![]() We found constitutive association between D2DR, VEGFR-2 and SHP-2. Whole-cell lysates from serum-starved HUVECs were immunoprecipitated with D2DR antibody and immunoblotted with antibodies against VEGFR-2 and SHP-2. Therefore, we performed coprecipitation experiments to identify specific protein tyrosine phosphatases that are involved in the dopamine-mediated VEGFR-2 dephosphorylation. ResultsĪs dopamine modulates VEGFR-2 tyrosine phosphorylation, we posited that certain phosphatases might be involved in this modulation. Here, we examine how dopamine specifically regulates one of the earliest steps of the angiogenic process, the VEGF-induced phosphorylation of VEGFR-2, and define a novel role of Src-homology-2-domain-containing protein tyrosine phosphatase 2 (SHP-2) in the dopamine-mediated anti-angiogenic pathway. Although it is known that dopamine can inhibit different signaling steps involved in VEGF-mediated angiogenesis, beginning at VEGFR-2 phosphorylation ( Basu et al., 2001 Bhattacharya et al., 2008b Sarkar et al., 2004), the pathophysiological implication of dopamine in the context of VEGF-induced angiogenesis is novel and requires further investigation. Dopamine, when applied to patients with septic shock, can effectively maintain the circulatory stability and promote restoration of renal function ( Dellinger et al., 2008). Dopamine has a role in the management of Parkinson's disease and is also involved in the etiopathogenesis of several neuropsychiatric diseases such as schizophrenia ( Egan and Weinberger, 1997 Goldstein and Deutch, 1992 Graybiel et al., 1990 Olanow and Tatton, 1999). Dopamine is produced in a wide variety of animals including both vertebrates and invertebrates, and activates the five types of dopamine receptors – D1, D2, D3, D4 and D5 – and their variants ( Neve et al., 2004). ![]() The role of the neurotransmitter dopamine in modulating VEGF-induced angiogenesis has been well defined ( Basu et al., 2001 Sarkar et al., 2004). The binding of VEGF to its receptors induces dimerization and subsequent receptor phosphorylation, which then leads to the activation of several intracellular downstream signaling pathways promoting angiogenesis ( Olsson et al., 2006 Sakurai et al., 2005 Takahashi et al., 2001 Zachary, 2003). VEGFR-2, the major positive signal transducer for both physiological and pathological angiogenesis ( Mukhopadhyay et al., 2004 Shibuya, 2006), is selectively expressed on vascular endothelial cells. The VEGF receptor family consists of three members, namely VEGFR-1, VEGFR-2 and VEGFR-3 ( Shibuya, 2008). As one of the most important cytokines, VEGF is widely expressed by a number of human and animal tumors and has the ability to regulate most of the steps in the angiogenic signal cascade ( Dvorak, 1990 Dvorak et al., 1999 Dvorak et al., 1979 Ferrara, 1999 Risau, 1997). Vascular endothelial growth factor (VEGF), also known as vascular permeability factor (VPF), is an important mediator of these angiogenic processes in both normal and diseased conditions. Our data establish a novel role for SHP-2 phosphatase in the dopamine-mediated regulation of VEGFR-2 phosphorylation.Īngiogenesis performs a crucial role in physiological conditions, including embryonic development, wound healing and organ regeneration, as well as in pathological processes such as diabetes retinopathies, atherosclerosis, tumorigenesis and tumor metastasis ( Carmeliet, 2003 Ferrara et al., 2003 Folkman, 1971 Folkman, 1995 Sueishi et al., 1997). We also observe that SHP-2 knockdown impairs the dopamine-regulated inhibition of VEGF-induced phosphorylation of VEGFR-2 and, subsequently, Src phosphorylation and migration. Active SHP-2 then dephosphorylates VEGFR-2 at Y951, Y996 and Y1059, but not Y1175. Dopamine administration leads to increased VEGF-induced phosphorylation of SHP-2 and this increased phosphorylation parallels the increased phosphatase activity of SHP-2. Dopamine pretreatment increases the translocation and colocalization of Src-homology-2-domain-containing protein tyrosine phosphatase (SHP-2) with D2DR at the cell surface. Here, we demonstrate that D2 dopamine receptor (D2DR) colocalizes with VEGFR-2 at the cell surface. ![]() ![]() In this study, we address the mechanism by which VEGFR-2 phosphorylation is regulated by dopamine. Our previous studies have shown that the neurotransmitter dopamine could inhibit VEGF-induced phosphorylation of VEGF receptor 2 (VEGFR-2), endothelial cell proliferation, migration, microvascular permeability, and thus, angiogenesis. Vascular endothelial growth factor (VEGF)-induced receptor phosphorylation is the crucial step for initiating downstream signaling pathways that lead to angiogenesis or related pathophysiological outcomes. ![]()
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