Existence of leptin receptor protein in chicken tissues: Isolation of a monoclonal antibody against chicken leptin receptor
Existence of leptin receptor protein in chicken tissues: Isolation of a monoclonal antibody against chicken leptin receptor
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DOI：
10.1016/j.ygcen.2007.01.021
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发表时间：
2007-05-01
影响因子：
2.7
通讯作者：
Ito, Y.
中科院分区：
文献类型：
--
作者：
Ohkubo, T.;Nishio, M.;Ito, Y.
瘦素受体属于I类细胞因子受体超家族，在哺乳动物体内具有多种生理功能。然而，瘦素系统在鸟类中知之甚少，因为鸟类中存在受体的天然配体的证据是有争议的。作为揭示瘦素在鸟类中的生理学意义的策略的一部分，我们分离了针对鸡瘦素受体（chLEPR）的单克隆抗体（mAb）。基于chLEPR的cDNA序列，在大肠杆菌中表达编码chLEPR胞质结构域的肽，并将其用于免疫小鼠以获得mAb。使用从用chLEPR表达载体转染的COS-7细胞制备的细胞提取物，通过Western印迹分析，抗chLEPR mAb识别在约180 kDa处迁移的蛋白质。通过Western blot分析，使用相同的单克隆抗体，免疫反应带迁移在180 kDa的鸡脑和来亨鸡雄性肝癌（LMH）细胞中检测到，这是在体外转染研究中观察到的大小相似。总之，在本研究中获得的chLEPR mAb交叉反应，至少，与长亚型chLEPR，表明LEPR mRNA表达在鸡组织中可能被翻译。chLEPR单克隆抗体，这还没有在其他地方描述，使我们能够探索受体在鸡组织中的蛋白质水平上的表达和定位。因此，该抗体将是研究和了解鸟类瘦素及其受体的调控和功能的有力工具。(c)2007爱思唯尔公司All rights reserved.
Leptin receptor belongs to the class I cytokine receptor superfamily, which mediates multiple physiological roles in mammals. However, the leptin system is poorly understood in birds, as the evidence for the existence of a natural ligand of the receptor in birds is controversial. As part of a strategy to reveal the physiological significance of leptin in birds, we isolated a monoclonal antibody (mAb) against a chicken leptin receptor (chLEPR). Based on the cDNA sequence for chLEPR, a peptide coding for the cytoplasmic domain of chLEPR was expressed in Escherichia coli and this was used to immunize mice to obtain the mAb. The anti-chLEPR mAb recognized proteins migrated at approximately 180 kDa by Western blot analysis using cellular extracts prepared from COS-7 cells transfected with chLEPR expression vector. By Western blot analysis using the same mAb, an immunoreactive band migrated at 180 kDa was detected in the chicken brain and Leghorn male hepatoma (LMH) cells, and which was similar to the size observed in the in vitro transfection study. Taken together, the chLEPR mAb obtained in the present study cross-reacted, at least, with long isoform chLEPR, suggesting that LEPR mRNA expressed in chicken tissues is likely to be translated. The chLEPR mAb, which has not been described elsewhere, enables us to explore the expression and localization of the receptor in the chicken tissues at the protein level. Therefore, this antibody would be a powerful tool in studying and understanding the regulation and function of leptin and its receptors in birds. (c) 2007 Elsevier Inc. All rights reserved.