Functional properties of leptin receptor isoforms internalization and degradation of leptin and ligand-induced receptor downregulation
Functional properties of leptin receptor isoforms internalization and degradation of leptin and ligand-induced receptor downregulation
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DOI：
10.2337/diabetes.48.2.279
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发表时间：
1999-02-01
期刊：
影响因子：
7.7
通讯作者：
Flier, JS
中科院分区：
文献类型：
--
作者：
Uotani, S;Bjorbæk, C;Flier, JS
瘦素受体长（ObRb）和短（ObRa）亚型被认为分别在介导瘦素信号传导以及瘦素的转运和降解中起重要作用。尽管已经报道了这些克隆受体种类介导信号转导的能力，但没有关于单个受体种类介导瘦素内化和降解或经历配体诱导的下调的能力的信息。因此，我们研究了这些参数在中国仓鼠卵巢（CHO）细胞稳定表达的瘦素受体的ObRa或ObRb亚型。我们确定，ObRa和ObRb介导的I-125标记的瘦素的内化的温度和涂层坑依赖性机制。ObRa和ObRb还通过溶酶体机制介导I-125-瘦素的降解，并且这在这些细胞中由ObRa更有效地介导。既不瘦素内化，也不降解ObRa的保守盒1基序的突变的影响。通过研究ObRa的缺失突变体，我们发现有效的内化依赖于位于ObRa胞内结构域的氨基酸8和29之间的基序。表达ObRa或ObRb的细胞在37 ° C暴露于未标记的瘦素90分钟产生可用的表面受体的下调，并且这种作用在表达ObRb的细胞中更大。而表达生长激素受体的CHO细胞暴露于地塞米松（DEX）或佛波醇肉豆蔻酸（PMA）后显示出明显的配体结合下调，PMA对ObRa或ObRb的表达没有影响，DEX使与表达ObRb的细胞的结合降低了15%。因此，两种瘦素受体亚型ObRa和ObRb通过包被的Pit依赖性机制介导瘦素内化，通过溶酶体途径介导瘦素降解，以及配体诱导的受体下调。两种受体亚型的差异能力可能与受体亚型在瘦素生物学中的不同作用有关。
Long (ObRb) and short (ObRa) leptin receptor isoforms are thought to play essential roles in mediating leptin signaling and the transport and degradation of leptin, respectively. Although the capacity of these cloned receptor species to mediate signal transduction has been reported, there is no information on the ability of individual receptor species to mediate leptin internalization and degradation or to undergo ligand-induced downregulation. We therefore studied these parameters in Chinese hamster ovary (CHO) cells stably expressing either ObRa or ObRb isoforms of the leptin receptor. We determined that both ObRa and ObRb mediated internalization of I-125-labeled leptin by a temperature- and coated pit-dependent mechanism. Both ObRa and ObRb also mediated degradation of I-125-leptin by a lysosomal mechanism, and this was more efficiently mediated by ObRa in these cells. Neither leptin internalization nor degradation by ObRa was affected by mutation of the conserved Box 1 motif. By studying deletion mutants of ObRa, we found that efficient internalization was dependent on a motif located between amino acids 8 and 29 of the intracellular domain of ObRa. Exposure of cells expressing ObRa or ObRb to unlabeled leptin for 90 min at 37 degrees C produced downregulation of available surface receptors, and this effect was of greater magnitude in cells expressing ObRb. Whereas CHO cells expressing the growth hormone receptor showed marked downregulation of ligand binding after exposure to dexamethasone (DEX) or phorbol myristic acid (PMA), PMA had no effect on expression of ObRa or ObRb, and DEX reduced binding to cells expressing ObRb by 15%. Thus, the two leptin receptor isoforms, ObRa and ObRb, mediate leptin internalization by a coated pit-dependent mechanism, leptin degradation by a lysosomal pathway, and ligand-induced receptor downregulation. The differential capacity of the two receptor isoforms may relate to the different roles of the receptor isoforms in the biology of leptin.