TRANSCRIPTION OF THE SODIUM MYO-INOSITOL COTRANSPORTER GENE IS REGULATED BY MULTIPLE TONICITY-RESPONSIVE ENHANCERS SPREAD OVER 50 KILOBASE PAIRS IN THE 5'-FLANKING REGION/

The sodium/myo-inositol cotransporter is a plasma membrane protein res ponsible for concentrative cellular accumulation of myo-inositol in a variety of tissues. When cells in kidney and brain are exposed to a hy perosmolar salt condition (hypertonicity) due to the operation of urin ary concentration mechanism and pathological conditions, respectively; they survive the stress of hypertonicity by raising the cellular conc entration of myo-inositol. Transcription of the sodium/myo-inositol co transporter gene is markedly stimulated in response to hypertonicity, leading to an increase in the activity of the cotransporter, which in turn drives the osmoprotective accumulation of myo-inositol. To unders tand the molecular mechanisms by which hypertonicity stimulates transc ription, we analyzed the BI-flanking region of the cotransporter gene for cis-acting regulatory sequences. We identified five tonicity-respo nsive enhancers that are scattered over 50 kilobase pairs. All the enh ancers are variations of the same type of enhancer interacting with th e transcription factor named tonicity-responsive enhancer binding prot ein. In vivo methylation experiments demonstrated that exposure of cel ls to hypertonicity increases the binding of tonicity-responsive enhan cer binding protein to the enhancer sites, indicating that all of thes e enhancers are involved in the transcriptional stimulation. We conclu de that the sodium/myo-inositol cotransporter gene is regulated by a l arge region (similar to 50 kilobase pairs) upstream of the gene.