MOLECULAR CHARACTERIZATION AND IN-SITU LOCALIZATION OF A MOUSE RETINAL TAURINE TRANSPORTER

Various ocular tissues have a higher concentration of taurine than pla sma. This taurine concentration gradient across the cell membrane is m aintained by a high-affinity taurine transporter. To understand the ph ysiological role of the taurine transporter in the retina, we cloned a taurine transporter encoding cDNA from a mouse retinal library, deter mined its biochemical and pharmacological properties, and identified t he specific cellular sites expressing the taurine transporter mRNA. Th e deduced protein sequence of the mouse retinal taurine transporter (m TAUT) revealed >93% sequence identity to the canine kidney, rat brain, mouse brain, and human placental taurine transporters. Our data sugge st that the mTAUT and the mouse brain taurine transporter may be varia nts of one another. The mTAUT synthetic RNA induced Na+- and Cl--depen dent [H-3]taurine transport activity in Xenopus laevis oocytes that sa turated with an average K-m of 13.2 mu M for taurine. Unlike the previ ous studies, we determined the rate of taurine uptake as the external concentration of Cl- was varied, a single saturation process with an a verage apparent equilibrium constant (KCl-) of 17.7 mM. In contrast, t he rate of taurine uptake showed a sigmoidal dependence when the exter nal concentration of Na+ was varied (apparent equilibrium constant, KN a+ similar to 54.8 mM). Analyses of the Na+- and Cl--concentration dep endence data suggest that at least two Na+ and one Cl- are required to transport one taurine molecule via the taurine transporter. Varying t he pH of the transport buffer also affected the rate of taurine uptake ; the rate showed a minimum between pH 6.0 and 6.5 and a maximum betwe en pH 7.5 and 8.0. The taurine transport was inhibited by various inhi bitors tested with the following order of potency: hypotaurine > beta- alanine > L-diaminopropionic acid > guanidinoethane sulfonate > beta-g uanidinopropionic acid > chloroquine > gamma-amino-butyric acid > 3-am ino-1-propanesulfonic acid (homo-taurine). Furthermore, the mTAUT acti vity was not inhibited by the inactive phorbol ester 4 alpha-phorbol 1 2,13-didecanoate but was inhibited significantly by the active phorbol eater phorbol 12-myristate 13-acetate, which was both concentration a nd time dependent. The cellular sites expressing the taurine transport er mRNA in the mouse eye, as determined by in situ hybridization techn ique, showed low levels of expression in many of the ocular tissues, s pecifically the retina and the retinal pigment epithelium. Unexpectedl y, the highest expression levels of taurine transporter mRNA were foun d instead in the ciliary body of the mouse eye.