Role of SUV3 Helicase in Maintaining Mitochondrial Homeostasis in Human Cells

Abstract

In yeast mitochondria, RNA degradation takes place through the coordinated activities of ySuv3 helicase and yDss1 exoribonuclease (mtEXO), whereas in bacteria, RNA is degraded via RNaseE, RhlB, PNPase, and enolase. Yeast lacking the Suv3 component of the mtEXO form petits and undergo a toxic accumulation of omega intron RNAs. Mammalian mitochondria resemble their prokaryotic origins by harboring a polyadenylation-dependent RNA degradation mechanism, but whether SUV3 participates in regulating RNA turnover in mammalian mitochondria is unclear. We found that lack of hSUV3 in mammalian cells subsequently yielded an accumulation of shortened polyadenylated mtRNA species and impaired mitochondrial protein synthesis. This suggests that SUV3 may serve in part as a component of an RNA degradosome, resembling its yeast ancestor. Reduction in the expression levels of oxidative phosphorylation components correlated with an increase in reactive oxygen species generation, whereas membrane potential and ATP production were decreased. These cumulative defects led to pleiotropic effects in mitochondria such as decreased mtDNA copy number and a shift in mitochondrial morphology from tubular to granular, which eventually manifests in cellular senescence or cell death. Thus, our results suggest that SUV3 is essential for maintaining proper mitochondrial function, likely through a conserved role in mitochondrial RNA regulation.Prokaryotes and eukaryotes both utilize RNA processing and degradation, albeit via different pathways, as one mechanism for controlling gene expression (1-4). The RNA degradosome of Escherichia coli, yeast, and mammalian cells have all conserved the ability to turnover RNA; however, the assembly of their multicomponent machineries differ (5, 6). In E. coli, RNA degradation is conducted by the cooperation of four principal enzymes (7). The first enzyme is an endoribonuclease, RNase E, which initiates the turnover of many RNAs by cleaving singlestranded RNA internally and is essential for cell growth (2,8,9). The second enzyme is a 50-kDa DEAD-box helicase, RhlB, which unwinds and translocates RNA substrates (10). RhlB facilitates the degradation of structured mRNA decay intermediates by the third 85-kDa enzyme, polynucleotide polymerase (PNPase).3 This process, requiring ATP hydrolysis, is believed to involve the local unwinding of structures that block PNPase, thereby ensuring rapid degradation by PNPase (11-13). The fourth enzyme, a 48-kDa glycolytic enzyme enolase, associates with RNase E in response to stress caused by the overabundance of phosphosugars (14, 15). However, enolase is a unique and mysterious contributor to the degradosome, as it is not an RNA-binding protein nor does it appear to be directly involved in RNA turnover.In Saccharomyces cerevisiae, mitochondrial RNA degradation also requires the activity of multifunctional components (mtEXO) (16), which consist mainly of two enzymes: ySuv3 and yDss1 (17)(18)(19)(20). Yeast Suv3 is a nuclear gene-encoded protein that localizes to mitochondria in ...

Uncoupling the Roles of the SUV3 Helicase in Maintenance of Mitochondrial Genome Stability and RNA Degradation

“…Membrane Potential and ATP Production Detection-Methods were adapted from a previous publication (30). Briefly, 5 ml of yeast cultures after depletion of SUV3 in glucose for 24 h were washed with 1ϫ PBS and incubated with 50 nM rhodamine 123 for 15 min.…”

“…Quantitative PCR was performed as described previously using an iCycler (Bio-Rad) (30,39). The relative mtDNA copy number was calculated as a ratio between the abundances of mitochondrial COX1/nuclear Actin1.…”

“…The extent to which the intron contributes to both processes also warrants a thorough investigation. Although it seems plausible that reopening of mRNAs may disrupt mitochondrial translation and affect respiratory chain function, leading to increased reactive oxygen species production that damages mtDNA 5 and contributes to the loss of mitochondrial DNA (30,35), a direct role of SUV3 in the maintenance of mitochondrial genomic stability cannot be ruled out.…”

“…The human mitochondrial degradosome, on the other hand, does not resemble the low eukaryotic counterpart because the yeast exoribonuclease, DSS1, has not been identified in humans (27)(28)(29). Instead, the 330-kDa human mitochondrial SUV3-PNPase complex of degradosome seems to function as an alternative form of E. coli degradosome lacking RNase E. The inactivation of SUV3 in the human system leads to disruption of mitochondrial homeostasis, which eventually results in cell senescence or death (30).…”

Conditional control of gene function by an invertible gene trap in zebrafish

“…It is an essential component of the mitochondrial degradosome complex that regulates mtRNA decay and mRNA surveillance (19). Human SUPV3L1 is important for maintaining mitochondrial homeostasis (20) and has been shown to degrade double-stranded RNA (21). However, SUPV3L1 also appears to function in the cytoplasm, interacting with the WRN and BLM DNA helicases to suppress mitotic homologous recombination (13).…”