Role for Protein Kinase A in the Neurospora Circadian Clock by Regulating White Collar-Independent frequency Transcription through Phosphorylation of RCM-1

Abstract

ABSTRACT Rhythmic activation and repression of clock gene expression is essential for the eukaryotic circadian clock functions. In the Neurospora circadian oscillator, the transcription of the frequency ( frq ) gene is periodically activated by the White Collar (WC) complex and suppressed by the FRQ-FRH complex. We previously showed that there is WC-independent frq transcription and its repression is required for circadian gene expression. How WC-independent frq transcription is regulated is not known. We show here that elevated protein kinase A (PKA) activity results in WC-independent frq transcription and the loss of clock function. We identified RCM-1 as the protein partner of RCO-1 and an essential component of the clock through its role in suppressing WC-independent frq transcription. RCM-1 is a phosphoprotein and is a substrate of PKA in vivo and in vitro . Mutation of the PKA-dependent phosphorylation sites on RCM-1 results in WC-independent transcription of frq and impaired clock function. Furthermore, we showed that RCM-1 is associated with the chromatin at the frq locus, a process that is inhibited by PKA. Together, our results demonstrate that PKA regulates frq transcription by inhibiting RCM-1 activity through RCM-1 phosphorylation.