A Screen To Identify Protein Phosphatases with Roles in Circadian Period, Temperature Compensation and Output in the Neurospora Circadian Clock

Abstract

The circadian clock is a highly conserved evolutionary advantage which allows organisms to anticipate regular changes in daily environmental conditions. Clocks from fungi to mammals rely on a transcription-translation feedback loop (TTFL) mechanism. Phosphorylation is understood to be a critical regulatory step for maintaining the period of the circadian clock and feedback loop closure. The role of kinases in the Neurospora clock has been examined extensively; however, phosphatases have not been systematically interrogated. By re-examining the Neurospora genome using current informatic tools we identified the 30 genes previously identified as encoding protein phosphatases as well as 13 novel genes, and we assessed the function of the core circadian clock in 39 non-essential phosphatases using a real-time luciferase reporter. We observed both period lengthening and shortening effects, which are not restricted to a single phosphatase family or fold. All but one deletion mutant maintained a rhythmic core clock. In addition, we observed a new temperature compensation defect in the previously studied knockout of phosphatase pph-4, the result of nutritional growth conditions.