Circadian Oscillators: Around the Transcription-Translation Feedback Loop and on to Output

Abstract

From cyanobacteria to mammals, organisms have evolved timing mechanisms to adapt to environmental changes in order to optimize survival and improve fitness. To anticipate these regular daily cycles, many organisms manifest ∼24h cell-autonomous oscillations that are sustained by transcription-translation-based or post-transcriptional negative-feedback loops that control a wide range of biological processes. With an eye to identifying emerging common themes among cyanobacterial, fungal, and animal clocks, some major recent developments in the understanding of the mechanisms that regulate these oscillators and their output are discussed. These include roles for antisense transcription, intrinsically disordered proteins, codon bias in clock genes, and a more focused discussion of post-transcriptional and translational regulation as a part of both the oscillator and output.

Keywords: PTO; TTFL; antisense.; clock; intrinsically disordered protein ;codon bias.

Figure 1

Minimal schematic representations of circadian model systems. A. Cyanobacteria. The Post Translational Oscillator (PTO) composed of KaiA, B, and C is seen at the top and output from it, which drives the Transcription Translation Feedback Loop (TTFL) as well as clock-controlled genes (ccgs), is at the bottom. Briefly, the core PTO oscillation derives from the autokinase-autophosphatase activity of KaiC. Kai A promotes the autokinase, and Kai B promotes the autophosphatase activity and traps KaiA into inactive complexes, crucial for sustained oscillations, before they are both lost from the KaiC complex. Output from the PTO proceeds through the kinase SasA and (via KaiB) through the phosphatase CikA; among the loci controlled is the KaiBC operon, thereby constituting a TTFL that supplies KaiB and KaiC. See text and Box 1 for details. B. Fungal and Animal Clocks. TTFLs comprising heterodimeric activators (blue ovals) and common parts of negative elements complexes (red ovals, deliberately simplified) are the core circadian oscillators. Their generalized actions within the clock and in driving output are shown below: Positive Elements (PEs) drive expression of the gene(s) encoding the Negative Elements (NEs). After translation these NE proteins are slowly post-translationally modified, eventually reaching a state where they become effective in forming a repressive complex that depresses the expression of the own gene(s). Although additional post-translational modification of the NEs leads to their degradation this does not contribute to the timing mechanism. The daily cycle of repression and de-repression of the PEs creates a means for output from the clock when the PEs drive expression of clock-controlled genes. Stars represent phosphorylations; the pink oval represents inactivated/degrading complex. See text and Box 2 for details.

Figure 2

Recent advancements in elucidating negative arm function. A. Negative arm noncoding antisense RNA creates a double negative feedback loop via either ribosome collision or creating a transcriptionally permissive state. B. Stability of an Intrinsically Disordered Protein (IDP) in the negative arm (red oval) is enhanced by its partner protein (grey moon), stabilizing the IDP until it is able to impart its function in the clock. C. Negative arm repression is conferred, in part, by negative arm proteins acting as a scaffold to bring into contact transcriptional repressors (green elements), whose components may be split between the positive arm and negative arm. D. Degradation of the negative arm proteins is not essential to release the clock from a repressive state. Likely, clock-specific phosphorylation regulates negative arm activity and degradation-specific phosphorylation regulates degradation of the negative arm half-life. Blue ovals represent the positive arm complexes, red ovals represent the negative arm complexes, stars represent phosphorylations; the pink oval represents inactivated/degrading complex, green shapes represent transcriptional repressors, orange ovals represent ribosomes, grey moons represent Nanny proteins.

Figure 3

Current models of Positive Arm operation. A. The heterodimeric transcription factor complex of the Positive Arm in fungi and mammals has been shown to act as a Pioneer transcription factor (TF), in some cases allowing secondary TFs to set the phase of ccgs. B. Post transcriptional mechanisms regulate the output of the clock, making it possible for genes that are rhythmically activated to be arrhythmic at the steady state mRNA level, as well as genes that are not rhythmically activated to be rhythmic at the mRNA steady state level.