(1) THE First demonstration of non-transcriptional circadian oscillations in eukaryotic cells
We published two papers (O’Neill & Reddy Nature 2011; and O’Neill et al. Nature 2011) demonstrating the existence of circadian oscillations in the absence of transcription in two completely different model systems - human red blood cells and an alga called Ostreococcus tauri. This work is important because it highlights that circadian oscillations can occur in eukaryotes without genetic feedback loops – there is no transcription in red blood cells, or in algae held in the dark.
Circadian clocks in red blood cells
RNA synthesis (mRNA transcription) is not needed for 24 hour oscillations in the alga Ostreococcus tauri.
(2) the pervasiveness of circadian redox oscillations in evolution
Our lab subsequently went on to demonstrate that redox oscillations in the form of rhythms in oxidation of the peroxiredoxin protein family are evolutionarily conserved in the three domains of life – bacteria, archaea and eukaryotes (Edgar et al. Nature 2012). This study has far reaching consequences for understanding how clocks arose, and the mechanisms by which they keep time. Before this study, it was thought that circadian clocks had evolved by convergent evolution in different species, whereas our work indicates that they may have instead evolved divergently, from a common origin.
(3) gating of virus infection by the circadian clock
Since viruses are obligate intracellular pathogens, they must infect cells of an organism to reproduce. In this paper, we showed that the clockwork regulates the infection of two unrelated and common types of virus, herpesvirus and influenza A, such that time of day impacts on infectivity (Edgar et al. PNAS 2016). Importantly, we demonstrated that the clock gene Bmal1 is critical in restraining viral infection by quite diverse intracellular pathways that were targeted differentially by the two types of virus. Loss of Bmal1 leads to increased kinetics and overall levels of infection both in vivo and in cell culture, i.e. in the absence of the immune system. This work thus highlights a novel host factor (the circadian clock) that needs to be carefully considered in infectious diseases and in vaccination programmes in order to optimize therapy and prevention.
(4) Glucose metabolism impacts on the clockwork
Metabolism and the clockwork are heavily intertwined. In this paper, we identified the pentose phosphate pathway (PPP) as an important clock regulator (Rey et al. Cell Metabolism 2016). The PPP is a key glucose-metabolising pathway regulates the central redox factor NADPH, and as such we suggest a mechanism by which NADPH controls the redox transcription factor NRF2 to direct 24-hour gene expression programmes. Importantly, we showed that the PPP regulates the clock in both invertebrates (fruit flies) and vertebrates (mammalian cells and tissue), indicating an evolutionarily conserved role for this pathway in clock regulation.
(5) Daily oscillations in chromatin architecture
In this paper (Valekunja et al. PNAS 2013), we demonstrated that huge proportions of the liver's epigenome underwent circadian regulation, and that particular activatory chromatin marks (histone methylation) underwent this rhythmic change. We went on to identify that the histone methyltransferase MLL3, which is part of a highly conserved family of such enzymes, plays an important part in bringing about this change. Importantly, loss of MLL3's enzymatic activity severely perturbs cellular circadian oscillations, implicating rhythmic histone methylation as an important contributor to circadian transcription.