Molecular and functional characterization of adrenomedullin receptors in pufferfish
Molecular and functional characterization of adrenomedullin receptors in pufferfish
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DOI：
10.1152/ajpregu.00507.2005
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发表时间：
2006-02-01
影响因子：
2.8
通讯作者：
Hirose, S
中科院分区：
文献类型：
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作者：
Nag, K;Kato, A;Hirose, S
在暗纹东方鲀（一种与红鳍东方鲀非常接近的广盐性鲀类，其拥有多达五个肾上腺髓质素基因（AM1 - 5））中对降钙素基因相关肽（CGRP）/肾上腺髓质素（AM）家族肽的受体进行了特性分析。CGRP和AM共有一种称为降钙素受体样受体（CLR）的G蛋白偶联核心受体，并且CLR的特异性由与受体活性修饰蛋白（RAMPs）的相互作用所决定。通过数据库挖掘，鉴定出三个CLR（CLR1 - 3）和五个RAMP（RAMP1 - 5），并且通过逆转录聚合酶链反应（RT - PCR）对它们全部进行了克隆，并通过在COS7细胞中以CLR - RAMP的所有可能组合进行功能性表达来对其特性进行分析。以下组合在生理浓度的CGRP、AM1（哺乳动物AM的直系同源物）、AM2和AM5作用下产生环磷腺苷（cAMP）：CLR1 - RAMP1/4（CGRP）、CLR1 - RAMP2/3/5（AM1）、CLR2 - RAMP2（AM1）、CLR1 - RAMP3（AM2）以及CLR1 - RAMP3（AM5）。通过Northern杂交分析发现它们的表达具有组织特异性和盐度依赖性。例如，CLR1 - RAMP5和CLR1 - RAMP2分别在鳃和肾脏中特异性表达，表明它们参与渗透调节。此外，在脾脏和卵巢中发现相对较高水平的CLR和RAMP，表明它们在免疫和雌性生殖系统中起作用。免疫组织化学显示以下类型的AM受体在暗纹东方鲀鳃和肾脏的如下部位（括号内所示）表达：CLR1 - RAMP5（鳃小片间血管）、CLR2 - RAMP2（鳃丝的柱细胞）以及CLR1 - RAMP2（肾近端小管细胞的顶端侧）。
The receptors for the calcitonin gene- related peptide (CGRP)/ adrenomedullin (AM) family peptides were characterized in the mefugu Takifugu obscurus, a euryhaline fugu species very close to Takifugu rubripes, which has as many as five adrenomedullin genes (AM1 - 5). CGRP and AM share a G protein- coupled core receptor called calcitonin receptor- like receptor (CLR), and the specificity of the CLR is determined by the interaction with receptor activity- modifying proteins (RAMPs). Through database mining, three CLRs (CLR1 - 3) and five RAMPs (RAMP1 - 5) were identified, and all of them were cloned by RT- PCR and characterized by functional expression in COS7 cells in every possible combination of CLR- RAMP. The following combinations generated cAMP in response to physiological concentrations of CGRP, AM1 (an ortholog of mammalian AM), AM2, and AM5: CLR1-RAMP1/ 4 (CGRP), CLR1- RAMP2/ 3/ 5 (AM1), CLR2RAMP2 (AM1), CLR1- RAMP3 (AM2), and CLR1- RAMP3 (AM5). Their expressions were found by Northern blot analysis to be tissue specific and salinity dependent. For example, CLR1- RAMP5 and CLR1- RAMP2 are expressed specifically in the gill and kidney, respectively, suggesting their involvement in osmoregulation. Furthermore, relatively high levels of CLRs and RAMPs were found in the spleen and ovary, suggesting roles in the immune and female reproductive systems. Immunohistochemistry revealed that AM receptors of the following types are expressed in the locations, indicated in brackets, of the mefugu gill and kidney: CLR1- RAMP5 (interlamellar vessels), CLR2- RAMP2 (pillar cells), and CLR1- RAMP2 (apical side of renal proximal tubule cells).