van den Hove Marie-France, Croizet-Berger Karine, Tyteca Donatienne, Selvais Charlotte, de Diesbach Philippe, Courtoy Pierre J
Université Catholique de Louvain, Christian de Duve Institute of Cellular Pathology, Cell Biology Unit, B-1200 Brussels, Belgium.
J Clin Endocrinol Metab. 2007 Jul;92(7):2803-10. doi: 10.1210/jc.2006-2351. Epub 2007 May 1.
We have previously reported that the TSH receptor/cAMP cascade enhances the coordinate expression of the rate-limiting endocytic catalysts, Rab5a and Rab7, which respectively promote thyroglobulin (Tg) internalization and transfer to lysosomes, thereby accelerating thyroid hormone secretion.
We address whether TSH further controls Rab5a activity by promoting its GTP-bound state.
We compared Rab5a activation in seven pairs of hyperactive and corresponding quiescent thyroid tissues; TSH effect was reproduced on polarized cultures of normal human thyrocytes.
We studied seven euthyroid patients bearing hyperactive autonomous adenomas; normal thyroid tissue for culture.
Rab5a GDP/GTP exchange factor activity [Rab5a-guanine nucleotide exchange factor (GEF)], expression of Rabex-5 (a Rab5a-GEF), and function of thyrocytes in vitro were the main outcome measures.
In autonomous adenomas, constitutive activation increased both total activity and sedimentability (membrane recruitment) of Rab5a-GEF, compared with perinodular tissues. Increased Rab5a-GEF activity correlated with increased expression of Rabex-5 and Rab5a, as well as with Tg store depletion. In polarized human thyrocyte monolayers, TSH did not affect total Rab5a-GEF activity after 2 h but promoted its membrane recruitment; after 4 d, TSH increased both Rab5a-GEF activity and Rabex-5 expression and recruitment onto membranes where Rabex-5 coimmunoprecipitated with Rabaptin-5 and Rab5a. Sedimentable Rab5a-GEF perfectly correlated with apical endocytosis and lysosomal transfer of 125I-Tg, and with basolateral secretion of 125I-derived hormones.
This study provides the first clinical and experimental evidence that regulation of the activity of a rate-limiting endocytic catalyst finely tunes a tightly controlled cellular function that ultimately governs whole body metabolism.
我们之前报道过，促甲状腺激素（TSH）受体/cAMP级联反应可增强限速内吞催化剂Rab5a和Rab7的协同表达，这两种催化剂分别促进甲状腺球蛋白（Tg）的内化并将其转运至溶酶体，从而加速甲状腺激素的分泌。
我们研究TSH是否通过促进Rab5a的GTP结合状态来进一步控制其活性。
我们比较了7对功能亢进及相应静止状态的甲状腺组织中Rab5a的激活情况；并在正常人甲状腺细胞的极化培养中再现TSH的作用。
我们研究了7例患有功能亢进自主性腺瘤的甲状腺功能正常患者；采用正常甲状腺组织进行培养。
Rab5a GDP/GTP交换因子活性[Rab5a-鸟嘌呤核苷酸交换因子（GEF）]、Rabex-5（一种Rab5a-GEF）的表达以及体外甲状腺细胞的功能是主要观察指标。
与结节周围组织相比，在自主性腺瘤中，组成性激活增加了Rab5a-GEF的总活性和沉降性（膜募集）。Rab5a-GEF活性增加与Rabex-5和Rab5a表达增加以及Tg储存耗竭相关。在极化的人甲状腺细胞单层中，TSH在2小时后不影响Rab5a-GEF的总活性，但促进其膜募集；4天后，TSH增加了Rab5a-GEF活性以及Rabex-5的表达和在膜上的募集，在膜上Rabex-5与Rabaptin-5和Rab5a进行共免疫沉淀。可沉降的Rab5a-GEF与125I-Tg的顶端内吞和溶酶体转运以及125I衍生激素的基底外侧分泌完全相关。
本研究提供了首个临床和实验证据，即限速内吞催化剂活性的调节可精细调节一个严格控制的细胞功能，该功能最终控制全身代谢。
