Characterization of the molecular mechanism underlying PI3Kgamma-dependent CFTR phosphorylation and stabilization at the plasma membrane

Abstract The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-dependent Cl- transporter whose dysfunction mostly affects the respiratory tract, resulting in an impaired mucociliary clearance with consequent airway obstruction and chronic inflammation. Decreased CFTR function can be found in obstructive airway diseases, such as asthma, chronic obstructive pulmonary disease (COPD) and cystic fibrosis (CF), a genetic disorder caused by mutations in the CFTR gene. In 2019 FDA approved a combination of CFTR modulators, named Trikafta®, that rescues the function of the most common CFTR mutant found in CF patients, F508del, characterized by defective protein folding and channel gating. However, Trikafta® rescues F508del-CFTR activity only up to 60% of physiological values, underlying the need for alternative treatments. One possible therapeutic strategy is to activate the β2-adrenergic receptor (AR)/cAMP axis, stimulating the CFTR activation and stabilization at the plasma membrane (PM). Here, we describe a cell-permeable peptide that, by disrupting the protein kinase A (PKA)-anchoring function of phosphoinositide 3-kinase γ (PI3Kγ), inhibits cAMP degradation by phosphodiesterases (PDEs) 4B and 4D and amplifies β2-AR/cAMP responses in airway cells. We demonstrated that the PI3Kγ mimetic peptide (PI3Kγ MP) increases cAMP levels in the vicinity of the CFTR, promoting the phosphorylation of a specific activating residue, S737. Moreover, the PI3Kγ MP rescues CFTR trafficking to the PM in a cAMP/PKA-independent manner, possibly involving ER Ca2+ store depletion and protein kinase D (PKD) 1 activation. Finally, we found that the PI3Kγ MP synergizes with CFTR modulators, doubling Cl- secretion and enhancing CFTR stabilization, suggesting a potential therapeutic application for obstructive airway diseases, especially CF.