CK2 is a key regulator of SLC4A2-mediated Cl−/HCO3 − exchange in human airway epithelia

Abstract

Transepithelial bicarbonate secretion by human airway submucosal glands and surface epithelial cells is crucial to maintain the pH-sensitive innate defence mechanisms of the lung. cAMP agonists stimulate HCO3 − secretion via coordinated increases in basolateral HCO3 − influx and accumulation, as well as CFTR-dependent HCO3 − efflux at the luminal membrane of airway epithelial cells. Here, we investigated the regulation of a basolateral located, DIDS-sensitive, Cl−/HCO3 − exchanger, anion exchanger 2 (AE2; SLC4A2) which is postulated to act as an acid loader, and therefore potential regulator of HCO3 − secretion, in human airway epithelial cells. Using intracellular pH measurements performed on Calu-3 cells, we demonstrate that the activity of the basolateral Cl−/HCO3 − exchanger was significantly downregulated by cAMP agonists, via a PKA-independent mechanism and also required Ca2+ and calmodulin under resting conditions. AE2 contains potential phosphorylation sites by a calmodulin substrate, protein kinase CK2, and we demonstrated that AE2 activity was reduced in the presence of CK2 inhibition. Moreover, CK2 inhibition abolished the activity of AE2 in primary human nasal epithelia. Studies performed on mouse AE2 transfected into HEK-293T cells confirmed almost identical Ca2+/calmodulin and CK2 regulation to that observed in Calu-3 and primary human nasal cells. Furthermore, mouse AE2 activity was reduced by genetic knockout of CK2, an effect which was rescued by exogenous CK2 expression. Together, these findings are the first to demonstrate that CK2 is a key regulator of Cl−-dependent HCO3 − export at the serosal membrane of human airway epithelial cells.Electronic supplementary materialThe online version of this article (doi:10.1007/s00424-017-1981-3) contains supplementary material, which is available to authorized users.

Protein kinase CK2: a potential therapeutic target for diverse human diseases

“…Numerous CK2 consensus motifs have been identified on SLC4A2 aminoacidic sequence; most importantly, the exposure of both Calu-3 cells (a human lung cancer cell line) and primary human nasal cells to the CK2 inhibitor CX-4945 led to the almost complete inhibition of channel activity. 230 In the same way, it was shown that also the activity of TMEM16A, a Ca 2+ -activated chloride channel, is positively regulated by CK2; in fact, the TMEM16A conductance in CFBE airway epithelial cells is strongly hampered by both the pharmacological inhibition (mediated by TBB and CX-4945) and the transient downregulation of the α′ subunit of the kinase. 231 These data further support the strong CK2 involvement in chloride and hydrogen carbonate homeostasis, that in the airway epithelia is fundamental for adequate solubilization and excretion of mucus, as well as for efficient antimicrobial action.…”

Drug Repurposing for Cystic Fibrosis: Identification of Drugs That Induce CFTR-Independent Fluid Secretion in Nasal Organoids

“…For instance, cAMP agonists activate the Cl − channel CLCN2 [ 34 ], SLC26A9 [ 35 ], the Cl − /HCO 3 − exchanger pendrin (SLC26A4) [ 36 ], the Na + /HCO 3 − cotransporter NBCE1 [ 37 ] and the H + /K + ATPase HKA2 (ATP12A) [ 29 , 38 ]. Additionally, it has been shown that cAMP modulates the expression of the Cl − /HCO 3 − exchanger type 2 (AE2 or SLC4A2) in airway epithelial cells [ 29 , 39 , 40 ]. Furthermore, cAMP mediates K + signaling, e.g., by activation of the basolateral membrane K + channel KCNQ1 [ 39 , 41 ], which is associated with chloride secretion [ 42 ].…”

Drug repurposing for Cystic Fibrosis: identification of drugs that induce CFTR-independent fluid secretion in nasal organoids

“…For instance, cAMP agonists activate the Cl - channel SLC26A9 [33], the Cl − /HCO3 − exchanger pendrin (SLC26A4) [34], the Na + /HCO3 - cotransporter NBCE1 [35], and the H + /K + ATPase HKA2 (ATP12A) [28,36]. Additionally, it has been shown that cAMP modulates the expression of the Cl - /HCO3 - exchanger type 2 (AE2 or SLC4A2) in airway epithelial cells [28,37,38]. Furthermore, cAMP mediates K + signaling, e.g.…”