Established and potential physiological roles of bicarbonate-sensing soluble adenylyl cyclase (sAC) in aquatic animals

Abstract

Soluble adenylyl cyclase (sAC) is a recently recognized source of the signaling molecule cyclic AMP (cAMP) that is genetically and biochemically distinct from the classic G-protein-regulated transmembrane adenylyl cyclases (tmACs). Mammalian sAC is distributed throughout the cytoplasm and it may be present in the nucleus and inside mitochondria. sAC activity is directly stimulated by HCO3(-), and sAC has been confirmed to be a HCO3(-) sensor in a variety of mammalian cell types. In addition, sAC can functionally associate with carbonic anhydrases to act as a de facto sensor of pH and CO2. The two catalytic domains of sAC are related to HCO3(-)-regulated adenylyl cyclases from cyanobacteria, suggesting the cAMP pathway is an evolutionarily conserved mechanism for sensing CO2 levels and/or acid/base conditions. Reports of sAC in aquatic animals are still limited but are rapidly accumulating. In shark gills, sAC senses blood alkalosis and triggers compensatory H(+) absorption. In the intestine of bony fishes, sAC modulates NaCl and water absorption. And in sea urchin sperm, sAC may participate in the initiation of flagellar movement and in the acrosome reaction. Bioinformatics and RT-PCR results reveal that sAC orthologs are present in most animal phyla. This review summarizes the current knowledge on the physiological roles of sAC in aquatic animals and suggests additional functions in which sAC may be involved.

Keywords: Acid/base; Carbonic anhydrase; Proton pump; V-ATPase; cAMP; pH sensing.

Fig. 1.

Intracellular cAMP-signaling microdomains. cAMP production may occur in discrete intracellular compartments such as (1) focal points throughout the cytoplasm, (2) the nucleus, (3) mitochondria, (4) the cell membrane vicinity and (5) internalized endosomes. Additional regulation might involve the movement of soluble adenylyl cyclase (sAC) between compartments (not shown). Each microdomain contains a source of cAMP [sAC or transmembrane adenylyl cyclase (tmAC)]; phosphodiesterases (PDEs) that degrade cAMP, thus acting as barriers for cAMP diffusion; and cAMP targets such as protein kinase A (PKA) or exchange protein activated by cAMP (EPAC) (not depicted). Production of cAMP by sAC is stimulated by increased [HCO₃⁻] (and in some cases Ca²⁺, see ‘Discovery of mammalian sAC’ and Fig. 2 for details). Production of cAMP by tmAC occurs in response to various extracellular ligands and it requires modulation by G-protein-coupled receptors and G-protein.

Fig. 2.

Mechanisms of sAC activation in vivo. (1) sAC in the cytoplasm can be stimulated by HCO₃⁻ from various sources. (a) Carbonic anhydrase (CA)-dependent hydration of external CO₂. (b) CA-dependent hydration of metabolic CO₂. (c) H⁺-extruding transporters (HE) such as V-type H⁺-ATPase or Na⁺/H⁺ exchangers from the cell may prevent slowing down of the CO₂ hydration reaction. (d,e) HCO₃⁻ that enters through membrane-transporting proteins such as electrogenic Na⁺/HCO₃⁻-cotransporters (NBCs), anion exchangers or cystic fibrosis transmembrane conductance regulator (CFTR) channels. (f) The entry of HCO₃⁻ across transporters, exchangers and channels can potentially be modulated by hormones. (2) sAC in the nucleus may be stimulated by HCO₃⁻ derived from all the sources listed above. (3) sAC in the cytoplasm may be stimulated by catalytic metals (e.g. Ca²⁺ in mammals), which enter the cell through voltage-dependent Ca²⁺ channels (VDCC) or potentially by Ca²⁺ released from the endoplasmic reticulum or mitochondria (not depicted). (4) sAC inside mitochondria may be stimulated by metabolically generated CO₂ through CA.

Fig. 3.

sAC-dependent sensing and compensation of blood alkalosis. During increased blood HCO₃⁻ and pH, (1) HCO₃⁻ in blood is dehydrated into CO₂ by extracellular carbonic anhydrase (CA_IV). (2) CO₂ enters the V-type H⁺-ATPase-rich cells, where it is hydrated back into H⁺ and HCO₃⁻ by intracellular CA II (CA_II). (3) The elevated intracellular HCO₃⁻ stimulates sAC to generate cAMP, which triggers the microtubule-dependent translocation of V-type H⁺-ATPase (blue icon) containing cytoplasmic vesicles to the basolateral membrane. Basolateral V-type H⁺-ATPase reabsorbs H⁺ into the blood to counteract the original alkalosis. (4) The excess HCO₃⁻ is secreted to seawater in exchange for chloride via a Pendrin-like anion exchanger (AE). Chloride is probably absorbed into the blood across a basolateral channel (CC).

Fig. 4.

Expression of sAC in dogfish shark tissues. (A,B) sAC protein detected by western immunoblot in rectal gland and red blood cells; the band matches the predicted ~110 kDa sAC protein. (C) sAC mRNA detected by RT-PCR in gill (positive control) and rectal gland. (D) Immunolocalization of sAC (brown) in rectal gland cells, showing cytoplasmic and potentially nuclear localization (nuclei stained in green). Methods followed those described previously (Tresguerres et al., 2010c).