RGS5/16-dependent phenotype regulation of arterial vascular smooth muscle cells
Pharmacology
Cell Biology
Final Report Abstract
G-proteins initiate and modulate a plethora of signaling cascades in vascular smooth muscle cells (VSMCs). Regulators of G-protein signaling (RGS) have the capacity to terminate their activity and thus balance the short- and long-term outcome of G-protein-transduced stimuli. Those include agonists such as endothelin 1 (ET-1) and biomechanical stress as an indirect trigger of G-protein activity. In this context, we revealed that hypertension induces the expression of both RGS5 and RGS16 (a homologue to RGS5) in arterial VSMCs from which RGS5 was shown to contribute to structural remodeling processes of the arterial wall. This study (originally proposed by Dr. Arnold) was intended to unravel the functional and mechanistic impact of RGS5 and RGS16 on balancing G-protein-mediated signals as a prerequisite for the activation of arterial VSMCs during exposure to biomechanical stress. The project was hampered by several unforeseen issues including restrictions dependent on the pandemic (breeding restrictions, shutdown of the laboratory) and limitations of some animal models. Collectively, our results suggest that Rgs5 acts as an artery subtype-specific regulator of G-proteindependent signaling. As evidenced by results from both global and SMC-specific Rgs5-deficient mice, RGS5 has little impact on the systemic blood pressure under physiological conditions or hypertension. However, in hypertensive Rgs5-/- mice, the capacity of arterial VSMCs to promote structural remodeling processes is limited. Interestingly, under control conditions loss of RGS5 elevated baseline activity of the MAP kinase ERK1/2 in arterial VSMCs, increased their sensitivity to vasocontractile agonists such as endothelin 1 (ET-1) but lowered the activity of RhoA. On the contrary, Rgs5 overexpression attenuated the Gαq/11-mediated calcium release, IP generation, the activity of Akt- and MAP kinase-related signaling cascades and contractile responses of VSMCs while promoting RhoA activity and growth arrest of proliferating VSMCs. Rgs16 shows low expression levels in mouse arteries under baseline conditions. In line with this observation, in silico analyses of scRNAseq data suggest no regulation of Rgs16 in activated VSMCs derived from arteriosclerotic lesions. Likewise, no cardiovascular phenotype is reported for Rgs16- deficient mice. In vitro, Rgs16 expression was only slightly stimulated in VSMCs by G-protein activating agonists such as ET-1 but rapidly downregulated by biomechanical stretch. Its overexpression promoted RhoA activity in biomechanically stressed VSMCs but not under control conditions. Considering that multiple RGS proteins contribute to the outcome of signaling processes in VSMCs, the overall functional relevance of RGS16 appears to be limited here as its specific impact on G-protein signaling may be covered by redundant functions of other members of the RGS family. Beyond the original scope of this project, we developed a novel 3D in vitro model for the study of VSMC contraction and relaxation that allowed us to reveal the capacity of RGS5 to limit ET-1-mediated MAP kinase activation and VSMC contraction. This model will also serve as a simple but versatile tool for future analyses of agonist-mediated contraction/relaxation of human and mouse VSMCs and complies with the 3R principle (Replace, Reduce, Refine) to support animal welfare.
Publications
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RGS5 Attenuates Baseline Activity of ERK1/2 and Promotes Growth Arrest of Vascular Smooth Muscle Cells. Cells, 10(7), 1748.
Demirel, Eda; Arnold, Caroline; Garg, Jaspal; Jäger, Marius Andreas; Sticht, Carsten; Li, Rui; Kuk, Hanna; Wettschureck, Nina; Hecker, Markus & Korff, Thomas
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Tracing G-Protein-Mediated Contraction and Relaxation in Vascular Smooth Muscle Cell Spheroids. Cells, 12(1), 128.
Garg, Jaspal; Sporkova, Alexandra; Hecker, Markus & Korff, Thomas
