In mammals, circadian rhythms are generated by delayed negative feedback, in which PERIOD (PER1-3) and CRYPTOCHROME (CRY1, 2) proteins gradually accumulate in the nucleus to suppress the transcription of their own genes. Although the importance of nuclear import and export signals for subcellular localization of clock proteins is well established, little is known about the dynamics of these processes as well as their importance for circadian rhythm generation. We show by pharmacological perturbations of oscillating cells that nuclear import and export are of critical importance for circadian period. Live-cell fluorescence microscopy revealed that nuclear import of the key circadian protein PER2 is fast and further accelerated by CRY1. Moreover, PER2 nuclear import is critically dependent on a specific nuclear receptor binding motif in PER2 that also mediates nuclear immobility. Nuclear export, however, is relatively slow supporting a model of PER2 nuclear accumulation by rapid import, slow export and substantial nuclear degradation.