CikA, an Input Pathway Component, Senses the Oxidized Quinone Signal to Generate Phase Delays in the Cyanobacterial Circadian Clock

Abstract

The circadian clock is a timekeeping system in most organisms that keeps track of the time of day. The rhythm generated by the circadian oscillator must be constantly synchronized with the environmental day/night cycle to make the timekeeping system truly advantageous. In the cyanobacterial circadian clock, quinone is a biological signaling molecule used for entraining and fine-tuning the oscillator, a process in which the external signals are transduced into biological metabolites that adjust the phase of the circadian oscillation. Among the clock proteins, the pseudo-receiver domain of KaiA and CikA can sense external cues by detecting the oxidation state of quinone, a metabolite that reflects the light/dark cycle, although the molecular mechanism is not fully understood. Here, we show the antagonistic phase shifts produced by the quinone sensing of KaiA and CikA. We introduced a new cyanobacterial circadian clock mixture that includes an input component in vitro. KaiA and CikA cause phase advances and delays, respectively, in this circadian clock mixture in response to the quinone signal. In the entrainment process, oxidized quinone modulates the functions of KaiA and CikA, which dominate alternatively at day and night in the cell. This in turn changes the phosphorylation state of KaiC—the central oscillator in cyanobacteria—ensuring full synchronization of the circadian clock. Moreover, we reemphasize the mechanistic input functionality of CikA, contrary to other reports that focus only on its output action.

Synchronization of the circadian clock to the environment tracked in real time

“…CikA’s name ( c ircadian i nput k inase) implies its important role in transmitting environmental signals to the clock. More recent work found that including CikA in the IVO reaction can enable delays in the KaiC phosphorylation peak when quinone is added at certain circadian times (CTs), while only phase advances were observed without CikA, consistent with a role in input ( 38 ). CikA and SasA both act in output pathways where they regulate the activity of the transcription factor, and additionally affect the period of the core oscillator and have features of oscillator components ( 23 , 33 , 39 ).…”

Shift in Conformational Equilibrium Underlies the Oscillatory Phosphoryl Transfer Reaction in the Circadian Clock

“…Recently, we proposed that a KaiC-alone hourglass can oscillate under the alternating Mg 2+ concentrations in vitro, and that this perhaps put the emergence of Mg 2+ homeostasis after that of the primeval timekeeper on the clock’s evolutionary timeline [ 14 ]. Since the endogenous Mg 2+ concentration was reported to be constant in S. elongatus [ 43 ], the emergence of a damped oscillator as a better timekeeping system than the cue-dependent hourglass fits well in the picture [ 39 ]. After going through multiple evolutionary steps, the current self-sustained circadian oscillator may have emerged as a biologically efficient way to exploit the periodic availability of solar energy.…”

“…KaiB binding‘s allosteric induction of A-loop burial also had a potential role in the entrainment process since maintaining the A-loop’s buried conformation during the falling phase of the clock was crucial not only for maintaining the KaiC–KaiB–CikA complex, but also because KaiA could not tether the A-loop in its exposed conformation and run the rhythm backward toward phosphorylation. If the A-loop became exposed at this stage, KaiA, not CikA, would have been inactivated by oxidized quinone, which would have advanced the phase instead of delaying it [ 43 , 45 , 46 ]. It is through this process that KaiB initiated the dephosphorylation phase at the correct timing of day and ensured that the CikA binding took place accordingly.…”

The Circadian Clock—A Molecular Tool for Survival in Cyanobacteria

“…CikA also has a PsR domain, which resembles that of KaiA [ 45 ], and oxidized quinone inactivates CikA in a similar manner. Therefore, now CikA and oxidized quinone are added into the standard KaiABC in vitro mixture in order to test whether CikA contributes to a phase delay in response to quinone [ 46 ]. During KaiC’s nighttime dephosphorylation phase, KaiA and CikA compete for the same binding site on KaiB.…”

“…Previously, the standard KaiABC oscillator mixture has been shown to be relatively immune in the biologically meaningful range of temperature between 25 °C and 35 °C, with a Q 10 value close to 1.0 [ 13 ]. In a similar manner, quinone entrainment with the addition of CikA into the mixture also turned out to compensate for temperature [ 46 ].…”