RGS2 Modulation of Angiotensin II Receptor Type 1 Trafficking and Signaling in Neuroblastoma Cells

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IntroductionAltered expression and function of brain angiotensin II receptor type 1 (AT1R) has been implicated in the pathophysiology of various neuropsychiatric diseases such as drug addiction and Alzheimer's disease. However, there is a knowledge gap in understanding the molecular mechanisms of altered AT1R trafficking and function. Regulators of G‐protein signaling (RGS) proteins are negative modulators of G‐protein coupled receptors (GPCRs). Mechanistically, RGS proteins terminate signaling by accelerating the hydrolysis of GTP bound to active G‐alpha subunits. Among all the RGS subtypes, our lab found that RGS2 co‐localizes with AT1R in the brain, suggesting a potential role of RGS2 regulation of AT1R signaling. Additionally, our lab previously found that RGS2 regulates the constitutive and agonist‐induced trafficking of dopamine D2 receptors. Therefore, this project assessed the role of RGS2 in regulation of AT1R trafficking and signaling using a neuroblastoma 2a (N2A) cell model. Knowledge learned from this project may be relevant to the physiological function of RGS2 in the brain.MethodsWe generated stable expression of the human Angiotensin Type I receptor (HA‐AT1R) in N2A cells. N2A‐AT1R cells were then transiently transfected with RGS2 siRNA to knock down RGS2 expression. Cells were then treated with vehicle or angiotensin II (AngII, 10 μM) for 2, 5, or 15 minutes for western blotting of phosphorylated Akt and total Akt. Immunocytochemistry and confocal microscopy were performed on cells treated with AngII for various times to measure the trafficking of AT1R.ResultsWe found that the knockdown of RGS2 enhanced AT1R‐mediated signaling by increasing Akt phosphorylation. RGS2 knockdown also showed to increase AngII‐induced internalization of AT1R. This is the first report on the role of RGS2 in regulation of AT1R trafficking. Further study is necessary to investigate the molecular mechanisms underlying RGS2 modulation of the signaling and trafficking of AT1R in neuronal cells.Support or Funding InformationThis work is supported by NIH R01DA042862, P50DA006634, F31AA025532This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.