“During mammalian aging, there are changes in abundance of noncoding RNAs including microRNAs, long noncoding RNAs, and circular RNAs.”
The aging of an organism is reflected not only in the function of its organs but also in the molecular signatures written into its cells. For years, scientists have cataloged the changes in protein-coding genes and various non-coding RNAs that occur as we grow older. However, one class of molecules—circular RNAs originating from the genome of our cellular power plants, the mitochondria—has remained largely unexplored.
A new research paper, titled “Aging-associated mitochondrial circular RNAs” published in Volume 18 of Aging-US by a multi-institutional team of researchers, provides the first detailed profile of these molecules and reveals a surprising link to cellular energy metabolism.
The team’s investigation demonstrates that a specific mitochondrial circular RNA, circMT-RNR2, is depleted in older individuals and plays a direct role in regulating the TCA cycle, the engine of cellular energy production.
The Discovery: A Mitochondrial Circular RNA Lost with Age
The researchers began by analyzing circular RNA junctions in peripheral blood mononuclear cells (PBMCs) from 11 young adults (average age 30) and 11 older adults (average age 64). Using RNA sequencing data, they identified hundreds of circular RNA species.
The most striking finding was the source of these molecules. In young individuals, the vast majority of circular RNA junctions originated from the mitochondrial chromosome (chrM). Specifically, the most abundant circular RNAs were derived from a mitochondrial ribosomal RNA gene called MT-RNR2. In older individuals, however, these same circular RNA junctions were sharply depleted—a loss of nearly 90%.
This age-associated decline was not just a statistical observation. When the team examined human fibroblasts (skin cells) as they aged in culture, they saw the same pattern: levels of circMT-RNR2 dropped progressively as the cells approached senescence, the point at which they permanently stop dividing.
The Regulator: An RNA-Binding Protein Called GRSF1
If circMT-RNR2 disappears with age, what controls its production? The team turned their attention to GRSF1, a protein known to localize to mitochondrial RNA granules—specialized compartments where mitochondrial RNAs are processed.
Using a split-GFP system, they confirmed that GRSF1 resides within mitochondria. They then performed a PAR-CLIP analysis, a technique that identifies precisely which RNAs a protein binds to. The results showed that GRSF1 binds directly to several mitochondrial transcripts, including both the linear and circular forms of MT-RNR2. A specific RNA motif—UGxxGGUU—was identified as the recognition sequence for GRSF1 on its target RNAs.
When the researchers depleted GRSF1 from human fibroblasts, circMT-RNR2 levels plummeted. This established GRSF1 as a critical factor for maintaining the abundance of this mitochondrial circular RNA.
The Function: Scaffolding the TCA Cycle
The discovery that a circular RNA is lost with age raised an obvious question: what does it actually do? Given that MT-RNR2 originates from the mitochondria, the team hypothesized it might be involved in mitochondrial metabolism.
They performed RNA immunoprecipitation assays to see if circMT-RNR2 interacts with metabolic enzymes. The results revealed that both linear and circular MT-RNR2 bind to two key enzymes of the TCA cycle: SUCLG1 (part of succinyl-CoA synthetase) and SDHA (a component of succinate dehydrogenase complex II).
This binding appears to have functional consequences. When the team depleted MT-RNR2 from cells, levels of the TCA cycle metabolites fumarate and alpha-ketoglutarate declined. Conversely, reintroducing circMT-RNR2 restored fumarate levels. The circular RNA seemed to be acting as a scaffold, helping to assemble or stabilize the enzyme complexes that drive the TCA cycle.
The Consequence: Suppressing Cellular Senescence
If circMT-RNR2 supports energy production, its loss should accelerate aging at the cellular level. To test this, the team measured markers of cellular senescence—p16 and p21—after manipulating GRSF1 and circMT-RNR2.
Depleting GRSF1, which reduced circMT-RNR2, caused a sharp increase in p16 and p21 mRNA levels. However, when they reintroduced circMT-RNR2 into these GRSF1-depleted cells, the senescence markers returned to normal. The circular RNA alone was sufficient to reverse the senescence phenotype.
Further analysis showed that GRSF1 depletion broadly suppressed mitochondrial transcripts, and reintroducing circMT-RNR2 partially rescued this defect. The model that emerges is one where GRSF1 promotes the production of circMT-RNR2, which then scaffolds TCA cycle enzymes to maintain efficient energy production and keep cells in a proliferating, non-senescent state.
Implications for Future Research
This study opens several new avenues for investigation. First, it establishes that mitochondria produce circular RNAs with distinct functions, expanding our understanding of mitochondrial biology. Second, it identifies GRSF1 as a key regulator of these molecules, linking RNA-binding proteins to mitochondrial metabolism.
The finding that a single circular RNA can influence the entire TCA cycle suggests that non-coding RNAs may play broader roles in metabolism than previously appreciated. The authors propose that circMT-RNR2 may act similarly to other scaffold non-coding RNAs, like NEAT1, which assemble metabolic enzymes to accelerate biochemical reactions.
The mechanism by which MT-RNR2 produces a circular RNA remains intriguing. Since the gene lacks introns, conventional back-splicing cannot explain its circularization. The authors speculate that trans-splicing—a process more common in plants and trypanosomes—may be at work, potentially mediated by GRSF1 within mitochondrial RNA granules.
Future Perspectives and Conclusion
This research does not claim to have fully mapped the landscape of mitochondrial circular RNAs or their functions. Rather, it offers a compelling proof-of-concept that these molecules exist, change with age, and have measurable biological effects.
By integrating transcriptomic profiling, biochemical analysis, and functional studies, the team demonstrates that circMT-RNR2 is depleted during human aging and senescence, that it is regulated by GRSF1, and that it supports the TCA cycle by scaffolding metabolic enzymes.
The perspective that emerges is one where the mitochondria are not just passive energy generators but active participants in the aging process through their non-coding RNA output. Continued research will be needed to determine whether other mitochondrial circular RNAs have similar functions, how precisely they are generated, and whether they might serve as therapeutic targets to preserve metabolic health in older age.
Click here to read the full research paper published in Aging-US.
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