Our daily rhythms in gene expression and metabolism are driven by the circadian oscillator, a biological timer composed of auto-regulatory transcriptional/translational feedback loops. However, molecular regulation and function of core clock components, in particular Period2 (Per2), are not fully understood. Two reporter mouse lines were previously generated, both expressing PER2:LUC fusion proteins from the endogenous Per2 promoter. Whereas the endogenous Per2 3'-UTR remains intact in Per2:Luc mice, it was replaced by an SV40 poly(A) signal in Per2:LucSV mice. Intriguingly, the latter exhibited significantly enhanced circadian amplitude and peak levels of PER2:LUC protein and bioluminescence. Further analysis identified a miR-24 binding site in the 3'-UTR, suggesting an important role of miR-24 in PER2 translation. Furthermore, robust induction of Per2 and Bmal1 transcript levels were observed in Per2:LucSV mice relative to Per2:Luc, suggesting a positive activation role of PER2 in its own transcription. Consistent with the predominant role of the clock in metabolic regulation, preliminary data also illustrated activation of several metabolic regulators in Per2:LucSV mice. Based on these interesting findings, it is hypothesized that PER2 protein levels are controlled by miR-24, and PER2 plays a positive role in Per2 transcription and circadian metabolic function.
Aim 1. Determine the pivotal role of miR-24 in PER2 translational regulation and mouse circadian behavior. The 3'-UTR miR-24 binding site in the Per2:Luc knock-in vector was mutated, and candidate targeted ES cell clones were obtained following electroporation. Mutant differentiated cells (Aim 1A) and knock-in mice (Aim 1B) will be derived to examine reporter rhythms, molecular clock and circadian behavior.
Aim 2. Delineate the molecular mechanism underlying the positive role of PER2 in Per2 auto- regulation. Based on previous ChIP-seq studies showing Per2 promoter recruitment of both positive (CBP) and negative (REV-ERBs) regulators, molecular studies will be conducted to investigate whether PER2 functions to relieve REV-ERB-dependent Per2 transcriptional repression (Aim 2A), and/or to potentiate CBP- mediated transcriptional activation (Aim 2B).
Aim 3. Determine the molecular function of PER2 in energy metabolism. Genetic disruption of the clock leads to insulin resistance and metabolic deficits. To address the reciprocal hypothesis whether enhanced PER2 and circadian rhythms confers metabolic protection, molecular and physiological studies will be conducted to determine whether insulin signaling, a central regulatory pathway for energy metabolism, is activated in Per2:LucSV mice (Aim 3A), and whether these mice are resistant to high-fat diet induced circadian and metabolic abnormalities (Aim 3B).

Public Health Relevance

The circadian clock plays a fundamental role in orchestrating daily rhythms in metabolism and physiology, thus safeguarding our well-being. Period2 (Per2) is a central circadian factor, but its regulation and function are not fully understood. The proposed mechanistic studies will reveal important molecular insights into the regulation and function of Per2 in the clock and energy metabolism, which will lead to better design of preventive and therapeutic strategies against clock-related metabolic diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM114424-02
Application #
9038374
Study Section
Cellular Signaling and Regulatory Systems Study Section (CSRS)
Program Officer
Sesma, Michael A
Project Start
2015-04-01
Project End
2020-03-31
Budget Start
2016-04-01
Budget End
2017-03-31
Support Year
2
Fiscal Year
2016
Total Cost
Indirect Cost
Name
University of Texas Health Science Center Houston
Department
Biochemistry
Type
Schools of Medicine
DUNS #
800771594
City
Houston
State
TX
Country
United States
Zip Code
77225
Chen, Zheng; Yoo, Seung-Hee; Takahashi, Joseph S (2018) Development and Therapeutic Potential of Small-Molecule Modulators of Circadian Systems. Annu Rev Pharmacol Toxicol 58:231-252
Xiang, Yu; Ye, Youqiong; Lou, Yanyan et al. (2018) Comprehensive Characterization of Alternative Polyadenylation in Human Cancer. J Natl Cancer Inst 110:379-389
Ye, Youqiong; Xiang, Yu; Ozguc, Fatma Muge et al. (2018) The Genomic Landscape and Pharmacogenomic Interactions of Clock Genes in Cancer Chronotherapy. Cell Syst 6:314-328.e2
Gloston, Gabrielle F; Yoo, Seung-Hee; Chen, Zheng Jake (2017) Clock-Enhancing Small Molecules and Potential Applications in Chronic Diseases and Aging. Front Neurol 8:100
Zhu, Wenyi; Krishna, Saritha; Garcia, Cristina et al. (2017) Daam2 driven degradation of VHL promotes gliomagenesis. Elife 6:
Nohara, Kazunari; Chen, Zheng; Yoo, Seung-Hee (2017) A Filtration-based Method of Preparing High-quality Nuclei from Cross-linked Skeletal Muscle for Chromatin Immunoprecipitation. J Vis Exp :
Hughes, Michael E; Abruzzi, Katherine C; Allada, Ravi et al. (2017) Guidelines for Genome-Scale Analysis of Biological Rhythms. J Biol Rhythms 32:380-393
Yoo, Seung-Hee; Kojima, Shihoko; Shimomura, Kazuhiro et al. (2017) Period2 3'-UTR and microRNA-24 regulate circadian rhythms by repressing PERIOD2 protein accumulation. Proc Natl Acad Sci U S A 114:E8855-E8864
Jung, Hoe-Yune; Lee, Dongyeop; Ryu, Hye Guk et al. (2017) Myricetin improves endurance capacity and mitochondrial density by activating SIRT1 and PGC-1?. Sci Rep 7:6237
He, Baokun; Nohara, Kazunari; Park, Noheon et al. (2016) The Small Molecule Nobiletin Targets the Molecular Oscillator to Enhance Circadian Rhythms and Protect against Metabolic Syndrome. Cell Metab 23:610-21

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