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Minicircle-encoded guide RNAs from Crithidia fasciculata
1Howard Hughes Medical Institute, University of California, Los Angeles 90095, USA.
RNA (New York, N.Y.)
|August 1, 1995
Summary
Guide RNAs (gRNAs) are encoded in kinetoplast minicircle DNA in Crithidia fasciculata, despite previous assumptions. Minor minicircle classes encode functional gRNAs essential for RNA editing.
Area of Science:
- Molecular Biology
- Genetics
- Parasitology
Background:
- Mitochondrial RNA editing via guide RNAs (gRNAs) is known in Crithidia fasciculata.
- Evidence for gRNA encoding within kinetoplast minicircle DNA was previously lacking.
- Concerns existed about the capacity of a single minicircle sequence class to encode diverse gRNAs.
Purpose of the Study:
- To investigate the encoding of gRNAs within kinetoplast minicircle DNA in Crithidia fasciculata.
- To identify specific gRNAs and their target cryptogenes (MURF4, RPS12).
- To determine the genomic location and sequence class of gRNA-encoding minicircles.
Main Methods:
- Determined genomic and mature edited sequences of MURF4 and RPS12 cryptogenes.
- Constructed a gRNA library from mitochondrial RNA.
- Analyzed minicircle DNA sequence classes and gRNA gene localization.
Main Results:
- Identified five specific gRNAs, three for RPS12 mRNA and two for MURF4 mRNA.
- Localized gRNA genes to specific minicircle molecules within variable regions.
- Found gRNA-encoding minicircles constitute minor sequence classes (approx. 2%).
Conclusions:
- Kinetoplast minicircle DNA molecules in Crithidia fasciculata encode gRNAs, confirming findings in other trypanosomatids.
- Specific minicircle sequence class copy numbers can vary significantly without impacting the RNA editing system.
- This study provides the first evidence for gRNA encoding in C. fasciculata minicircle DNA.