Critical factors for precise and efficient RNA cleavage by RNase Y in Staphylococcus aureus

Area of Science:

• Molecular Biology
• Microbiology
• Genetics

Background:

• Cellular processes rely on precise gene regulation, with mRNA degradation being crucial.
• RNase Y, an endoribonuclease in Firmicutes (e.g., Bacillus subtilis, Staphylococcus aureus), plays a key role in RNA degradation.
• The specific molecular interactions dictating RNase Y's RNA targeting and cleavage sites remain largely uncharacterized.

Purpose of the Study:

• To identify conserved RNase Y target transcripts in Staphylococcus aureus and Bacillus subtilis.
• To elucidate the molecular determinants governing RNase Y's RNA cleavage specificity and positioning.
• To demonstrate the sufficiency of identified sequence and structural elements for RNase Y-mediated cleavage.

Main Methods:

• Comparative transcript analysis to identify homologous RNase Y targets in S. aureus and B. subtilis.
• Utilizing identified homologous transcript pairs as model systems to study RNase Y function.
• Experimental manipulation of RNA sequences and structures to assess their impact on RNase Y cleavage.
• Conversion of a non-target transcript into an RNase Y target using identified sequence elements.

Main Results:

• Identified conserved RNase Y target transcripts common to both S. aureus and B. subtilis.
• Demonstrated functional overlap in RNase Y activity between the two bacterial species.
• Established that RNA's primary nucleotide sequence downstream of the cleavage site and secondary structure influence cleavage efficiency.
• Showed that downstream secondary structure broadly localizes cleavage, while the upstream nucleotide fine-tunes positioning.
• Confirmed that identified sequence elements are sufficient to confer RNase Y-dependent cleavage to non-target RNAs.

Conclusions:

• RNase Y's targeting and cleavage are primarily dictated by specific RNA sequence and secondary structure features.
• These findings reveal fundamental principles of RNA recognition and processing by RNase Y.
• The identified elements provide a basis for predicting and engineering RNase Y activity, with implications for understanding bacterial gene regulation.