The cellular reactive oxygen species (ROS) mediate stability and amplification of circadian rhythm

生物钟是由许多反馈回路组成的。丝状真菌粗毛脉孢霉有一个主要的反馈环，由频率(FRQ)/白领(WC)-1和-2振荡器(FWO)和假定的无FRQ振荡器(FLO)组成，如WC依赖的Flo(WC-Flo)。细胞内的ROS被认为与生物钟有关。然而，这种参与背后的机制仍不清楚。在这里，我们显示了细胞内ROS浓度和ROS破坏过氧化氢酶(CAT)活性以昼夜节律的方式振荡。ROS的幅度随着周期的延长而增加，此后保持不变，而细胞CAT活性的幅度则下降。细胞内的ROS是由NADPH氧化酶(NOX)产生的，并通过使用CAT的FLOS振荡，但其幅度受FWO的调节。CAT-1的表达在没有FRQ的情况下振荡，但在WC-1或WC-2中不振荡。此外，遗传和生化证据表明，较高的ROS浓度促进了WCC的转录功能，并增强了分生孢子带。由WCC和CAT-1通过ROS构成的反馈回路可能是WC-Flo的候选方案。这些发现表明，细胞内的RO连接了FWO和FLOO等振荡器，使它们能够进行交流并稳定昼夜节律。

Circadian clocks are composed of a number of feedback loops. The filamentous fungus Neurospora crassa has a major feedback loop that consists of the FREQUENCY (FRQ)/WHITE COLLAR (WC) -1 and -2 oscillator (FWO) and presumptive FRQ-less oscillators (FLOs) such as WC-dependent FLO (WC-FLO). Intracellular ROS are thought to be involved in the circadian clocks. However, the mechanisms underlying this involvement remain unknown. Here, we show that cellular ROS concentrations and the ROS destroying catalases (CATs) activities oscillate in a circadian manner. The amplitudes of ROS increase with each cycle and remain steady thereafter, while those of cellular CAT activity decrease. The cellular ROS are generated by NADPH oxidase (NOX) and oscillate via FLOs using CATs, but their amplitudes are regulated by FWO. cat-1 expression oscillates in the absence of frq but does not in wc-1 or wc-2. Furthermore, genetic and biochemical evidence reveals that a higher ROS concentration promotes the WCC's transcriptional function and enhances conidiation banding. The feedback loop composed of the WCC and CAT-1 via ROS may be a candidate for the WC-FLO. These findings suggest that cellular ROS connect oscillators such as the FWO and FLOs, allowing them to communicate and stabilize the circadian rhythm.