tRNA nuclear transport: Defining the critical regions of human tRNAimet by point mutagenesis

Tobian, Janet; Drinkard, L. C.; Zasloff, Michael

doi:10.1016/0092-8674(85)90171-0

Cited by 113 publications

(64 citation statements)

References 16 publications

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“…Nuclear export of tRNA requires that the tRNA adopt the correct tertiary structure (Tobian et al 1985). Therefore, to avoid disruption of the tertiary structure of the imported glutamine tRNA during mutational analysis, the sequences that differ between the imported and nonimported glutamine tRNAs were substituted as structural blocks (boxed in Fig.…”

Section: Design Of Mutant Trnasmentioning

confidence: 99%

The anticodon is the signal sequence for mitochondrial import of glutamine tRNA in Tetrahymena.

Rusconi¹,

Cech²

1996

Genes Dev.

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The import of nuclear-encoded RNAs into mitochondria is required for proper mitochondrial function in most organisms. However, the mechanisms used to achieve RNA import are largely unknown. In particular, the RNA elements that direct import have not been identified in any organism. In Tetrahymena, only one of three nuclear-encoded glutamine accepting tRNAs is imported into mitochondria. We transform Tetrahymena with marked glutamine tRNAs and quantitate their level of accumulation in mitochondria. Of several isostructural nucleotide substitutions tested, alteration of the anticodon sequence uniquely abolishes import. Furthermore, substitution of a single anticodon nucleotide (UUA-,UUG) confers import on a normally nonimported glutamine tRNA. Thus, the anticodon functions as a mitochondrial localization signal and is both necessary and sufficient for tRNA import. Given the prior evidence that neither the cytoplasmic nor the mitochondrial glutaminyl-tRNA synthetase distinguishes between the imported and nonimported glutamine tRNAs with respect to aminoacylation, we propose that some mitochondrial import factor distinct from a synthetase recognizes the anticodon of the imported glutamine tRNA.

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Section: Design Of Mutant Trnasmentioning

confidence: 99%

The anticodon is the signal sequence for mitochondrial import of glutamine tRNA in Tetrahymena.

Rusconi¹,

Cech²

1996

Genes Dev.

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“…mRNA export is facilitated by its 5' m7G cap (10, 11). Exit of tRNAs and ribosomal subunits appears to be receptor mediated (9,12).…”

Section: Introductionmentioning

confidence: 99%

“…mRNA export is facilitated by its 5' m7G cap (10, 11). Exit of tRNAs and ribosomal subunits appears to be receptor mediated (9,12).Because colloidal gold coated with poly(A), tRNA, or 5S RNA microinjected into the nucleus exits via nuclear pores (13) and because exit of tRNA and 5S RNA is inhibited by antibodies that react with pore complexes (14), it is likely that mRNA also exits via pores. Maximal mRNA release from isolated animal cell nuclei requires ATP (15-17).…”

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confidence: 99%

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A conditional yeast mutant deficient in mRNA transport from nucleus to cytoplasm.

Kadowaki

Zhao

Tartakoff³

1992

Proc. Natl. Acad. Sci. U.S.A.

