a methylguanosine cap in Pre 1 but not in Pre 3 of tRNA Leu CAA . Thus, our observations are consistent with the previous finding 31 . The lack of capping of Pre 3 in mammals suggests a possible regulatory mechanism in tRNA biogenesis-for example, 5′ capping of Pre 1 might prevent tRNA splicing for generation of Pre 3 or enhance degradation of the capped Pre 1.…”

“…1 and Supplementary Tables 1 and 2a), indicating that 3′ end maturation precedes 5′ processing when RNase P is not functional. Although the 5′ terminus of pre-tRNA is thought to bear a triphosphate 26,[30][31][32][33][34] , we detected few fragments with 5′-triphosphate (pppAACp) structures in either pre-tRNA (Fig. 2b).…”

“…Several groups have described the 5′ termini of pre-tRNAs in eukaryotes 26,[30][31][32][33][34] . Pre-tRNAs transcribed from an expression vector in cell extracts or in Xenopus oocytes have 5′ triphosphate or diphosphate at their 5′ termini 34 .…”

“…Pre-tRNAs transcribed from an expression vector in cell extracts or in Xenopus oocytes have 5′ triphosphate or diphosphate at their 5′ termini 34 . Naturally occurring pre-tRNAs, including pre-tRNA Leu HeLa cells 31 . RNA fingerprinting analyses revealed that the 5′ termini of tRNA Leu CAA precursors bear triphosphate.…”

APhaseolus vulgaris mitochondrial tRNALeu is identical to its cytoplasmic counterpart: sequencing andin vivo transcription of the gene corresponding to the cytoplasmic tRNALeu

“…There is an unknown modification (N) found at position 34 ("wobble" position of the anticodon) in both mt tRNA Leu and cyt tRNA Leu species. Two other reports [22,23] have revealed the presence of an unidentified modification at position 34 in tRNA Leu (NAA) of HeLa cells and of the rat turnout Morris hepatoma which has been identified as a derivative of C. In our case N is also a derivative of C (see below) but it is not possible to determine whether the modification we report here is in fact the same as that found in HeLa cells and rat tumour Morris hepatoma.…”