Peroxiredoxins are conserved markers of circadian rhythms
Summary (3 min read)
Peroxiredoxins are conserved markers of
- The molecular phylogeny of mechanisms driving these rhythms has been difficult to dissect because identified clock genes and proteins are not conserved across the domains of life: Bacteria, Archaea and Eukaryota.
- Therefore, across phylogenetic kingdoms, there are apparently no common ‘clock’ components, suggesting that daily timekeeping evolved independently within different lineages.
- The authors previously characterized circadian cycles of peroxiredoxin oxidation using antiserum directed against the oxidized active site, which recognizes both over- (PRX-SO2) and hyper- (PRX-SO3) oxidized forms9,10,14.
- 7MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, UK.
Peroxiredoxin rhythms in eukaryotes
- Using the PRX-SO2/3 antiserum, the authors first examined circadian time courses from a range of eukaryotes under constant conditions (that is, in the absence of external timing cues).
- To extend these findings beyond vertebrates, the authors examined peroxiredoxin rhythms in the fruitfly Drosophila melanogaster.
- The authors pooled whole heads from insects maintained in constant darkness over two circadian cycles after they had been stably entrained to 12 h light, 12 h dark cycles.
- Again, circadian oscillations in PRX-SO2/3 immunoreactivity were observed, as well as in the clock protein Timeless (TIM) (Fig. 2b).
- Similarly, seedlings from the plant Arabidopsis thaliana exhibited robust PRX-SO2/3 oscillations in free-running conditions of constant light, which were also seen in the filamentous fungus Neurospora crassa—another well-characterized clock model system (Fig. 2c and Supplementary Fig. 2).
Peroxiredoxin rhythms in prokaryotes
- Having observed ,24 h peroxiredoxin oxidation rhythms in organisms with nucleated cells, the authors next sought to examine representative prokaryotes from each major domain—Bacteria and Archaea.
- Because all three Kai proteins are expressed together in only a small number of bacterial species, and in no known archaea18, this system cannot represent a general prokaryotic clock mechanism.
- The authors postulated that, regardless of the timekeeping mechanism, the peroxiredoxin oxidation cycles they observe reflect an absolutely conserved rhythmic cellular output.
- This indicates the presence of an underlying capacity for circadian timing in both mutant strains, which was clearly perturbed by the absence of functional transcriptional feedback circuitry (Fig. 4a).
- To establish the wider relevance of these findings, the authors also examined similar mutants in the fungus Neurospora crassa.
Implications for clock evolution
- The authors have observed ,24 h cycles of peroxiredoxin oxidation–reduction in all domains of life and consequently, the possibility that cellular rhythms share a common molecular origin seems increasingly plausible.
- Approximately 2.5 billion years ago, photosynthetic bacteria acquired the capacity for photo-dissociation of water, leading to the geologically rapid accumulation of molecular oxygen during the Great Oxidation Event (GOE), when anaerobic life underwent a catastrophic decline35.
- If there was considerable pressure for the co-evolution of metabolic/ROS pathways with cellular timekeeping systems, then evidence for this should exist in the phylogenetic trees of their component mechanisms.
- See Supplementary Table 9 for cycle period estimates (by harmonic regression) and detailed statistics (by ANOVA), as well as Supplementary Fig. 10 for TIM and FRQ immunoblots for fruitfly and fungus, respectively.
- This suggests that similarities in the evolutionary profiles of these cellular mechanisms go beyond those that would be expected simply based on the time since a common ancestry, because even highly conserved proteins had considerably inferior correlations to peroxiredoxin (Supplementary Fig. 6 and Supplementary Table 8)42.
Concluding remarks
- It has long been recognized that oxygen-sensing PAS-domain proteins are involved in the clockwork of many eukaryotes, but the rationale behind this has remained elusive23,43–45.
- In light of their current findings, the authors speculate that sensing and responding to oxidative cycles in cellular environments could have driven the evolution of circadian rhythms, and maintained the intrinsic link between clocks and metabolism (Fig. 6).
- A direct prediction therefore, is that organisms that lack ROS detoxification systems will not have circadian rhythms.
- At least one such class of organism exists on Earth, an example being the hyperthermophilic archaea Methanopyri (NCBI taxonomy accession 183988).
