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The RNA component of mitochondrial ribonuclease P from Aspergillus nidulans
1Department of Microbiology, College of Natural Sciences, Seoul National University, Korea.
Abstract:
Several RNA molecules that copurified with Aspergillus nidulans mitochondrial ribonuclease (RNase) P were identified [Lee, Y C., Lee, B. J., Hwang, D. S. & Kang, H. S. (1996) Eur J. Biochem. 235, 289-296], and their partial sequences were determined. Using an oligonucleotide probe, we cloned and mapped the gene encoding this putative RNA component of RNase P (RNase P-RNA), situated between URFA3 (unidentified reading frame A3) and cobA (apocytochrome b) genes in the mitochondrial genome of A. nidulans. The gene is extremely (A+T)-rich and contains two regions of sequence similarity conserved among the known mitochondrial RNase P-RNAs and the eubacterial RNase P-RNAs. The determination of 5' and 3' termini by primer extension and sequencing indicated that the length of the RNA transcript is 232 nucleotides. Northern-blot analysis revealed that its only subcellular location was the mitochondria. Two RNase P-RNA fragments of 110 nucleotides and 80 nucleotides, each containing one of the two conserved regions, could be recovered from the nuclease-treated enzyme without significant loss of activity. The sizes of these fragments appeared to be the minimum lengths required for the vitro activity of the enzyme.
Insights
Researchers identified the gene for mitochondrial ribonuclease (RNase) P RNA in Aspergillus nidulans. This RNA component is essential for the enzyme's activity, with specific fragments retaining catalytic function.
Area of Science:
- Molecular Biology
- Mitochondrial Genetics
- Enzymology
Background:
- Ribonuclease P (RNase P) is a crucial enzyme involved in RNA processing.
- Mitochondrial RNase P in eukaryotes has been less characterized compared to its bacterial counterparts.
- Previous studies identified RNA molecules associated with Aspergillus nidulans mitochondrial RNase P.
Purpose of the Study:
- To clone and map the gene encoding the RNA component of Aspergillus nidulans mitochondrial RNase P (RNase P-RNA).
- To characterize the sequence and structure of the A. nidulans mitochondrial RNase P-RNA.
- To investigate the functional significance of conserved regions within the RNase P-RNA.
Main Methods:
- Oligonucleotide probe hybridization for gene cloning and mapping.
- Primer extension and sequencing for determining RNA termini and length.
- Northern-blot analysis for subcellular localization.
- Enzymatic assays with nuclease-treated enzyme fragments.
Main Results:
- The gene for A. nidulans mitochondrial RNase P-RNA was successfully cloned and mapped within the mitochondrial genome.
- The gene is highly (A+T)-rich and encodes a 232-nucleotide RNA transcript localized exclusively to mitochondria.
- Conserved sequence regions within the RNA are essential for catalytic activity, as demonstrated by functional fragments.
Conclusions:
- The identified gene encodes the functional RNA subunit of Aspergillus nidulans mitochondrial RNase P.
- The study provides insights into the structure-function relationship of mitochondrial RNase P-RNA.
- This work contributes to understanding the evolution and mechanisms of RNA processing in mitochondria.