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Articles in PresS, published online ahead of print November 20, 2001
Am J Physiol Renal Physiol, 10.1152/ajprenal.00264.2001
Submitted on August 20, 2001
Accepted on December 31, 1969
1 School of Biological Sciences, University of Manchester, Manchester, United Kingdom
2 Queens Medical Centre, Institute of Genetics, University of Nottingham, Nottingham, United Kingdom
* To whom correspondence should be addressed. E-mail: RFENTON{at}MAN.AC.UK.
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 PAC 'shotgun' sequencing, we have determined the structure of the mouse UT-A gene. The gene is greater than 300kb in length, contains 24 exons and has two distinct promoters. Flanking the 5'-region of the gene is the UT-A
promoter that regulates transcription of UT-A1 and UT-A3. The second promoter, termed UT-Aß, is present in intron 13 and regulates transcription of UT-A2. cAMP agonists (100µM dibutryl cAMP, 25µM forskolin, 0.5mM 3-isobutyl-1-methylxanthine) increased the activity of a 2.2kb UT-A promoter construct 6.2-fold (0.026±0.003 to 0.160 ± 0.004, RLU/µg protein) and a 2.4kb UT-Aß promoter construct 9.5-fold (0.020±0.002 to 0.190 ± 0.043) above untreated controls. Interestingly, only the UT-Aß promoter contained consensus sequences for CREs and deletion of these elements abolished cAMP-sensitivity. Increasing the tonicity of culture medium from 300mOsm to 600mOsm with NaCl caused a significant increase (0.060 ± 0.004 to 0.095 ± 0.010) in UT-A promoter activity, but had no effect on the UT-Aß promoter. A TonE response element was identified in UT-A and is suggested to be responsible for mediating this effect. Levels of UT-A2 and UT-A3 mRNA were increased in thirsted mice compared to euhydrated controls; indicating that the activities of both promoters are elevated during prolonged antidiuresis.
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