Sodium regulation of GAF domain function.

Cyclic nucleotide PDEs (phosphodiesterases) regulate cellular levels of cAMP and cGMP by controlling the rate of degradation. Several mammalian PDE isoforms possess N-terminal GAF (found in cGMP PDEs, Anabaena adenylate cyclases and Escherichia coli FhlA; where FhlA is formate hydrogen lyase transcriptional activator) domains that bind cyclic nucleotides. Similarly, the CyaB1 and CyaB2 ACs (adenylate cyclases) of the cyanobacterium Anabaena PCC 7120 bind cAMP through one (CyaB1) or two (CyaB2) N-terminal GAF domains and mediate autoregulation of the AC domain. Sodium inhibits the activity of CyaB1, CyaB2 and mammalian PDE2A in vitro through modulation of GAF domain function. Furthermore, genetic ablation of cyaB1 and cyaB2 gives rise to Anabaena strains defective in homoeostasis at limiting sodium. Sodium regulation of GAF domain function has therefore been conserved since the eukaryotic/prokaryotic divergence. The GAF domain is the first identified protein domain to directly sense and signal changes in environmental sodium.

Soluble adenylyl cyclase as an evolutionarily conserved bicarbonate sensor.

Spermatozoa undergo a poorly understood activation process induced by bicarbonate and mediated by cyclic adenosine 3',5'-monophosphate (cAMP). It has been assumed that bicarbonate mediates its effects through changes in intracellular pH or membrane potential; however, we demonstrate here that bicarbonate directly stimulates mammalian soluble adenylyl cyclase (sAC) activity in vivo and in vitro in a pH-independent manner. sAC is most similar to adenylyl cyclases from cyanobacteria, and bicarbonate regulation of cyclase activity is conserved in these early forms of life. sAC is also expressed in other bicarbonate-responsive tissues, which suggests that bicarbonate regulation of cAMP signaling plays a fundamental role in many biological systems.

A new family of adenylyl cyclase genes in the male germline of Drosophila melanogaster.

We describe the cloning and characterization of a new gene family of adenylyl cyclase related genes in Drosophila. The five adenylyl cyclase-like genes that define this family are clearly distinct from previously known adenylyl cyclases. One member forms a unique locus on chromosome 3 whereas the other four members form a tightly clustered, tandemly repeated array on chromosome 2. The genes on chromosome 2 are transcribed in the male germline in a doublesex dependent manner and are expressed in postmitotic, meiotic, and early differentiating sperm. These genes therefore provide the first evidence for a role for the cAMP signaling pathway in Drosophila spermatogenesis. Expression from this locus is under the control of the always early, cannonball, meiosis arrest, and spermatocyte arrest genes that are required for the G2/M transition of meiosis I during spermatogenesis, implying a mechanism for the coordination of differentiation and proliferation. Evidence is also provided for positive selection at the locus on chromosome 2 which suggests this gene family is actively evolving and may play a novel role in spermatogenesis.

Restricted expression of a truncated adenylyl cyclase in the cephalic furrow of Drosophila melanogaster.

We have identified a novel isoform of adenylyl cyclase, DAC78C, in Drosophila melanogaster that encodes two structurally distinct proteins in a developmentally restricted manner. The protein corresponding to one transcript is potently activated by G protein and protein kinase C and is expressed ubiquitously. The protein corresponding to the second transcript is expressed in a dynamic pattern in gastrulation stage embryos; it is restricted to the cephalic furrow and dorsal transverse folds, active regions of cell movement of unknown function in the Drosophila embryo. We propose that DAC78C and the cAMP pathway play an important role in directing these morphogenetic movements, and that this gene may provide clues to the functional significance of these structures in gastrulation.

Cytosolic adenylyl cyclase defines a unique signaling molecule in mammals.

Mammals have nine differentially regulated isoforms of G protein-responsive transmembrane-spanning adenylyl cyclases. We now describe the existence of a distinct class of mammalian adenylyl cyclase that is soluble and insensitive to G protein or Forskolin regulation. Northern analysis indicates the gene encoding soluble adenylyl cyclase (sAC) is preferentially expressed in testis. As purified from rat testis cytosol, the active form of sAC appears to be a fragment derived from the full-length protein, suggesting a proteolytic mechanism for sAC activation. The two presumptive catalytic domains of sAC are closely related to cyanobacterial adenylyl cyclases, providing an evolutionary link between bacterial and mammalian signaling molecules.

Cloning and characterization of a Drosophila adenylyl cyclase homologous to mammalian type IX.

A novel Drosophila adenylyl cyclase (AC) was identified by PCR using degenerate primers specific for the known metazoan ACs. The full-length cDNA predicts a protein displaying significant sequence homology with mammalian Type IX AC (AC9). The abundance and size of the message for the Drosophila AC9 homolog (DAC9) changes through development. Biochemical analysis of DAC9 confirms it encodes a functional enzyme which can be activated by forskolin or G protein. Together with the Drosophila Type I AC homolog encoded by the learning and memory gene, rutabaga, the molecular identification of DAC9 demonstrates there is a family of Drosophila AC isoforms reflecting at least part of the diversity of mammalian AC isoforms.