The role of soluble adenylyl cyclase in sensing and regulating intracellular pH.

Soluble adenylyl cyclase (sAC) differs from transmembrane adenylyl cyclases (tmAC) in many aspects. In particular, the activity of sAC is not regulated by G-proteins but by the prevailing bicarbonate concentrations inside cells. Therefore, sAC serves as an exquisite intracellular pH sensor, with the capacity to translate pH changes into the regulation of localization and/or activity of cellular proteins involved in pH homeostasis. In this review, we provide an overview of literature describing the regulation of sAC activity by bicarbonate, pinpointing the importance of compartmentalization of intracellular cAMP signaling cascades. In addition, examples of processes involving proton and bicarbonate transport in different cell types, in which sAC plays an important regulatory role, were described in detail.

Soluble adenylyl cyclase, the cell-autonomous member of the family.

Soluble adenylyl cyclase (sAC) is the evolutionarily most ancient of a set of 10 adenylyl cyclases (Adcys). While Adcy1 to Adcy9 are cAMP-producing enzymes that are activated by G-protein coupled receptors (GPCRs), Adcy10 (sAC) is an intracellular adenylyl cyclase. sAC plays a pivotal role in numerous cellular processes, ranging from basic physiological functions to complex signaling cascades. As a distinct member of the adenylyl cyclase family, sAC is not activated by GPCRs and stands apart due to its unique characteristics, regulation, and localization within cells. This minireview aims to honour Ulli Brandt, the outgoing Executive Editor of our journal, Biochimica Biophysica Acta (BBA), and longstanding Executive Editor of the BBA section Bioenergetics. We will therefore focus this review on bioenergetic aspects of sAC and, in addition, review some important recent general developments in the field of research on sAC.

Cholangiocytes express an isoform of soluble adenylyl cyclase that is N-linked glycosylated and secreted in extracellular vesicles.

Soluble adenylyl cyclase (sAC)-derived cAMP regulates various cellular processes; however, the regulatory landscape mediating sAC protein levels remains underexplored. We consistently observed a 85 kD (sAC85 ) or 75 kD (sAC75 ) sAC protein band under glucose-sufficient or glucose-deprived states, respectively, in H69 cholangiocytes by immunoblotting. Deglycosylation by PNGase-F demonstrated that both sAC75 and sAC85 are N-linked glycosylated proteins with the same polypeptide backbone. Deglycosylation with Endo-H further revealed that sAC75 and sAC85 carry distinct sugar chains. We observed release of N-linked glycosylated sAC (sACEV ) in extracellular vesicles under conditions that support intracellular sAC85 (glucose-sufficient) as opposed to sAC75 (glucose-deprived) conditions. Consistently, disrupting the vesicular machinery affects the maturation of intracellular sAC and inhibits the release of sACEV into extracellular vesicles. The intracellular turnover of sAC85 is extremely short (t1/2 ~30 min) and release of sACEV in the medium was detected within 3 h. Our observations support the maturation and trafficking in cholangiocytes of an N-linked glycosylated sAC isoform that is rapidly released into extracellular vesicles.

Soluble adenylyl cyclase regulates the cytosolic NADH/NAD+ redox state and the bioenergetic switch between glycolysis and oxidative phosphorylation.

The evolutionarily conserved soluble adenylyl cyclase (sAC, ADCY10) mediates cAMP signaling exclusively in intracellular compartments. Because sAC activity is sensitive to local concentrations of ATP, bicarbonate, and free Ca2+, sAC is potentially an important metabolic sensor. Nonetheless, little is known about how sAC regulates energy metabolism in intact cells. In this study, we demonstrated that both pharmacological and genetic suppression of sAC resulted in increased lactate secretion and decreased pyruvate secretion in multiple cell lines and primary cultures of mouse hepatocytes and cholangiocytes. The increased extracellular lactate-to-pyruvate ratio upon sAC suppression reflected an increased cytosolic free [NADH]/[NAD+] ratio, which was corroborated by using the NADH/NAD+ redox biosensor Peredox-mCherry. Mechanistic studies in permeabilized HepG2 cells showed that sAC inhibition specifically suppressed complex I of the mitochondrial respiratory chain. A survey of cAMP effectors revealed that only selective inhibition of exchange protein activated by cAMP 1 (Epac1), but not protein kinase A (PKA) or Epac2, suppressed complex I-dependent respiration and significantly increased the cytosolic NADH/NAD+ redox state. Analysis of the ATP production rate and the adenylate energy charge showed that inhibiting sAC reciprocally affects ATP production by glycolysis and oxidative phosphorylation while maintaining cellular energy homeostasis. In conclusion, our study shows that, via the regulation of complex I-dependent mitochondrial respiration, sAC-Epac1 signaling regulates the cytosolic NADH/NAD+ redox state, and coordinates oxidative phosphorylation and glycolysis to maintain cellular energy homeostasis. As such, sAC is effectively a bioenergetic switch between aerobic glycolysis and oxidative phosphorylation at the post-translational level.