Calcineurin: An essential regulator of sleep revealed by biochemical, chemical biological, and genetic approaches

Research into mechanisms underlying sleep traditionally relies on electrophysiology and genetics. Because sleep can only be measured on whole animals by behavioral observations and physical means, no sleep research was initiated by biochemical and chemical biological approaches. We used phosphorylation sites of kinases important for sleep as targets for biochemical and chemical biological approaches. Sleep was increased in mice carrying a threonine-to-alanine substitution at residue T469 of salt-inducible kinase 3 (SIK3). Our biochemical purification and photo-crosslinking revealed calcineurin (CaN) dephosphorylation, both in vitro and in vivo, of SIK3 at T469 and S551, but not T221. Knocking down CaN regulatory subunit reduced daily sleep by more than 5 h, exceeding all known mouse mutants. Our work uncovered a critical physiological role for CaN in sleep and pioneered biochemical purification and chemical biology as effective approaches to study sleep.