Buisson B, Laflamme L, Bottari S P, de Gasparo M, Gallo-Payet N, Payet M D
Service of Endocrinology, Faculty of Medicine, University of Sherbrooke, Québec, Canada.
J Biol Chem. 1995 Jan 27;270(4):1670-4. doi: 10.1074/jbc.270.4.1670.
In non-differentiated NG108-15 cells, both angiotensin II (Ang II) (100 nM) and CGP 42112 (100 nM) decreased the T-type calcium current amplitude by 24 +/- 2% and 21 +/- 3%, respectively. cGMP is not a mediator of the Ang II effect, since loading of cells with 50 microM cGMP did not prevent the inhibitory effects of Ang II. The effects of Ang II involves a non-identified GTPase activity since incubation with GDP beta S (3 mM) completely reversed the inhibitory effect of Ang II while GTP gamma S mimicked its effect. However, Ang II binding was not affected by GTP gamma S, and the effect of Ang II was not modified in pertussis toxin-treated cells. The inhibitory effect of Ang II on the T-type Ca2+ current involves a phosphotyrosine phosphatase activity since sodium orthovanadate prevented the effects of Ang II, although microcystin-LR, a selective Ser/Thr phosphatase 1 and 2A inhibitor, did not modify the effect of Ang II. These results provide the first evidence of a modulation of membrane conductance by Ang II through the AT2 receptor and demonstrate the involvement of a phosphotyrosine phosphatase and a G protein in the AT2 transduction mechanism in NG108-15 cells. Moreover, our data suggest that phosphotyrosine phosphatase activation is proximal to receptor occupation, since sodium orthovanadate inhibits both GTPase activity and T-type current blockage induced by Ang II or CGP 42112, while GTP gamma S inhibition of the T-type calcium current is not impaired.
在未分化的NG108 - 15细胞中，血管紧张素II（Ang II）（100 nM）和CGP 42112（100 nM）分别使T型钙电流幅度降低了24±2%和21±3%。cGMP不是Ang II作用的介质，因为用50 microM cGMP处理细胞并不能阻止Ang II的抑制作用。Ang II的作用涉及一种未明确的GTP酶活性，因为与GDPβS（3 mM）共同孵育可完全逆转Ang II的抑制作用，而GTPγS可模拟其作用。然而，GTPγS并不影响Ang II的结合，且在百日咳毒素处理的细胞中Ang II的作用未改变。Ang II对T型Ca2+电流的抑制作用涉及一种磷酸酪氨酸磷酸酶活性，因为原钒酸钠可阻止Ang II的作用，尽管微囊藻毒素 - LR（一种选择性丝氨酸/苏氨酸磷酸酶1和2A抑制剂）并未改变Ang II的作用。这些结果首次证明了Ang II通过AT2受体对膜电导的调节作用，并证明了磷酸酪氨酸磷酸酶和G蛋白参与了NG108 - 15细胞中AT2转导机制。此外，我们的数据表明磷酸酪氨酸磷酸酶的激活发生在受体被占据之后，因为原钒酸钠可抑制由Ang II或CGP 42112诱导的GTP酶活性和T型电流阻断，而GTPγS对T型钙电流的抑制作用并未受损。
