Role of receptor activity modifying protein 1 in function of the calcium sensing receptor in the human TT thyroid carcinoma cell line.
Role of receptor activity modifying protein 1 in function of the calcium sensing receptor in the human TT thyroid carcinoma cell line.
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DOI：
10.1371/journal.pone.0085237
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Skerry TM
中科院分区：
文献类型：
--
作者：
Desai AJ;Roberts DJ;Richards GO;Skerry TM
钙敏感受体（CaSR）通过感受血清Ca 2+的微小变化和调节促钙激素的分泌在钙稳态中起作用。在转染的细胞中已经显示，被称为受体活性修饰蛋白（RAMP）的辅助蛋白，特别是RAMP 1和3，是CaSR细胞表面运输所必需的。这些效应仅在转染细胞中得到证实，因此其生理相关性尚不清楚。在这里，我们详细探讨了CaSR/RAMP相互作用，并表明在甲状腺癌细胞中，RAMP 1是CaSR运输所必需的。此外，我们表明，正常的RAMP 1功能所需的配体的细胞内反应。具体来说，为了证实早期的研究与标记的构建体，并提供定量化学计量分析的额外好处，我们使用荧光共振能量转移显示RAMP 1和3的伴侣蛋白CaSR的细胞表面的能力相同，虽然RAMP 3更有效地与受体相互作用。此外，在细胞表面上的CaSR复合物中观察到RAMP 3比RAMP 1更高的分数，表明RAMPs与CaSR的比例不同。为了确定这些发现在内源性表达系统中的相关性，我们评估了RAMP 1 siRNA敲低在甲状腺髓样癌TT细胞（其组成型表达RAMP 1，但不表达RAMP 3）中的作用，并测量了响应于CaSR配体西那卡塞和新霉素的信号传导的显著50%衰减。使用特异性抗体阻断RAMP 1诱导TT细胞中响应西那卡塞的CaSR介导的信号传导的浓度依赖性降低，这表明RAMP 1除了在受体运输中的已知作用外，还在调节CaSR信号传导中具有新的功能作用。这些数据提供的证据表明，RAMPs交通的CaSR作为更高级别的低聚物，并发挥作用的CaSR信号，即使在细胞表面定位已经发生。
The Calcium Sensing Receptor (CaSR) plays a role in calcium homeostasis by sensing minute changes in serum Ca2+ and modulating secretion of calciotropic hormones. It has been shown in transfected cells that accessory proteins known as Receptor Activity Modifying Proteins (RAMPs), specifically RAMPs 1 and 3, are required for cell-surface trafficking of the CaSR. These effects have only been demonstrated in transfected cells, so their physiological relevance is unclear. Here we explored CaSR/RAMP interactions in detail, and showed that in thyroid human carcinoma cells, RAMP1 is required for trafficking of the CaSR. Furthermore, we show that normal RAMP1 function is required for intracellular responses to ligands. Specifically, to confirm earlier studies with tagged constructs, and to provide the additional benefit of quantitative stoichiometric analysis, we used fluorescence resonance energy transfer to show equal abilities of RAMP1 and 3 to chaperone CaSR to the cell surface, though RAMP3 interacted more efficiently with the receptor. Furthermore, a higher fraction of RAMP3 than RAMP1 was observed in CaSR-complexes on the cell-surface, suggesting different ratios of RAMPs to CaSR. In order to determine relevance of these findings in an endogenous expression system we assessed the effect of RAMP1 siRNA knock-down in medullary thyroid carcinoma TT cells, (which express RAMP1, but not RAMP3 constitutively) and measured a significant 50% attenuation of signalling in response to CaSR ligands Cinacalcet and neomycin. Blockade of RAMP1 using specific antibodies induced a concentration-dependent reduction in CaSR-mediated signalling in response to Cinacalcet in TT cells, suggesting a novel functional role for RAMP1 in regulation of CaSR signalling in addition to its known role in receptor trafficking. These data provide evidence that RAMPs traffic the CaSR as higher-level oligomers and play a role in CaSR signalling even after cell surface localisation has occurred.
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影响因子：
2.9
作者：
Huang, Yun;Zhou, Yubin;Castiblanco, Adriana;Yang, Wei;Brown, Edward M.;Yang, Jenny J.
通讯作者：
Yang, Jenny J.
影响因子：
4.8
作者：
Bomberger, JM;Spielman, WS;Parameswaran, N
通讯作者：
Parameswaran, N
DOI：
10.1152/ajprenal.2000.279.6.f1083
发表时间：
2000-12-01
影响因子：
4.2
作者：
Handlogten, ME;Shiraishi, N;Miller, RT
通讯作者：
Miller, RT
影响因子：
7.8
作者：
Gerbino, Andrea;Ruder, Warren C;Curci, Silvana;Pozzan, Tullio;Zaccolo, Manuela;Hofer, Aldebaran M
通讯作者：
Hofer, Aldebaran M
影响因子：
4.5
作者：
Faurskov, B;Bjerregaard, HF
通讯作者：
Bjerregaard, HF