AT2 receptors recruit c-Src, SHP-1 and FAK upon activation by Ang II in PND15 rat hindbrain

The functional role of AT2 receptors is unclear and it activates unconventional signaling pathways, which in general do not involve a classical activation of a G-protein. In the present study, we aimed to investigate the transduction mechanism of AT2 Ang II receptors in PND15 rat hindbrain membrane preparations, which represents a physiological developmental condition. To determine whether Ang II AT2 receptors induced association to SHP-1 in rat hindbrain, co-immunoprecipitation assays were performed. Stimulation of Ang II AT2 receptors induced both a transient tyr-phosphorylation and activation of SHP-1. The possible participation of c-Src in Ang II-mediated SHP-1 activation, we demonstrated by recruitment of c-Src in immunocomplexes obtained with anti AT2 or anti-SHP-1 antibodies. The association of SHP-1 to c-Src was inhibited by PD123319 and the c-Src inhibitor PP2. Similarly, SHP-1 activity determined in AT2-immunocomplexes was inhibited by PD123319 and the c-Src inhibitor PP2. Following stimulation with Ang II, AT2 receptors recruit c-Src, which was responsible for SHP-1 tyr-phosphorylation and activation. Since AT2 receptors are involved in neuron migration, we tested the presence of FAK in immunocomplexes. Surprisingly, AT2-immunocomplexes contained mainly the 85 kDa fragment of FAK. Besides, p125FAK associated to SHP-1. In summary, we demonstrated the presence of an active signal transduction mechanism in PND15 rat hindbrain, a developmental stage critical for cerebellar development. In this model, we showed a complex containing AT2/SHP-1/c-Src/p85FAK, suggesting a potential role of Ang II AT2 receptors in cerebellar development and neuronal differentiation.

► Ang II AT2 receptors induced a transient tyr-phosphorylation and activation of SHP-1.
► AT2 receptors recruit c-Src, which induced SHP-1 tyr-phosphorylation and activation.
► We observed the formation of a complex containing AT2/SHP-1/c-Src/p85FAK.
► Our observation suggests that a functional complex was formed following activation.