The relatively recent focus on the widespread occurrence and abundance of circular RNAs (circRNA) in the human cell nucleus has sparked an intensive interest in their existence and possible roles in cell gene expression and physiology. The presence of circRNAs in mammalian mitochondria, however, has been under-explored. Mitochondrial mRNAs differ from those produced from nuclear genes because they lack introns and are transcribed as poly-cistronic transcripts that are endonucleolytically cleaved, leaving transcripts with very small 5′ and 3′ UTRs. Circular RNAs have been identified in the semi-autonomous organelles of single-celled organisms and plants but their purpose has not been clearly demonstrated. The goal of our project was to test the hypothesis, processed mRNAs are circularized in vertebrate mitochondria as a necessary RNA processing step prior to translation. Mitochondrial mRNAs were isolated from the human cell line HEK293 and evidence of circularization sought by treating RNA with RNAse-R and then amplifying putative 3′-5′ junction sites. Sequence results demonstrated the occurrence of mRNA circularization within each coding region of the mitochondrial genome. However, in most cases the circRNAs carried coding regions that had been truncated, suggesting they were not translatable. Quantification of the circularized versions of the mRNAs revealed they comprise a small portion (~10%) of the total mRNA. These findings demonstrate that mRNA circularization occurs in mammalian mitochondria but it does not appear to play a role in making translatable mRNAs.
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Mitochondrial mRNA Processing in the Chlorophyte Alga Pediastrum duplex and Streptophyte Alga Chara vulgaris Reveals an Evolutionary Branch in Mitochondrial mRNA Processing
Mitochondria carry the remnant of an ancestral bacterial chromosome and express those genes with a system separate and distinct from the nucleus. Mitochondrial genes are transcribed as poly-cistronic primary transcripts which are post-transcriptionally processed to create individual translationally competent mRNAs. Algae post-transcriptional processing has only been explored in Chlamydomonas reinhardtii (Class: Chlorophyceae) and the mature mRNAs are different than higher plants, having no 5′ UnTranslated Regions (UTRs), much shorter and more variable 3′ UTRs and polycytidylated mature mRNAs. In this study, we analyzed transcript termini using circular RT-PCR and PacBio Iso-Seq to survey the 3′ and 5′ UTRs and termini for two green algae, Pediastrum duplex (Class: Chlorophyceae) and Chara vulgaris (Class: Charophyceae). This enabled the comparison of processing in the chlorophyte and charophyte clades of green algae to determine if the differences in mitochondrial mRNA processing pre-date the invasion of land by embryophytes. We report that the 5′ mRNA termini and non-template 3′ termini additions in P. duplex resemble those of C. reinhardtii, suggesting a conservation of mRNA processing among the chlorophyceae. We also report that C. vulgaris mRNA UTRs are much longer than chlorophytic examples, lack polycytidylation, and are polyadenylated similar to embryophytes. This demonstrates that some mitochondrial mRNA processing events diverged with the split between chlorophytic and streptophytic algae.
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- Award ID(s):
- 1919491
- NSF-PAR ID:
- 10298147
- Date Published:
- Journal Name:
- Plants
- Volume:
- 10
- Issue:
- 3
- ISSN:
- 2223-7747
- Page Range / eLocation ID:
- 576
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/plants10030576
