USP2 regulates the intracellular localization of PER1 and circadian gene expression.

Endogenous 24-h rhythms in physiology are driven by a network of circadian clocks located in most tissues. The molecular clock mechanism is based on feedback loops involving clock genes and their protein products. Posttranslational modifications, including ubiquitination, are important for regulating the clock feedback mechanism. Recently, we showed that the deubiquitinating enzyme ubiquitin-specific peptidase 2 (USP2) associates with clock proteins and deubiquitinates PERIOD1 (PER1) but does not affect its overall stability. Mice devoid of USP2 display defects in clock function. Here, we show that USP2 regulates nucleocytoplasmic shuttling and nuclear retention of PER1 and its repressive role on the clock transcription factors CLOCK and BMAL1. The rhythm of nuclear entry of PER1 in Usp2 knockout mouse embryonic fibroblasts (MEFs) was advanced but with reduced nuclear accumulation of PER1. Although Per1 mRNA expression rhythm remained intact in the Usp2 KO MEFs, the expression profiles of other core clock genes were altered. This was also true for the expression of clock-controlled genes (e.g., Dbp, Tef, Hlf, E4bp4). A similar phase advance of PER1 nuclear localization rhythm and alteration of clock gene expression profiles were also observed in livers of Usp2 KO mice. Taken together, our results demonstrate a novel function of USP2 in the molecular clock in which it regulates PER1 function by gating its nuclear entry and accumulation.