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Mechanical control of cAMP signaling through integrins is mediated by the heterotrimeric Gαs protein

Mechanical stresses that are preferentially transmitted across the cell surface via transmembrane integrin receptors activate gene transcription by triggering production of intracellular chemical second messengers, such as cAMP. Here we show that the sensitivity of the cAMP signaling pathway to mechanical stresses transferred across β1 integrins is mediated by force‐dependent activation of the heterotrimeric G protein subunit Gαs within focal adhesions at the site of stress application. Gαs is recruited to focal adhesions that form within minutes following clustering of β1 integrins induced by cell binding to magnetic microbeads coated with activating integrin ligands, and β1 integrin and Gαs co‐precipitate when analyzed biochemically. Stress application to activated β1 integrins using magnetic twisting cytometry increases Gαs recruitment and activates these large G proteins within focal adhesions, as measured by binding of biotinylated azido‐anilido‐GTP, whereas application of similar stresses to inactivated integrins or control histocompatibility antigens has little effect. This response is relevant physiologically as application of mechanical strain to cells bound to flexible extracellular matrix‐coated substrates induce translocation of phospho‐CREB to the nucleus, which can be attenuated by inhibiting Gαs activity, either using the inhibitor melittin or suppressing its expression using siRNA. Although integrins are not typical G protein‐coupled receptors, these results show that integrins focus mechanical stresses locally on heterotrimeric G proteins within focal adhesions at the site of force application, and transduce mechanical stimuli into an intracellular cAMP signaling response by activating Gαs at these membrane signaling sites. J. Cell. Biochem. 106: 529–538, 2009. © 2009 Wiley‐Liss, Inc.