Transcriptional regulation of NHE3 and SGLT1 by the circadian clock protein Per1 in proximal tubule cells.

We have previously demonstrated that the circadian clock protein period (Per)1 coordinately regulates multiple genes involved in Na(+) reabsorption in renal collecting duct cells. Consistent with these results, Per1 knockout mice exhibit dramatically lower blood pressure than wild-type mice. The proximal tubule is responsible for a majority of Na(+) reabsorption. Previous work has demonstrated that expression of Na(+)/H(+) exchanger 3 (NHE3) oscillates with a circadian pattern and Na(+)-glucose cotransporter (SGLT)1 has been demonstrated to be a circadian target in the colon, but whether these target genes are regulated by Per1 has not been investigated in the kidney. The goal of the present study was to determine if Per1 regulates the expression of NHE3, SGLT1, and SGLT2 in the kidney. Pharmacological blockade of nuclear Per1 entry resulted in decreased mRNA expression of SGLT1 and NHE3 but not SGLT2 in the renal cortex of mice. Per1 small interfering RNA and pharmacological blockade of Per1 nuclear entry in human proximal tubule HK-2 cells yielded the same results. Examination of heterogeneous nuclear RNA suggested that the effects of Per1 on NHE3 and SGLT1 expression occurred at the level of transcription. Per1 and the circadian protein CLOCK were detected at promoters of NHE3 and SGLT1. Importantly, both membrane and intracellular protein levels of NHE3 and SGLT1 were decreased after blockade of nuclear Per1 entry. This effect was associated with reduced activity of Na(+)-K(+)-ATPase. These data demonstrate a role for Per1 in the transcriptional regulation of NHE3 and SGLT1 in the kidney.

Small intestinal glucose and sodium absorption through calcium-induced calcium release and store-operated Ca2+ entry mechanisms.

BACKGROUND AND PURPOSE: Luminal glucose enhances intestinal Ca2+ absorption through apical Cav 1.3 channels necessary for GLUT2-mediated glucose absorption. As these reciprocal mechanisms are not well understood, we investigated the regulatory mechanisms of intestinal [Ca2+ ]cyt and SGLT1-mediated Na+ -glucose co-transports. EXPERIMENTAL APPROACH: Glucose absorption and channel expression were examined in mouse upper jejunal epithelium using an Ussing chamber, immunocytochemistry and Ca2+ and Na+ imaging in single intestinal epithelial cells. KEY RESULTS: Glucose induced jejunal Isc via Na+ -glucose cotransporter 1 (SGLT1) operated more efficiently in the presence of extracellular Ca2+ . A crosstalk between luminal Ca2+ entry via plasma Cav 1.3 channels and the ER Ca2+ release through ryanodine receptor (RYR) activation in small intestinal epithelial cell (IEC) or Ca2+ -induced Ca2+ release (CICR) mechanism was involve in Ca2+ -mediated jejunal glucose absorption. The ER Ca2+ release through RyR triggered basolateral Ca2+ entry or store-operated Ca2+ entry (SOCE) mechanism and the subsequent Ca2+ entry via Na+ /Ca2+ exchanger 1 (NCX1) were found to be critical in Na+ -glucose cotransporter-mediated glucose absorption. Blocking RyR, SOCE and NCX1 inhibited glucose induced [Na+ ]cyt and [Ca2+ ]cyt in single IEC and protein expression and co-localization of STIM1/Orai1, RyR1 and NCX1 were detected in IEC and jejunal mucosa. CONCLUSION AND IMPLICATIONS: Luminal Ca2+ influx through Cav 1.3 triggers the CICR through RyR1 to deplete the ER Ca2+ , which induces the basolateral STIM1/Orai1-mediated SOCE mechanism and the subsequent Ca2+ entry via NCX1 to regulate intestinal glucose uptake via Ca2+ signalling. Targeting these mechanisms in IEC may help to modulate blood glucose and sodium in the metabolic disease.

A gut microbial metabolite of ginsenosides, compound K, induces intestinal glucose absorption and Na(+) /glucose cotransporter 1 gene expression through activation of cAMP response element binding protein.

SCOPE: The Na(+) /glucose cotransporter 1 (SGLT1) plays a crucial role in glucose uptake in intestinal epithelial cells (IECs), which has been shown essential in ameliorating intestinal inflammation. Ginseng has historically been used to treat inflammatory disorders. Understanding the regulatory mechanism of ginseng-mediated induction of SGLT1 gene expression in human intestinal cells is therefore important. METHODS AND RESULTS: We demonstrate that ginsenoside compound K (CK) enhances SGLT1-mediated glucose uptake in mice and human intestinal Caco-2 cells. Transient transfection analysis using SGLT1 promoter-luciferase reporters demonstrated that the presence of an essential cAMP response element (CRE) is required for CK-mediated induction of SGLT1 gene expression. The ChIP assays indicated that increased CRE-binding protein (CREB) and CREB-binding protein (CBP) binding to the SGLT1 promoter in CK-treated cells is associated with an activated chromatin state. Our result showed that the increased CREB phosphorylation is directly correlated with SGLT1 expression in IECs. Further studies indicated that the epidermal growth factor receptor (EGFR) signaling pathway is involved in the CK-mediated effect. CONCLUSION: These findings provide a novel mechanism for the CK-mediated upregulation of SGLT1 expression through EGFR-CREB signaling activation, which could contribute to reducing gut inflammation.