Gαi3 signaling is associated with sexual dimorphic expression of the clock-controlled output gene Dbp in murine liver.

Madhurendra Singh,Fabian Kuck,Hans Reinke,Katja Pexa,Roland P Piekorz,Antje Lindecke,Dennis Stibane,Laura Bergmann,Karl Köhrer,Hakima Ezzahoini,Charlotte Von Gall,Linda Janke,Constanze Wiek,Helmut Hanenberg,Alexander Lang

doi:10.18632/oncotarget.25727

Abstract

The albumin D-box binding protein (DBP) is a member of the PAR bZip (proline and acidic amino acid-rich basic leucine zipper) transcription factor family and functions as important regulator of circadian core and output gene expression. Gene expression of DBP itself is under the control of E-box-dependent binding by the Bmal1-Clock heterodimer and CRE-dependent binding by the cAMP responsive element binding protein (CREB). However, the signaling mechanism mediating CREB-dependent regulation of DBP expression in the peripheral clock remains elusive. In this study, we examined the role of the GPCR (G-protein-coupled receptor)/Gαi3 (Galphai3) controlled cAMP-CREB signaling pathway in the regulation of hepatic expression of core clock and clock-regulated genes, including Dbp. Analysis of circadian gene expression revealed that rhythmicity of hepatic transcript levels of the majority of core clock (including Per1) and clock-regulated genes were not affected by Gαi3 deficiency. Consistently, the period length of primary Gαi3 deficient tail fibroblasts expressing a Bmal1-Luciferase reporter was not affected. Interestingly, however, Gαi3 deficient female but not male mice showed a tendentiously increased activation of CREB (nuclear pSer133-CREB) accompanied by an advanced peak in Dbp gene expression and elevated mRNA levels of the cytochrome P450 family member Cyp3a11, a target gene of DBP. Accordingly, selective inhibition of CREB led to a strongly decreased expression of DBP and CYP3A4 (human Cyp3a11 homologue) in HepG2 liver cells. In summary, our data suggest that the Gαi3-pCREB signalling pathway functions as a regulator of sexual-dimorphic expression of DBP and its xenobiotic target enzymes Cyp3a11/CYP3A4.

Highlights

The circadian clock system in mammals is hierarchically organized comprising a central master clock and peripheral clocks [1, 2]
The cAMP-cAMP responsive element binding protein (CREB) signaling pathway, that is under the control of heterotrimeric Gs and heterotrimeric Gαiβγ protein (Gi) proteins, regulates the expression of core clock and clock-regulated genes, including Per1 and D-box binding protein (DBP) [16, 17]
This study provides novel insight into the role of the heterotrimeric G protein Gαi3 as upstream regulator of the cAMP-pCREB signalling pathway in rhythmic gene expression in the liver

Summary

Introduction

The circadian clock system in mammals is hierarchically organized comprising a central master clock and peripheral clocks [1, 2]. Core molecular clockwork components encoded by socalled clock genes regulate positively (Clock, Bmal1) or negatively (Cry1/Cry and Per1/Per2) rhythmic expression of their target genes (i.e., clock output genes) through transcriptional control via so-called E-box elements [11]. Important clock output genes include members of the family of PAR-domain basic leucine zipper (PAR bZip) transcription factors, i.e. hepatic leukemia factor (HLF), thyrotrophic embryonic factor (TEF), and albumin D-box binding protein (DBP) [12]. The latter activates gene expression of Per1/2 via DBP binding sites (socalled D-box elements) modulating the levels of core clock gene products [13, 14]. PAR bZip proteins control the circadian expression of hepatic enzymes and regulators involved in endobiotic and xenobiotic biotransformation and drug metabolism [15], including isoenzymes of the cytochrome P450 family of monooxygenases

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Conclusion