Bicarbonate-sensing soluble adenylyl cyclase is an essential sensor for acid/base homeostasis

Abstract

pH homeostasis is essential for life, yet it remains unclear how animals sense their systemic acid/base (A/B) status. Soluble adenylyl cyclase (sAC) is an evolutionary conserved signaling enzyme that produces the second messenger cAMP in response to bicarbonate ions (HCO 3 − ). We cloned the sAC ortholog from the dogfish, a shark that regulates blood A/B by absorbing and secreting protons (H + ) and HCO 3 − at its gills. Similar to mammalian sAC, dogfish soluble adenylyl cyclase (dfsAC) is activated by HCO 3 − and can be inhibited by two structurally and mechanistically distinct small molecule inhibitors. dfsAC is expressed in the gill epithelium, where the subset of base-secreting cells resides. Injection of inhibitors into animals under alkaline stress confirmed that dfsAC is essential for maintaining systemic pH and HCO 3 − levels in the whole organism. One of the downstream effects of dfsAC is to promote the insertion of vacuolar proton pumps into the basolateral membrane to absorb H + into the blood. sAC orthologs are present throughout metazoans, and mammalian sAC is expressed in A/B regulatory organs, suggesting that systemic A/B sensing via sAC is widespread in the animal kingdom.

How to get into bones: proton pump and carbonic anhydrase inOsedaxboneworms

“…These antibodies recognize the epitope 'MQNAFRSLED' [27], which is more than 80 per cent conserved from cnidarians to mammals. These antibodies specifically cross-react with VHA A from a wide range of species, including mammals [28], fish [26] and mollusc tissues (M. Tresguerres 2012, personal observation).…”

“…Preliminary transcriptomic analyses demonstrate this epitope is 100 per cent conserved in VHA B from Osedax frankpressi (H. Tresguerres 2012, unpublished observation). The antibodies specifically recognize VHA B from fish, shark and mollusc tissues by Western blot, as well as 'VHA-rich cells' in shark gills by immunohistochemistry [26]. Antibodies against mouse VHA A were a kind gift from Dr Dennis Brown (Harvard University, MA, USA).…”

“…Both methods yielded identical results. Sections were attached to glass slides, rehydrated and processed for immunofluorescence as previously described [26]. After blocking non-specific binding sites (2% normal goat serum in 10 mM PBS, pH 7.4), sections were incubated overnight at 48C with antibodies against VHA or CA2 (3 mg ml 21 in PBS).…”

“…-secreting, H þ -absorbing cells such as gill cells from hagfish [31,32], shark [26,[33][34][35] and trout [36], and b-intercalated cells from mammalian kidney [21,37,38].…”

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“…It involves regulatory hypoxia response elements (HRE), factors (e.g. hypoxia inducible factor or HIF), hydroxylases and proteases and acid/base homoeostasis adenylyl cyclases (Gracey et al, 2001;Wenger, 2002;Law et al, 2006;Rytkönen et al, 2007;Tresguerres et al, 2010;Chepelev and Gilmore, 2011).…”