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Shear Stress Induces G Protein‐Coupled Receptor (GPCR)‐Independent Heterotrimeric G Protein Activation in Endothelial Cells
Author(s) -
Paz Nathaniel,
Melchior Benoit,
Frangos John
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.1029.7
Subject(s) - g protein coupled receptor , mechanotransduction , heterotrimeric g protein , chemistry , microbiology and biotechnology , g protein , receptor , internalization , biophysics , biology , biochemistry
Mechanotransduction is the process by which mechanical forces, such as shear stress, are converted into biochemical responses within the cell. The molecular mechanisms by which endothelial cells (ECs) sense and transduce shear stress into biological signals have not been fully elucidated. G proteins and G protein‐coupled receptors (GPCRs) have been hypothesized independently to mediate mechanotransduction. In this study, we developed a novel read‐out for in situ detection of GPCR/Gα q/11 activation, based on the proximity ligation assay (PLA). We adapted this approach to examine the role of specific GPCR/Gα q/11 pairs in shear stress‐induced EC mechanotransduction. We demonstrated that S1P stimulation causes a rapid dissociation of Gα q/11 from its receptor, S1P 3 , followed by a dramatic increase in association of S1P 3 with both GRK2 and β‐arrestin in human coronary artery ECs (HCAECs), indicative of GPCR/Gα q/11 activation and receptor desensitization/internalization. The S1P 3 antagonist, CAY10444, as well as the G protein inhibitor, GDP‐β‐S, prevented this activation in S1P‐treated cells. Interestingly, shear stress led to the rapid dissociation of Gα q/11 from S1P 3 . However, associations between S1P 3 and GRK2 and S1P 3 and β‐arrestin were not markedly increased over time. Pre‐treatment of cells with GDP‐β‐S prevented the dissociation of Gα q/11 from S1P 3 in response to shear stress. In contrast, treatment of cells with AlF 4 ‐ resulted in increased dissociation of Gα q/11 from S1P 3, but no increase in association between S1P 3 and either GRK2 or β‐arrestin. Altogether, these findings support the notion that Gα q/11 participates in the sensing/transducing of shear stress independent of GPCR activation in ECs.