Abstract
A number of diverse cell-surface proteins are anchored to the cytoskeleton via scaffold proteins. Na+/H+ exchanger regulatory factor-1 (NHERF1), encoded by the Slc9a3r1 gene, functions as a scaffold protein, which is implicated in the regulation of membrane expression of various cell-surface proteins. Here, we demonstrate that the circadian clock component PERIOD2 (PER2) modulates transcription of the mouse Slc9a3r1 gene, generating diurnal accumulation of NHERF1 in the mouse liver. Basal expression of Slc9a3r1 was dependent on transcriptional activation by p65/p50. PER2 bound to p65 protein and prevented p65/p50-mediated transactivation of Slc9a3r1. The time-dependent interaction between PER2 and p65 underlay diurnal oscillation in the hepatic expression of Slc9a3r1/NHERF1. The results of immunoprecipitation experiments and liquid chromatography-mass spectrometry analysis of mouse liver revealed that NHERF1 time-dependently interacted with fatty acid transport protein-5 (FATP5). Temporary accumulation of NHERF1 protein stabilized plasmalemmal localization of FATP5, thereby enhancing hepatic uptake of fatty acids at certain times of the day. Our results suggest an unacknowledged role for PER2 in regulating the diurnal expression of NHERF1 in mouse liver. This machinery also contributed to diurnal changes in the ability of hepatic cells to uptake fatty acids.
Highlights
A number of biological and physiological processes are subjected to diurnal regulation
Since several mechanisms have been suggested in the regulation of the NF-κB signaling pathway by circadian clock genes[19,20], we focused on NF-κB-RE and performed transient transcription assays by constructing Slc9a3r1 luciferase (Luc)-reporter vectors containing the NF-κB-RE of the mouse Slc9a3r1 gene spanning from −200 to +168 bp
The reporter constructs Slc9a3r1(−200/+168)::Luc were co-transfected with expression vectors encoding CLOCK/BMAL1, PER1, PER2, peroxisome proliferator-activated receptor-α (PPARα)/RXRα, RORα, REV-ERBα, D-site binding protein (DBP), or E4BP4
Summary
Received: 25 August 2017 Accepted: 1 June 2018 Published: xx xx xxxx regulatory factor-1 and its scaffolding function. Na+/H+ exchanger regulatory factor-1 (NHERF1), encoded by the Slc9a3r1 gene, functions as a scaffold protein, which is implicated in the regulation of membrane expression of various cell-surface proteins. Our results suggest an unacknowledged role for PER2 in regulating the diurnal expression of NHERF1 in mouse liver This machinery contributed to diurnal changes in the ability of hepatic cells to uptake fatty acids. The transcription of REV-ERBα is activated by CLOCK/BMAL1, and its transactivation is repressed by PER and CRY proteins, resulting in circadian oscillations in the expression of REV-ERBα encoded by the Nr1d1 gene. We found that the expression of Slc9a3r1/NHERF1 was controlled by the molecular component of the circadian clock PER2 This circadian transcriptional repressor time-dependently suppressed the p65/p50-mediated transactivation of Slc9a3r1, thereby generating diurnal accumulation of NHERF1 protein. Our results suggest a previously unacknowledged role of PER2 in regulating the diurnal expression of NHERF1, contributing to diurnal changes in hepatic fatty acid uptake
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