The Phosphorylation of CREB at Serine 133 Is a Key Event for Circadian Clock Timing and Entrainment in the...

Within the suprachiasmatic nucleus (SCN)-the locus of the master circadian clock- transcriptional regulation via the CREB/CRE pathway is implicated in the functioning of the molecular clock timing process, and is a key conduit through which photic input entrains the oscillator. One event driving CRE-mediated transcription is the phosphorylation of CREB at serine 133 (Ser¹³³). Indeed, numerous reporter gene assays have shown that an alanine point mutation in Ser¹³³ reduces CREB-mediated transcription. Here, we sought to examine the contribution of Ser¹³³ phosphorylation to the functional role of CREB in SCN clock physiology in vivo. To this end, we used a CREB knock-in mouse strain, in which Ser¹³³ was mutated to alanine (S/A CREB). Under a standard 12 h light-dark cycle, S/A CREB mice exhibited a marked alteration in clock-regulated wheel running activity. Relative to WT mice, S/A CREB mice had highly fragmented bouts of locomotor activity during the night phase, elevated daytime activity, and a delayed phase angle of entrainment. Further, under free-running conditions, S/A CREB mice had a significantly longer tau than WT mice and reduced activity amplitude. In S/A CREB mice, light-evoked clock entrainment, using both Aschoff type 1 and 6 h "jet lag" paradigms, was markedly reduced relative to WT mice. S/A CREB mice exhibited attenuated transcriptional drive, as assessed by examining both clock-gated and light-evoked gene expression. Finally, SCN slice culture imaging detected a marked disruption in cellular clock phase synchrony following a phase-resetting stimulus in S/A CREB mice. Together, these data indicate that signaling through CREB phosphorylation at Ser¹³³ is critical for the functional fidelity of both SCN timing and entrainment.