Ramamoorthy S, Del Monte M A, Leibach F H, Ganapathy V
Department of Biochemistry and Molecular Biology, Medical College of Georgia, Augusta 30912-2100.
Curr Eye Res. 1994 Jul;13(7):523-9. doi: 10.3109/02713689408999884.
The molecular identity and calmodulin-mediated regulation of the taurine transporter were investigated in a human retinal pigment epithelial cell line (HRPE). Reverse transcription-polymerase chain reaction amplification of HRPE cell mRNA using primer specific for a taurine transporter cloned from human placenta yielded a product of expected size (approximately 0.9 kb) which hybridized to the placental cDNA probe under high stringency conditions. The nucleotide sequence of the product was identical to the sequence of the portion of the placental taurine transporter cDNA flanked by the specific primers. The taurine transporter expressed in the HRPE cell line thus appears to be identical to the transporter cloned from the placenta. Treatment of the HRPE cells with a selective calmodulin antagonist CGS 9343 B (CGS) led to a marked decrease in taurine transport activity. This effect could be reproduced with W-7, another calmodulin antagonist. The inhibition caused by CGS occurred rapidly (t1/2 approximately 10 min). Treatment of the cells with CGS did not affect the transport of leucine, and amino acid not recognized by the taurine transporter as a substrate. The CGS-induced inhibition of taurine transport was accompanied by a decrease in the maximal velocity of the transporter with no detectable change in the substrate affinity. The steady state levels of the transporter mRNA however remained unaffected by CGS treatment. It is concluded that the HRPE cell line expressed a taurine transporter identical to the transporter describe in the human placenta and that the function of this transporter is regulated by calmodulin-dependent processes.
在人视网膜色素上皮细胞系（HRPE）中研究了牛磺酸转运体的分子特性及钙调蛋白介导的调节作用。使用从人胎盘中克隆的牛磺酸转运体特异性引物对HRPE细胞mRNA进行逆转录-聚合酶链反应扩增，得到预期大小（约0.9 kb）的产物，该产物在高严格条件下与胎盘cDNA探针杂交。产物的核苷酸序列与特异性引物侧翼的胎盘牛磺酸转运体cDNA部分的序列相同。因此，HRPE细胞系中表达的牛磺酸转运体似乎与从胎盘中克隆的转运体相同。用选择性钙调蛋白拮抗剂CGS 9343 B（CGS）处理HRPE细胞导致牛磺酸转运活性显著降低。用另一种钙调蛋白拮抗剂W-7也可重现这种效应。CGS引起的抑制作用迅速发生（t1/2约为10分钟）。用CGS处理细胞不影响亮氨酸的转运，亮氨酸是牛磺酸转运体不识别为底物的一种氨基酸。CGS诱导的牛磺酸转运抑制伴随着转运体最大速度的降低，而底物亲和力没有可检测到的变化。然而，转运体mRNA的稳态水平不受CGS处理的影响。结论是，HRPE细胞系表达的牛磺酸转运体与在人胎盘中描述的转运体相同，并且该转运体的功能受钙调蛋白依赖性过程的调节。