103

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Transport of mRNA from nucleus to cytoplasm is critical for eukaryotic gene expression; however, the mechanism of export is unknown. Selection and screening procedures have therefore been used to obtain a family of temperature-sensitive conditional mutants of Saccharomyces cerevisiae that accumulate poly(A)+ RNA in the nucleus when incubated at 37C, as judged by in situ hybridization. In one such mRNA ransport mutant, mtrl-1, RNA synthesis continues, the export of poly(A)+ RNA is inhibited, intranuclear poly(A)+ is remarkably stable, and protein synthesis gradually stops. Thus, there is no tight coupling between RNA synthesis and export. The export lesion is reversible. Although mRNA export is clearly not a default option, neither inhibition of protein synthesis, inhibition of mRNA splicing, nor inhibition of poly(A)-binding protein function blocks export of the average poly(A)+, as judged by in situ hybridization. Further analysis of the family of mtr mutants should help map the path of RNA transport.Gene expression in eukaryotes requires the rapid and selective export of mRNA from the nucleus to the cytoplasm. Most pre-mRNAs acquire a 5' m7G cap and are cleaved and polyadenylylated to generate their 3' end. mRNAs have few other common structural features; however, specific proteins are associated with both extremities of intranuclear premRNAs (1, 2), and intranuclear mRNAs associate with a number of additional proteins (3,4) as well as the karyoskeleton (5).Export of 5S RNA and rRNAs requires association with specific proteins (6)(7)(8) and the export of tRNA is highly sensitive to base changes, possibly for this reason (9). mRNA export is facilitated by its 5' m7G cap (10, 11). Exit of tRNAs and ribosomal subunits appears to be receptor mediated (9,12).Because colloidal gold coated with poly(A), tRNA, or 5S RNA microinjected into the nucleus exits via nuclear pores (13) and because exit of tRNA and 5S RNA is inhibited by antibodies that react with pore complexes (14), it is likely that mRNA also exits via pores. Maximal mRNA release from isolated animal cell nuclei requires ATP (15-17).In Saccharomyces cerevisiae several features of processing of pre-mRNA are relatively simple: only the 5' extremity of mRNA is methylated (18), only few mRNAs undergo splicing (19,20), and the 3' poly(A) tail of yeast mRNA is somewhat shorter than in animal cells (1, 21). Although nuclear pores of yeast have been characterized (22)(23)(24), and although temperature sensitive (ts) splicing mutants (prp mutants) have been produced (19,20,(25)(26)(27), only a single ts mutant has been claimed to affect mRNA export (28). This mutant, rnal, has multiple defects in covalent processing of RNA. The corresponding gene product is cytoplasmic (29). (ii) Selection B. Cells were incubated 3 hr at 37°C in lysine-free SD medium supplemented with the lysine analogue S-2-aminoethyl-L-cysteine and the proline analogue L-azetidine-2-carboxylate. The incubation was continued at 37°C for 24 hr; then cells were inoculated to YEPD plates. The...

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“…tRNAs are transcribed by RNA polymerase III. Following transcription and maturation, tRNAs are rapidly transported from the nucleus to the cytoplasm (4,5).…”

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confidence: 99%

Unique pathway of expression of an opal suppressor phosphoserine tRNA.

Lee¹,

Peña²,

Tobian³

et al. 1987

Proc. Natl. Acad. Sci. U.S.A.

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An opal suppressor phosphoserine tRNA gene is present in single copy in the genomes of higher vertebrates. We have shown that the product of this gene functions as a suppressor in an in vitro assay, and we have proposed that it may donate a modified amino acid directly to protein in response to specific UGA codons. In this report, we show through in vitro and in vivo studies that the human and Xenopus opal suppressor phosphoserine tRNAs are synthesized by a pathway that is, to the best of our knowledge, unlike that of any known eukaryotic tRNA. The primary transcript of this gene does not contain a 5'-leader sequence; and, therefore, transcription of this suppressor is initiated at the first nucleotide within the coding sequence. The 5'-terminal triphosphate, present on the primary transcript, remains intact through 3'-terminal maturation and through subsequent transport of the tRNA to the cytoplasm. The unique biosynthetic pathway of this opal suppressor may underlie its distinctive role in eukaryotic cells.

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tRNA nuclear transport: Defining the critical regions of human tRNAimet by point mutagenesis

Cited by 113 publications

References 16 publications

The anticodon is the signal sequence for mitochondrial import of glutamine tRNA in Tetrahymena.

The anticodon is the signal sequence for mitochondrial import of glutamine tRNA in Tetrahymena.

A conditional yeast mutant deficient in mRNA transport from nucleus to cytoplasm.

Unique pathway of expression of an opal suppressor phosphoserine tRNA.

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