Angiotensin II Activates pp60c-src in Vascular Smooth Muscle Cells

Division of Cardiology, University of Washington, Seattle 98195, USA.
Circulation Research (Impact Factor: 11.02). 01/1996; 77(6):1053-9. DOI: 10.1161/01.RES.77.6.1053
Source: PubMed


The angiotensin II type-1 (AT1) receptor, a G protein-coupled receptor, lacks intrinsic kinase activity. However, recent data show that angiotensin II (Ang II) stimulates tyrosine phosphorylation of phospholipase C-gamma 1 (PLC-gamma 1), Stat91 (one of the signal transducers and activators of transcription), and paxillin in vascular smooth muscle cells. The tyrosine kinases responsible for these phosphorylation events are unknown. Src family kinases have been shown to phosphorylate PLC-gamma 1 and to be activated by G protein-coupled receptors. We hypothesized that pp60c-src associates with the AT1 receptor and is activated after Ang II stimulation of smooth muscle cells. We immunoprecipitated pp60c-src from Ang II-stimulated vascular smooth muscle cells and measured pp60c-src activity by autophosphorylation and by phosphorylation of enolase. Both assays demonstrated an approximately threefold increase in pp60c-src activity within 1 minute. A similar increase in Ang II-stimulated pp60c-src activity was observed in Chinese hamster ovary cells transfected with the AT1 receptor but not in untransfected cells. These data are the first to show that pp60c-src is activated by Ang II. To determine if pp60c-src associated with the AT1 receptor, the AT1 receptor was immunoprecipitated (with two different antibodies), and Western blots were performed with two different anti-pp60c-src antibodies. No pp60c-src was detected. In addition, direct interaction between the AT1 receptor and pp60c-src could not be demonstrated by using a glutathione S-transferase (GST)-AT1 fusion protein to bind proteins from cell lysates stimulated by Ang II.(ABSTRACT TRUNCATED AT 250 WORDS)

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    • "Several earlier studies on AT1aR signalling have described angiotensin II mediated tyrosine phosphorylation [21]–[25]. Proteins harbouring these are often low abundant [2] and in addition current enrichment methods favour peptides phosphorylated on serines or threonines [18]. This leads to tyrosine phosphorylation being significantly underreported; in the two studies the fraction of tyrosine sites was 0.7% and 3.3% for the studies by Christensen and co-workers and Xiao and co-workers respectively (Figure 2B). "
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    • "Studies in a variety of Ang II target cell types have shown that the Ang II activation of different pathways is time dependent. For example, activation of the G-protein-dependent pathway and generation of IP 3 occurs in seconds, while MAPK and JAK/STAT activation occurs in minutes to hours after initial AT1R activation (Ishida et al., 1995; Schmitz et al., 1998). While many of the Ang II signaling cascades have been defined, an understanding of the downstream changes in gene expression has moved ahead very slowly, often through the definition of one gene-target at a time. "
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    • "There is also an increasing recognition for a critical role of reactive oxygen species (ROS) generation in triggering the responses of Ang II (Zafari et al. 1998; Zhang et al. 2005) and ET-1 (Cheng et al. 2003; Daou and Srivastava 2004). Both receptor and nonreceptor protein tyrosine kinases (PTK) have been shown to be activated by Ang II (Du et al. 1996; Ishida et al. 1995; Eguchi et al. 1999; Rocic et al. 2001) and ET-1 (Iwasaki et al. 1999; Kodama et al. 2002; Robin et al. 2002; Daou and Received 17 August 2006. Published on the NRC Research Press Web site at on 7 March 2007. "
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