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Am J Physiol Renal Physiol 282: F630-F638, 2002. First published November 20, 2001; doi:10.1152/ajprenal.00264.2001
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Vol. 282, Issue 4, F630-F638, April 2002

Structure and characterization of the mouse UT-A gene (Slc14a2)

R. A. Fenton1, C. A. Cottingham1, G. S. Stewart1, A. Howorth1, J. A. Hewitt2, and C. P. Smith1

1 School of Biological Sciences, University of Manchester, Manchester M13 9PT; and 2 Institute of Genetics, Queen's Medical Centre, Nottingham University, Nottingham NG7 2UH, United Kingdom

The movement of urea across plasma membranes is modulated by facilitated urea transporter proteins. These proteins are the products of two closely related genes, termed UT-A (Slc14a2) and UT-B (Slc14a1). By genomic library screening and P1 artificial chromosome "shotgun" sequencing, we have determined the structure of the mouse UT-A gene. The gene is >300 kb in length, contains 24 exons, and has 2 distinct promoters. Flanking the 5'-region of the gene is the UT-Aalpha promoter that regulates transcription of UT-A1 and UT-A3. The second promoter, termed UT-Abeta , is present in intron 13 and regulates transcription of UT-A2. cAMP agonists (100 µM dibutryl cAMP, 25 µM forskolin, 0.5 mM IBMX) increased the activity of a 2.2-kb UT-Aalpha promoter construct 6.2-fold [from 0.026 ± 0.003 to 0.160 ± 0.004, relative light units (RLU)/µg protein] and a 2.4-kb UT-Abeta promoter construct 9.5-fold (from 0.020 ± 0.002 to 0.190 ± 0.043 RLU/µg protein) above that in untreated controls. Interestingly, only the UT-Abeta promoter contained consensus sequences for CREs and deletion of these elements abolished cAMP sensitivity. Increasing the tonicity of culture medium from 300 to 600 mosmol/kgH2O with NaCl caused a significant increase (from 0.060 ± 0.004 to 0.095 ± 0.010 RLU/µg protein) in UT-Aalpha promoter activity but had no effect on the UT-Abeta promoter. A tonicity-responsive enhancer was identified in UT-Aalpha and is suggested to be responsible for mediating this effect. Levels of UT-A2 and UT-A3 mRNA were increased in thirsted mice compared with control animals, indicating that the activities of both promoters are likely to be elevated during prolonged antidiuresis.




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