Genomic organization and expression of the mouse equilibrative, nitrobenzylthioinosine-sensitive nucleoside transporter 1 (ENT1) gene

We have cloned and characterized the genomic structure of the mouse gene fo r the NBMPR-sensitive equilibrative nucleoside transporter (mENT1), which i s located on chromosome 17C, About 12-kb of genomic DNA was sequenced inclu ding the promoter region, 12 exons, 11 introns, and the 3'-untranslated reg ion. All exon-intron junction sequences conform to the GT/AG rule. Primer e xtension analysis demonstrated a transcription initiation site located 252 bp upstream of the translation start site. Analysis of the 2.5-kb 5'-flanki ng sequence shows putative binding sites for several transcription factors, including GATA-1, IRF-2, Pit-1, myogenin, CREB, Sp-1, Ap-2, MAZ, and GR. W e demonstrated that mouse ENT1 mRNA was highly expressed in heart, spleen, lung, liver, and testis. Lower levels of expression were detected in brain and kidney. Functional analysis of the 5'-flanking region showed that the n ucleotide sequence from -652 to -111 contains cis-regulatory elements that promote gene expression. We found two Sp-1 binding sites (-296/-303, -307/ -313) and two MAZ binding sites (-353/-359, -522/ -528) in this region. Luc iferase assay results suggest that MAZ and Sp-l transcription factors are i mportant positive regulators of transcription for the ENT1 gene in NG108-15 cells. (C) 2000 Academic Press.