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Both yeast W double-stranded RNA and its single-stranded form 20S RNA are linear
N Rodriguez-Cousiño1, R Esteban
1Instituto de Microbiología Bioquímica, Consejo Superior de Investigaciones Científicas/Universidad de Salamanca, Spain.
Nucleic Acids Research
|June 11, 1992
Summary
This study demonstrates that both the W double-stranded RNA (dsRNA) and 20S single-stranded RNA (ssRNA) in yeast are linear molecules. These findings clarify the structure of these important RNA species under stress conditions.
Area of Science:
- Molecular Biology
- Yeast Genetics
- RNA Biology
Background:
- Most yeast strains harbor a cytoplasmic 2.5 kb double-stranded RNA (dsRNA) molecule known as W.
- W (+) strands were previously proposed to be identical to 20S RNA, a single-stranded RNA (ssRNA) induced by stress.
- A recent hypothesis suggested 20S RNA might be circular.
Purpose of the Study:
- To investigate and clarify the structural nature of W dsRNA and 20S RNA in yeast.
- To experimentally determine if W dsRNA and 20S RNA are linear or circular.
Main Methods:
- Stoichiometric labeling of W dsRNA strands using 32P-pCp and T4 RNA ligase to assess linearity.
- Site-specific cleavage of 20S RNA using RNase H and a complementary oligodeoxynucleotide to confirm linearity.
Main Results:
- Stoichiometric labeling confirmed the presence of both RNA strands in W dsRNA, indicating linearity.
- The 3' end nucleotides of both W dsRNA strands were predominantly cytosine (C) or adenine (A).
- RNase H cleavage of 20S RNA produced two distinct fragments, directly supporting its linear structure.
Conclusions:
- Both W dsRNA and 20S RNA in yeast are definitively linear molecules.
- This research refutes the recent proposal of circular 20S RNA.
- Establishes the linear nature of these RNAs, crucial for understanding their function and replication in yeast.