van den Hove Marie-France, Croizet-Berger Karine, Jouret François, Guggino Sandra E, Guggino William B, Devuyst Olivier, Courtoy Pierre J
Cell Biology Unit-ICP/UCL 7541, 75 av Hippocrate, B-1200 Brussels, Belgium.
Endocrinology. 2006 Mar;147(3):1287-96. doi: 10.1210/en.2005-1149. Epub 2005 Nov 23.
Genetic inactivation of ClC-5, a voltage-gated chloride channel prominently expressed in the kidney, leads to proteinuria because of defective apical endocytosis in proximal tubular cells. Because thyroid hormone secretion depends on apical endocytosis of thyroglobulin (Tg), we investigated whether ClC-5 is expressed in the thyroid and affects its function, using Clcn5-deficient knockout (KO) mice. We found that ClC-5 is highly expressed in wild-type mouse thyroid ( approximately 40% of mRNA kidney level). The protein was immunolocalized at the apical pole of thyrocytes. In Percoll gradients, ClC-5 overlapped with plasma membrane and early endosome markers, but best codistributed with the late endosomal marker, Rab7. ClC-5 KO mice were euthyroid (normal T4 and TSH serum levels) but developed a goiter with parallel iodine and Tg accumulation (i.e. normal Tg iodination level). When comparing ClC-5 KO with wild-type mice, thyroid 125I uptake after 1 h was doubled, incorporation into Tg was decreased by approximately 2-fold, so that trichloroacetic acid-soluble 125I increased approximately 4-fold. Enhanced 125I- efflux upon perchlorate and presence of 125I-Tg as autoradiographic rings at follicle periphery demonstrated delayed iodide organification. Endocytic trafficking of 125I-Tg toward lysosomes was not inhibited. Expression of pendrin, an I-/Cl- exchanger involved in apical iodide efflux, was selectively decreased by 60% in KO mice at mRNA and protein levels. Thus, ClC-5 is well expressed in the thyroid but is not critical for apical endocytosis, contrary to the kidney. Instead, the goiter associated with ClC-5 KO results from impaired rate of apical iodide efflux by down-regulation of pendrin expression.
ClC-5是一种在肾脏中显著表达的电压门控氯通道，其基因失活会因近端肾小管细胞顶端内吞作用缺陷而导致蛋白尿。由于甲状腺激素的分泌依赖于甲状腺球蛋白（Tg）的顶端内吞作用，我们使用Clcn5基因敲除（KO）小鼠研究了ClC-5是否在甲状腺中表达并影响其功能。我们发现ClC-5在野生型小鼠甲状腺中高度表达（约为肾脏mRNA水平的40%）。该蛋白在甲状腺细胞的顶端极进行免疫定位。在Percoll梯度中，ClC-5与质膜和早期内体标记物重叠，但与晚期内体标记物Rab7的共分布最佳。ClC-5基因敲除小鼠甲状腺功能正常（血清T4和TSH水平正常），但出现甲状腺肿大，同时伴有碘和Tg蓄积（即Tg碘化水平正常）。将ClC-5基因敲除小鼠与野生型小鼠进行比较时，1小时后甲状腺125I摄取量增加了一倍，Tg中的掺入量减少了约2倍，因此三氯乙酸可溶性125I增加了约4倍。高氯酸盐处理后125I外流增强，且在滤泡周边出现125I-Tg作为放射自显影片环，表明碘化物有机化延迟。125I-Tg向溶酶体的内吞运输未受抑制。参与顶端碘外流的I-/Cl-交换体pendrin的表达在基因敲除小鼠的mRNA和蛋白水平上选择性降低了60%。因此，与肾脏相反，ClC-5在甲状腺中表达良好，但对顶端内吞作用并不关键。相反，与ClC-5基因敲除相关的甲状腺肿大是由于pendrin表达下调导致顶端碘外流速率受损所致。
