Department of Neuroscience, Istituto Superiore di Sanità, 00161 Rome, Italy.
National Centre for Drug Research and Evaluation, Istituto Superiore di Sanità, 00161 Rome, Italy.
Biomolecules. 2023 Sep 5;13(9):1350. doi: 10.3390/biom13091350.
(1) Background: Recently, we found that adenosine A receptor (AR) stimulation results in an increase in STEP phosphatase activity. In order to delve into the mechanism through which AR stimulation induced STEP activation, we investigated the involvement of mGluR since it is well documented that AR and mGluR physically and functionally interact in several brain areas. (2) Methods: In a neuroblastoma cell line (SH-SY5Y) and in mouse hippocampal slices, we evaluated the enzymatic activity of STEP by using a para-nitrophenyl phosphate colorimetric assay. A co-immunoprecipitation assay and a Western blot analysis were used to evaluate STEP/mGluR binding. (3) Results: We found that the AR-dependent activation of STEP was mediated by the mGluR. Indeed, the AR agonist CGS 21680 significantly increased STEP activity, and this effect was prevented not only by the AR antagonist ZM 241385, as expected, but also by the mGluR antagonist MPEP. In addition, we found that mGluR agonist DHPG-induced STEP activation was reversed not only by the mGluR antagonist MPEP but also by ZM 241385. Finally, via co-immunoprecipitation experiments, we found that mGluR and STEP physically interact when both receptors are activated (4) Conclusions: These results demonstrated a close functional interaction between mGlu and A receptors in the modulation of STEP activity.
(1) 背景：最近，我们发现腺苷 A 受体 (AR) 刺激会导致 STEP 磷酸酶活性增加。为了深入研究 AR 刺激诱导 STEP 激活的机制，我们研究了 mGluR 的参与，因为有充分的文献证明 AR 和 mGluR 在几个脑区存在物理和功能相互作用。(2) 方法：在神经母细胞瘤细胞系 (SH-SY5Y) 和小鼠海马切片中，我们通过使用对硝基苯磷酸比色测定法来评估 STEP 的酶活性。通过共免疫沉淀测定和 Western blot 分析来评估 STEP/mGluR 结合。(3) 结果：我们发现 AR 依赖性的 STEP 激活是由 mGluR 介导的。事实上，AR 激动剂 CGS 21680 显著增加了 STEP 活性，这种效应不仅被预期的 AR 拮抗剂 ZM 241385 阻止，而且还被 mGluR 拮抗剂 MPEP 阻止。此外，我们发现 mGluR 激动剂 DHPG 诱导的 STEP 激活不仅被 mGluR 拮抗剂 MPEP 逆转，而且还被 ZM 241385 逆转。最后，通过共免疫沉淀实验，我们发现当两种受体都被激活时，mGlu 和 STEP 物理相互作用。(4) 结论：这些结果表明 mGlu 和 A 受体在调节 STEP 活性方面存在密切的功能相互作用。
