Nucleotide Sequence Requirements for Self-cleavage of Neurospora VS RNA

• Published in: Journal of molecular biology Vol. 232; no. 2; pp. 351 - 361
• Main Authors: Guo, Hans C.T., De Abreu, Diane M., Tillier, Elisabeth R.M., Saville, Barry J., Olive, Joan E., Collins, Richard A.
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 1993; Elsevier
• Subjects: Analytical, structural and metabolic biochemistry; Base Sequence; Biological and medical sciences; catalytic RNA; chemical reactions; Consensus Sequence; DNA Mutational Analysis; Fundamental and applied biological sciences. Psychology; genbank/neumtvs (sic); Genetic Variation; horizontal transfer; mitochondria; Mitochondria - metabolism; mitochondrial plasmid; Molecular Sequence Data; Mutagenesis, Insertional; Neurospora; Neurospora - genetics; Neurospora - metabolism; neurospora intermedia; Neurospora sitophila; Nucleic acids; nucleotide sequences; Phylogeny; Plasmids - genetics; ribozyme; ribozymes; RNA; RNA Processing, Post-Transcriptional; RNA structure; RNA, Catalytic - genetics; RNA, Catalytic - metabolism; RNA, Fungal - genetics; RNA, Fungal - metabolism; Rna, ribonucleoproteins; Sequence Analysis, DNA; Sequence Homology, Nucleic Acid; Structure-Activity Relationship; catalytic RNA; horizontal transfer; RNA structure; mitochondrial plasmid; ribozyme; Fungi; Horizontal transfer; Ribozyme; Site directed mutagenesis; Deletion; Neurospora sitophila; Cleavage; Ascomycetes; Autocatalysis; In vitro; Structure function relationship; Thallophyta
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1006/jmbi.1993.1395

We have used several complementary approaches to investigate the minimal contiguous sequence required for the in vitro self cleavage reaction performed by Neurospora VS RNA. Deletion analysis and site-directed mutagenesis revealed that only a single nucleotide is required upstream of the self-cleavage site, and that the identity of this nucleotide is not critical. This distinguishes VS RNA from all currently known ribozymes except hepatitis delta virus RNA. The shortest contiguous sequence capable of cleavage contains 153 nt downstream of the cleavage site. Linker insertion mutagenesis suggests that much of this downstream sequence is important for self-cleavage. Comparative sequence analysis of the VS plasmid from six natural isolates supports the importance in vivo of the minimal region determined by in vitro methods. Also, phylogenetic analysis raises the possibility of a recent horizontal transfer of the VS plasmid from Neurospora intermedia to Neurospora sitophila.