USP2-45 Is a Circadian Clock Output Effector Regulating Calcium Absorption at the Post-Translational Level

PLoS One. 2016 Jan 12;11(1):e0145155. doi: 10.1371/journal.pone.0145155. eCollection 2016.

Abstract

The mammalian circadian clock influences most aspects of physiology and behavior through the transcriptional control of a wide variety of genes, mostly in a tissue-specific manner. About 20 clock-controlled genes (CCGs) oscillate in virtually all mammalian tissues and are generally considered as core clock components. One of them is Ubiquitin-Specific Protease 2 (Usp2), whose status remains controversial, as it may be a cogwheel regulating the stability or activity of core cogwheels or an output effector. We report here that Usp2 is a clock output effector related to bodily Ca2+ homeostasis, a feature that is conserved across evolution. Drosophila with a whole-body knockdown of the orthologue of Usp2, CG14619 (dUsp2-kd), predominantly die during pupation but are rescued by dietary Ca2+ supplementation. Usp2-KO mice show hyperabsorption of dietary Ca2+ in small intestine, likely due to strong overexpression of the membrane scaffold protein NHERF4, a regulator of the Ca2+ channel TRPV6 mediating dietary Ca2+ uptake. In this tissue, USP2-45 is found in membrane fractions and negatively regulates NHERF4 protein abundance in a rhythmic manner at the protein level. In clock mutant animals (Cry1/Cry2-dKO), rhythmic USP2-45 expression is lost, as well as the one of NHERF4, confirming the inverse relationship between USP2-45 and NHERF4 protein levels. Finally, USP2-45 interacts in vitro with NHERF4 and endogenous Clathrin Heavy Chain. Taken together these data prompt us to define USP2-45 as the first clock output effector acting at the post-translational level at cell membranes and possibly regulating membrane permeability of Ca2+.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Absorption, Physiological*
  • Animals
  • Calcium / metabolism*
  • Circadian Clocks*
  • Clathrin Heavy Chains / metabolism
  • Cryptochromes / metabolism
  • Drosophila melanogaster / metabolism
  • HEK293 Cells
  • Homeostasis
  • Humans
  • Hypercalciuria / metabolism
  • Intestinal Mucosa / metabolism
  • Locomotion
  • Male
  • Membranes / metabolism
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Models, Biological
  • Phosphoproteins / metabolism
  • Protein Binding
  • Protein Processing, Post-Translational*
  • Sodium-Hydrogen Exchangers / metabolism
  • Ubiquitin Thiolesterase
  • Ubiquitin-Specific Proteases / metabolism*
  • Up-Regulation

Substances

  • Cryptochromes
  • Phosphoproteins
  • Sodium-Hydrogen Exchangers
  • sodium-hydrogen exchanger regulatory factor
  • Clathrin Heavy Chains
  • Ubiquitin Thiolesterase
  • Ubiquitin-Specific Proteases
  • Usp2 protein, mouse
  • Calcium

Grant support

This work was supported by: Swiss National Science Foundation: www.snf.ch, Grant 310030_141013/1 to OS, Grant 31003A_138350 to VLK, Grant 31003A_130387 to EN; Grant NCCR Kidney.ch; the European Research Council: http://erc.europa.eu/, ERC-2010-StG-260988 to FG, ERC-StG-311194 to EN; and COST Action ADMIRE BM1301 to OS. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.