The non-classical nuclear import carrier Transportin 1 modulates circadian rhythms through its effect on PER1 nuclear localization

PLoS Genet. 2018 Jan 29;14(1):e1007189. doi: 10.1371/journal.pgen.1007189. eCollection 2018 Jan.

Abstract

Circadian clocks are molecular timekeeping mechanisms that allow organisms to anticipate daily changes in their environment. The fundamental cellular basis of these clocks is delayed negative feedback gene regulation with PERIOD and CRYPTOCHROME containing protein complexes as main inhibitory elements. For a correct circadian period, it is essential that such clock protein complexes accumulate in the nucleus in a precisely timed manner, a mechanism that is poorly understood. We performed a systematic RNAi-mediated screen in human cells and identified 15 genes associated with the nucleo-cytoplasmic translocation machinery, whose expression is important for circadian clock dynamics. Among them was Transportin 1 (TNPO1), a non-classical nuclear import carrier, whose knockdown and knockout led to short circadian periods. TNPO1 was found in endogenous clock protein complexes and particularly binds to PER1 regulating its (but not PER2's) nuclear localization. While PER1 is also transported to the nucleus by the classical, Importin β-mediated pathway, TNPO1 depletion slowed down PER1 nuclear import rate as revealed by fluorescence recovery after photobleaching (FRAP) experiments. In addition, we found that TNPO1-mediated nuclear import may constitute a novel input pathway of how cellular redox state signals to the clock, since redox stress increases binding of TNPO1 to PER1 and decreases its nuclear localization. Together, our RNAi screen knocking down import carriers (but also export carriers) results in short and long circadian periods indicating that the regulatory pathways that control the timing of clock protein subcellular localization are far more complex than previously assumed. TNPO1 is one of the novel players essential for normal circadian periods and potentially for redox regulation of the clock.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Active Transport, Cell Nucleus / genetics
  • Cell Nucleus / metabolism*
  • Circadian Rhythm / genetics*
  • HEK293 Cells
  • Humans
  • Period Circadian Proteins / metabolism*
  • Protein Transport / genetics
  • Tumor Cells, Cultured
  • beta Karyopherins / genetics
  • beta Karyopherins / physiology*

Substances

  • PER1 protein, human
  • Period Circadian Proteins
  • TNPO1 protein, human
  • beta Karyopherins

Grant support

This work was funded by the Deutsche Forschungsgemeinschaft (Collaborative Research Center 740; Projects D2 and C3). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.