A large-scale functional RNAi screen reveals a role for CK2 in the mammalian circadian clock

Area of Science:

• Chronobiology
• Molecular Biology
• Biochemistry

Background:

• Post-translational modifications, particularly phosphorylation, are critical for mammalian circadian rhythm generation and dynamics.
• The precise mechanisms by which phosphorylation of the Period 2 (PER2) protein controls circadian period and phase remain largely unknown.

Purpose of the Study:

• To identify kinases involved in PER2 phosphorylation and their role in the mammalian circadian clock.
• To elucidate the function of casein kinase 2 (CK2) in regulating circadian oscillations.

Main Methods:

• High-throughput RNA interference (RNAi)-based genetic screen to identify PER2-phosphorylating kinases.
• In vivo and in vitro biochemical assays to study CK2-PER2 interactions and phosphorylation.
• Pharmacological inhibition of CK2 activity.
• Analysis of circadian rhythm dynamics upon CK2 subunit silencing or inhibition.

Main Results:

• Casein kinase 2 (CK2) was identified as a novel PER2-phosphorylating kinase and component of the mammalian circadian clock.
• CK2 silencing or inhibition disrupted circadian rhythms.
• CK2 binds to PER2 in vivo, phosphorylates it at N-terminal residues in vitro, and supports nuclear PER2 accumulation.
• Mutation of CK2 phosphorylation sites on PER2 decreased its stability and mimicked the effects of CK2 inhibition on oscillation dynamics.

Conclusions:

• CK2 is a crucial regulator of the mammalian circadian clock through its phosphorylation of PER2.
• PER2 phosphorylation and stabilization by CK2 play a role in setting the clock speed.
• This study proposes a novel mechanism where PER2 phosphorylation and stabilization can modulate clock speed in opposing directions based on the phase of action.