Abstract 330: AT1R Conformation Mediates Mechanical Stress-Induced β-Arrestin Signaling
Wei Tang1, Howard A. Rockman1,2,3 1Departments of Medicine, 2Departments of Cell Biology, 3Departments of Molecular Genetics and Microbiology, Duke University Medical Center, DUMC 3104, 226 CARL Building, Durham, NC 27710, USA.
Introduction: Recent work has shown that mechanical stretch induces β-arrestin-biased Angiotensin II Type 1 Receptor (AT1R) prosurvival cellular signaling. However, the precise molecular details for this are not fully understood. We set out to explore whether the conformation of the AT1R determines stretch-induced cellular signaling and whether this mechanism of stretch-mediated beta-arrestin signaling activates cell cycle progression.
Methods: We used HEK 293 cells stably expressing AT1Rs with and without treatment with the nonionic triblock membrane sealant poloxamer 188 (P188). P188 is known to integrate into biological membranes and affect the local microviscosity of the membrane. Mechanical stretch was induced by hypotonic osmotic stress. β-arrestin signaling was examined by the level of phosphorylated ERK (pERK) with and without siRNA targeted to β-arrestin1/2, and cell cycle progression by the level of phosphorylated retinoblastoma (Rb).
Results: Both Angiotensin II (AngII) and osmotic stress increased pERK. The increase in pERK by osmotic stress was abrogated in the presence of β-arrestin1/2 siRNA. P188 had no effect on AngII induced pERK, but completely blocked the increase in pERK with osmotic stress. Both AngII and osmotic stress caused the rapid increase phosphorylated Rb at the Ser780 site, which is known to be phosphorylated by cyclin-dependent protein kinase 4.
Conclusion: Altering membrane viscosity blocks stretch-induced AT1R signaling indicating a possible role for receptor conformation in stretch-mediated β-arrestin signaling and cell cycle progression. Ongoing studies will directly measure receptor affinity states under ligand-activated vs. mechanical stretch-activated conditions.
- © 2012 by American Heart Association, Inc.