Distinct expression and ligand-binding profiles of two constitutively active GPR17 splice variants.
Distinct expression and ligand-binding profiles of two constitutively active GPR17 splice variants.
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DOI：
10.1111/j.1476-5381.2009.00633.x
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发表时间：
2010-03
影响因子：
7.3
通讯作者：
Rosenkilde MM
中科院分区：
文献类型：
--
作者：
Benned-Jensen T;Rosenkilde MM
在人类和非人类灵长类动物中，7 TM受体GPR 17以两种同种型存在，仅N-末端的长度不同。其中，只有短亚型先前已被表征。因此，我们研究了两种剪接变体的基因表达和配体结合谱，并进一步揭示和表征了两种亚型的组成型活性。使用定量RT-PCR测定几个脑区域以及心脏和肾脏中hGPR 17同种型的表达水平。采用CREB报告基因测定和[35 S]-GTPγS结合来评估组成型活性和UDP、UDP-葡萄糖和-半乳糖以及半胱氨酰白三烯LTC 4和LTD 4的活化。分别使用同源竞争结合和抗体喂养实验进一步测试白三烯结合和内化的诱导。短同种型（hGPR 17-S）在脑中比长同种型（hGPR 17-L）表达更丰富（8至23倍），而在心脏和肾脏中观察到相反的情况。如先前报道的，尿嘧啶核苷酸以微摩尔效力激活hGPR 17-S。然而，观察到hGPR 17-L的效力低得多，EC 50增加50至170倍。此外，与以前的报道相反，这两种亚型都没有被半胱氨酰白三烯激活或结合。最后，通过Gαi证实两种受体都具有组成性活性。我们提出了GPR 17的第一个亚型特异性表征，并表明亚型之间存在差异，在表达模式和药理学特征。反过来，我们的研究结果表明，这两种人类亚型可能具有组织特异性功能。
In humans and non-human primates, the 7TM receptor GPR17 exists in two isoforms differing only by the length of the N-terminus. Of these, only the short isoform has previously been characterized. Hence, we investigated gene expression and ligand-binding profiles of both splice variants and furthermore uncovered and characterized constitutive activity of both isoforms. Expression levels of the hGPR17 isoforms were determined in several brain regions as well as heart and kidney using quantitative RT-PCR. A CREB reporter assay and [35S]-GTPγS binding were employed to assess the constitutive activity and the activation by UDP, UDP-glucose and -galactose and the cysteinyl leukotrienes LTC4 and LTD4. Leukotriene binding and induction of internalization were furthermore tested using homologous competition binding and antibody-feeding experiments respectively. The short isoform (hGPR17-S) was expressed more abundantly (eight- to 23-fold) in the brain than the long isoform (hGPR17-L), whereas the opposite was observed in heart and kidney. As previously reported, the uracil nucleotides activated hGPR17-S with micromolar potencies. However, much lower potencies were observed for hGPR17-L with a 50- to 170-fold increase in EC50. Furthermore, contrary to previous reports, neither of the isoforms was activated or bound by the cysteinyl leukotrienes. Finally, both receptors were demonstrated to be constitutively active through Gαi. We present the first isoform-specific characterization of GPR17 and show that differences exist between the isoforms, in both expression pattern and pharmacological profile. In turn, our results indicate that the two human isoforms might serve tissue-specific functions.
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影响因子：
--
作者：
Kostich, WA;Chen, AR;Largent, BL
通讯作者：
Largent, BL
影响因子：
4.8
作者：
Chambers, JK;Macdonald, LE;Livi, GP
通讯作者：
Livi, GP
影响因子：
--
作者：
Holst, B;Cygankiewicz, A;Schwartz, TW
通讯作者：
Schwartz, TW
影响因子：
3.6
作者：
Benned-Jensen, Tau;Rosenkilde, Mette M.
通讯作者：
Rosenkilde, Mette M.
DOI：
10.1006/bbrc.1995.1328
发表时间：
1995-03-08
影响因子：
3.1
作者：
MIYAKE, A
通讯作者：
MIYAKE, A