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Antisense noncoding mitochondrial RNA-2 gives rise to miR-4485-3p by Dicer processing in vitro
Nicole Farfán1,2, Nicole Sanhueza1,3, Macarena Briones1
1Fundación Ciencia & Vida/Andes Biotechnologies SpA, Santiago, Chile.
Background:
The antisense noncoding mitochondrial RNAs (ASncmtRNAs) derive from the mitochondrial 16S gene. Knockdown of these transcripts with chemically-modified antisense oligonucleotides induces proliferative arrest, apoptosis and invasiveness reduction in tumor but not normal cells. One of these transcripts, ASncmtRNA-2, contains the complete and identical sequence of hsa-miR-4485-3p and, upon knockdown of this transcript, there is a strong increase in levels of this miRNA, suggesting ASncmtRNA-2 as a source for miR-4485-3p, which is supported by several evidences from our group and others, in the ex vivo setting.
Results:
Here we show that incubation of in vitro-transcribed ASncmtRNA-2 with recombinant Dicer produces RNA fragments corresponding to hsa-miR-4485-3p, showing that Dicer binds to and processes ASncmtRNA-2, strongly supporting the hypothesis that ASncmtRNA-2 acts as a precursor for miR-4485-3p.
Conclusion:
The in vitro results presented here strengthen the hypothesis that miR-4485-3p is derived from ASncmtRNA-2 by Dicer processing. Since miR-4485-3p is classified as a tumor suppressor miRNA, this evidence strengthens the application of ASncmtRNA knockdown for cancer therapy.
Insights
Antisense noncoding mitochondrial RNAs (ASncmtRNAs) like ASncmtRNA-2 can be processed by Dicer into tumor-suppressing microRNAs (miRNAs). This finding supports ASncmtRNA knockdown as a potential cancer therapy.
Area of Science:
- Mitochondrial genetics
- RNA biology
- Cancer therapeutics
Background:
- Antisense noncoding mitochondrial RNAs (ASncmtRNAs) originate from the mitochondrial 16S gene.
- ASncmtRNA knockdown inhibits tumor cell proliferation, induces apoptosis, and reduces invasiveness.
- ASncmtRNA-2 shares sequence identity with hsa-miR-4485-3p, with knockdown increasing miRNA levels.
Purpose of the Study:
- To investigate the processing of ASncmtRNA-2 into hsa-miR-4485-3p.
- To confirm ASncmtRNA-2 as a precursor for hsa-miR-4485-3p via Dicer processing.
- To strengthen the rationale for ASncmtRNA-based cancer therapies.
Main Methods:
- In vitro transcription of ASncmtRNA-2.
- Incubation with recombinant Dicer enzyme.
- Analysis of RNA fragments produced by Dicer processing.
Main Results:
- Recombinant Dicer processed in vitro-transcribed ASncmtRNA-2.
- RNA fragments corresponding to hsa-miR-4485-3p were generated.
- Dicer binding and processing of ASncmtRNA-2 were confirmed.
Conclusions:
- In vitro data support ASncmtRNA-2 as a precursor for hsa-miR-4485-3p via Dicer processing.
- hsa-miR-4485-3p functions as a tumor suppressor miRNA.
- ASncmtRNA knockdown represents a promising strategy for cancer treatment.
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