The intervening sequence excised from the ribosomal RNA precursor of Tetrahymena contains a 5-terminal guanosine residue not encoded by the DNA

Abstract

The ribosomal RNA precursor of Tetrahymena thermophila contains a 0.4 kilobase intervening sequence that is excised as a linear RNA molecule ("IVS RNA") and subsequently cyclized. In vitro transcription in isolated nuclei was used to accumulate the IVS RNA labeled at its 5' end was subjected to sequencing gel analysis and terminal nucleotide analysis. In addition, uniformly labeled IVS RNA was cleaved with RNAase T1, and the resulting oligonucleotides were studied by two-dimensional fingerprinting. The IVS RNA was found to be a unique molecule with no discernible terminal heterogeneity. The 5'-terminal nucleotide is a guanosine that is not present at the corresponding point in the DNA sequence, determined by N. Kan and J. Gall (see adjoining paper). This nucleotide is added to the IVS during splicing [Cech, Zaug, and Grabowski (1981) Cell 27, 487-496]. Based on the sequences near the ends of the RNA, the remainder of the RNA sequence is colinear with that of the DNA. The IVS RNA has 5'-monophosphate and 3'-hydroxyl termini. Comparison of these results to those obtained previously for yeast tRNA intervening sequences leads us to conclude that the splicing mechanisms are fundamentally different for these two classes of transcripts.