- Given its distinct anoxic environmental niche and methanogenic metabolism46, there may never have been a selective evolutionary pressure to develop circadian timekeeping as the authors know it.
METHODS SUMMARY
- A. thaliana, D. melanogaster, H. salinarum sp. NRC-1, M. musculus, N. crassa, O. tauri and S. elongatus were bred, grown or cultured in appropriate conditions, synchronized by specific methods normally used in each organisms, and then sampled under constant conditions of either darkness or light, depending on the organism.
- Immunoblotting was performed after protein transfer to nitrocellulose membranes.
- Interspecies plots were generated, which contain a simplified representation of the correlation between the interprotein distances in phylogenetic trees for each protein being compared.
- Statistical comparisons between Mirrortree correlation coefficients were performed as detailed above.
- C, Co-evolution of cyanobacteria oscillator components and peroxiredoxin proteins.
Published online 16 May 2012.
- The adaptive value of circadian clocks; an experimental assessment in cyanobacteria.
- The oligomeric conformation of peroxiredoxins links redox state to function.
METHODS
- Equal protein loading was confirmed by gel electrophoresis and Coomassie staining of gels loaded with equal volumes of lysate from each time point in each replicate set.
- Circadian time (CT) 0 was operationally defined as the nadir in bioluminescence signal, and CT12 was taken to be at the peak.
- Correlation coefficients obtained for analyses of KaiC distance versus other conserved bacterial proteins (including KaiB, KaiC and PRX2) were compared by converting them to a normally distributed metric using Fisher’s r-to-z transformation: r0~ 1 2 ln 1zr 1{r in which r is the Mirrortree correlation coefficient; and r9 is the Fisher-transformed correlation coefficient.
CORRECTIONS & AMENDMENTS
- In the author list of this Article, the names of Gerben van Ooijen and Maria Olmedo should also have been asterisked, indicating their equal contributions.
- This error has been corrected in the HTML and PDF versions of the original paper.
- *These authors contributed equally to this work.
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104 citations
Cites background from "Peroxiredoxins are conserved marker..."
...As it preceded the appearance of the latter mechanism in the course of evolution, it represents the oldest circadian oscillatory mechanism identified so far at the cellular level [43]....
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103 citations
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Cites background from "Peroxiredoxins are conserved marker..."
...Indeed, cyanobacteria are remarkable organisms: their evolution of photosynthesis led to the creation of chloroplasts and higher plants, but even before then they transformed life on Earth with the Great Oxygenation Event (GOE) [10]....
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...This reciprocal coupling of electrical firing and transcription likely contributes to the stability of the SCN clock mechanism, and the metabolic demands of such rhythms are reflected by a circadian cycle of PRDX superoxidation, which peaks in the SCN in circadian night [14]....
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...This oscillation in redox-regulatory capacity is also seen in the archaeon Halobacterium salinarum, identifying its ancient evolutionary origin, which may well have been in response to the challenges posed by increased oxygenation at the GOE [14]....
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101 citations
Cites background from "Peroxiredoxins are conserved marker..."
...5 billion years, and suggests that circadian rhythmicity has conferred an evolutionary advantage since bacteria first became able to photo-dissociate water (thereby producing the selective pressure of atmospheric oxygen) [7]....
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References
2,723 citations
"Peroxiredoxins are conserved marker..." refers background in this paper
...Importantly, we note that (1) during the GOE, rhythms of O2 production/consumption and ROS generation would have been driven by the solar cycle, as they are today; (2) metabolic/oxidation rhythms seem to be present in every organism with a circadian clock, all of which are aerobes, and these rhythms persist in the absence of transcriptional cycles; and (3) circadian timekeeping confers a selective advantage when it facilitates anticipation of environmental change (noxious or otherwise)....
[...]
2,010 citations
1,870 citations
1,276 citations
"Peroxiredoxins are conserved marker..." refers background in this paper
...Importantly, we note that (1) during the GOE, rhythms of O2 production/consumption and ROS generation would have been driven by the solar cycle, as they are today; (2) metabolic/oxidation rhythms seem to be present in every organism with a circadian clock, all of which are aerobes, and these rhythms persist in the absence of transcriptional cycles; and (3) circadian timekeeping confers a selective advantage when it facilitates anticipation of environmental change (noxious or otherwise)....
[...]
1,261 citations