Characterization of the human renal Na+-sulphate cotransporter gene (NAS1) promoter

Abstract

Sulphate (SO(4)(2-)) plays an essential role during growth, development, and cellular metabolism. Recently, we have isolated the human renal Na(+)-SO(4)(2-) cotransporter (hNaSi-1) that is implicated in the regulation of serum SO(4)(2-) levels. To gain an insight into hNaSi-1 regulation, our aims were to clone and characterize functionally the hNaSi-1 gene ( NAS1) promoter. We PCR-amplified 3742 bp of the NAS1 5'-flanking region, which is 64% AT-rich and contains numerous putative cis-acting elements. The NAS1 transcription start site was mapped to 25 bp upstream from the translation start site. NAS1 promoter truncations fused to luciferase gene constructs transfected into renal LLC-PK1, MDCK and OK cells allowed us to establish that the first 169 bp of the NAS1 promoter are sufficient for basal transcription. Furthermore, the NAS1 promoter conferred responsiveness to the polycyclic aromatic hydrocarbon 3-methylcholanthrene (3-MC), but not to thyroid hormone (T(3)) or vitamin D [1,25-(OH)(2)D(3)]. Site-directed mutagenesis of the NAS1 promoter identified a functional xenobiotic response element at -2,052, which conferred 3-MC responsiveness. The human NAS1 gene promoter is not responsive to Vitamin D or T(3), unlike the mouse Nas1 promoter with which it shares approximately 40% sequence similarity, but is transactivated by 3-MC, suggesting that the control of renal SO(4)(2-) reabsorption via the regulation of NAS1 transcription may be important for maintaining the sulphation potential for kidney polycyclic aromatic hydrocarbon metabolism.

Physiological Roles of Mammalian Sulfate Transporters NaS1 and Sat1

“…NaS1 promoter studies showed up-regulation by vitamin D, thyroid hormone and the xenobiotic 3-methycholanthrene (Dawson and Markovich 2002;Lee et al 2005;Lee and Markovich 2004) and down-regulated by glucocorticoids (Beck and Markovich 2000).…”

Structure, organization and tissue expression of the pig SLC13A1 and SLC13A4 sulfate transporter genes

“…GATA1, GATA binding factor 1; Oct-1, octamer factor 1; HNF4, hepatic nuclear factor 4; GMEB2, glucocorticoid modulatory element binding protein 2; and GATA3, GATA binding factor 3. (B) Relative locations of each transcription-factor binding site were compared to those previously reported in the 5’-flanking region of human SLC13A1 and mouse Slc13a1 [24] , [25] .

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“…To determine the nucleotide sequences of intron/exon junctions, transcription initiation start sites, and the 5’-flanking regions, we aligned the curated pig SLC13A1 ( XM_013985680.1 ) and SLC13A4 ( XM_003134643.3 ) mRNA sequences with pig genome sequence ( NC_010460.3 ). Putative transcription factor binding motifs within the first 1000 nucleotides of the 5’-flanking regions of pig SLC13A1 and SLC13A4 were identified using MatInspector software [23] , and compared to the published SLC13A1 and SLC13A4 gene promoter findings for human and mouse [16] , [24] , [25] , [26] . Amino acid sequences of pig SLC13A1 ( XP_013841134.1 ) and SLC13A4 ( XP_003134691.1 ) were aligned to human and mouse homologue proteins using ClustalW software [27] .…”

The SLC13 gene family of sodium sulphate/carboxylate cotransporters

“…The mNaS1 gene (Nas1) has been shown to be upregulated significantly by vitamin D and thyroid hormone by binding to the vitamin D 3 -responsive element (VDRE, at 525 bp) and triiodothyronine-responsive element (T 3 RE, at 436) on the Nas1 promoter, respectively [10] and down-regulated by glucocorticoids [27], whereas the hNaS1 gene (Nas1) is up-regulated significantly by the xenobiotic 3-methycholanthrene by binding to a xenobiotic-responsive element (XRE, at 2052 bp) on the Nas1 promoter [23].…”