Another heritage from the RNA world: self-excision of intron sequence from nuclear pre-tRNAs.

AUTOR(ES)

Weber, U

RESUMO

The intervening sequences of nuclear tRNA precursors are known to be excised by tRNA splicing endonuclease. We show here that a T7 transcript corresponding to a pre-tRNA(Tyr) from Arabidopsis thaliana has a highly specific activity for autolytic intron excision. Self-cleavage occurs precisely at the authentic 3'-splice site and at the phosphodiester bond one nucleotide downstream of the authentic 5'-splice site. The reaction results in fragments with 2',3'-cyclic phosphate and 5'-OH termini. It is resistant to proteinase K and/or SDS treatment and is not inhibited by added tRNA. The self-cleavage depends on Mg2+ and is stimulated by spermine and Triton X-100. A set of sequence variants at the cleavage sites has been analysed for autolytic intron excision and, in parallel, for enzymatic in vitro splicing in wheat germ S23 extract. Single-stranded loops are a prerequisite for both reactions. Self-cleavage not only occurs at pyrimidine-A but also at U-U bonds. Since intron self-excision is only about five times slower than the enzymatic intron excision in a wheat germ S23 extract, we propose that the splicing endonuclease may function by improving the preciseness and efficiency of an inherent pre-tRNA self-cleavage activity.

Documentos Relacionados

• Targeting genes for self-excision in the germ line
• Plant nonsense suppressor tRNA(Tyr) genes are expressed at very low levels in vitro due to inefficient splicing of the intron-containing pre-tRNAs.
• Wheat germ splicing endonuclease is highly specific for plant pre-tRNAs.
• Substrate recognition by RNase P and by the catalytic M1 RNA: identification of possible contact points in pre-tRNAs.
• Protein splicing of the yeast TFP1 intervening protein sequence: a model for self-excision.