Abstract
Aquaporin-5 (AQP5) is a water channel protein that is selectively expressed in respiratory, salivary, and lacrimal tissues. In order to establish the tissue-specific transcriptional programs that underlie its lung- and salivary-specific expression, a 4.5-kilobase pair DNA fragment encompassing the 5'-flanking region of the rat AQP5 gene has been characterized in detail. A major transcription start site utilized in lung and salivary glands has been localized downstream of a TATAA-like motif. Transient transfection assays of -4.3- and -1.7-AQP5-luciferase constructs in AQP5-expressing lung (MLE-15) and salivary (Pa-4) cells and nonexpressing fibroblast (NIH3T3) and epithelial (HeLa) cells demonstrate preferential transcriptional enhancement of reporter activities in MLE-15 and Pa-4 cells. Transient transfection assays of a series of 5' --> 3' deletion constructs of -4.3-AQP5-luciferase suggest that a common salivary and lung enhancer is located between nucleotides -274 and -139, and a lung-specific enhancer is located between nucleotides -894 and -710. There is one putative lung-specific repressor located in the region of nucleotides -1003/-894 and a common lung and salivary repressor located at nucleotides -503/-385. Moreover, 3' --> 5' deletions up to -171 and -127 base pairs almost abolish transcriptional activation in salivary and lung cells, respectively. Together, our findings indicate that the combination of enhancer/repressor elements within the proximal 5'-flanking region of rat AQP5 gene dictates its restricted expression in both lung and salivary cells.
Highlights
The aquaporins constitute a family of homologous intrinsic membrane proteins that function as highly selective water channels and are strongly expressed in tissues the function of which involves rapid water movement across the cell membrane [1,2,3]
AQP5 steady-state mRNA levels increase as a function of time in cultured alveolar epithelial cells, with the maximal increase occurring between days 3 and 5 (Fig. 2A)
We demonstrate that the increase in AQP5 expression that occurs during transdifferentiation of primary cultured AT2 cells toward the AT1 cell phenotype is likely regulated, at least in part, at the transcriptional level
Summary
The aquaporins constitute a family of homologous intrinsic membrane proteins that function as highly selective water channels and are strongly expressed in tissues the function of which involves rapid water movement across the cell membrane [1,2,3]. Expression of many of the aquaporins is regulated in a highly tissue- and cell-specific manner (4 – 6). The restricted expression of AQP5 in type I pneumocytes in distal lung [11], together with its ability to confer mercury-sensitive osmotic water permeability when expressed in Xenopus oocytes [7], implies a role for AQP5 in transepithelial water movement across the alveolar epithelium. In the context of the role of AQP5 as a differentiation marker for AT1 cells and in conjunction with its putative role in water transport in AT1 cells, establishment of AQP5 transcriptional control mechanisms and characterization of its cis-elements that underlie its cell- and tissue-specific expression were undertaken
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