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Pathways of adenylate cyclase stimulation

Receptor-mediated activation of adenylyl cyclase (ACA) in Dictyostelium requires CRAC protein. Upon translocation to the membrane, this pleckstrin homology (PH) domain protein stimulates ACA and thereby mediates developmental aggregation. Thus CRAC acts upstream of ACA. CRAC may also have roles later in development since CRAC-null cells can respond to chemotactic signals and participate in developmental aggregation when admixed with wild-type cells, but they do not complete development within such chimeras. To test whether the role of CRAC in postaggregative development is related to the activation of ACA, chemotactic aggregation was bypassed in CRAC-null cells by activating the cAMP-dependent protein kinase (PKA). While such strains form mounds, they do not complete fruiting body morphogenesis or form spores. Expression of CRAC in the prespore cells of these strains rescues sporulation and fruiting body formation. This later function of CRAC does not appear to require its PH domain since the C-terminal portion of the protein (CRAC-DeltaPH) can substitute for full-length CRAC in promoting spore cell formation and morphogenesis. No detectable ACA activation is observed in any of the CRAC-null strains rescued by PKA activation and expression of CRAC-DeltaPH. The development of CRAC-null ACA-null double mutants can be rescued by the activation of PKA, together with the expression of CRAC-DeltaPH. Thus, there appears to be a required function for CRAC in postaggregative development that is independent of its previously described function in the ACA activation pathway. How CRAC executes its function in prespore cell differentiation and morphogenesis during postaggregative development remains unknown. Possibilities include the participation of CRAC in a PKA-independent pathway required for sporulation, or the additional utilization of CRAC in the PKA pathway 'downstream' of PKA-C (Wang, 1999).

The type I adenylyl cyclase is directly stimulated by Ca2+ and calmodulin in vivo. In this study, sensitivity of type I adenylyl cyclase to beta-adrenergic agonists or glucagon was examined when intracellular Ca2+ was elevated by Ca2+ ionophore or carbachol. Although previous studies have shown that this enzyme can be directly stimulated by activated Gs in vitro, it is not stimulated by Gs-coupled receptors in vivo. However, the enzyme is stimulated by Gs-coupled receptors in vivo when it is activated by intracellular Ca2+. For example, the Ca2+ ionophore A23187 stimulates the enzyme 3 fold; isoproterenol alone does not stimulate the enzyme, but the combination of the two stimulates type I adenylyl cyclase 13-fold in vivo. Similarly, 500 nM glucagon alone does not stimulate the enzyme but the combination of A23187 and glucagon activates the enzyme 90 fold. Synergistic stimulation of type I adenylyl cyclase activity is also obtained with combinations of carbachol and isoproterenol or glucagon. This phenomenon is not observed with a mutant enzyme that is insensitive to Ca2+ and calmodulin, suggesting that conformational changes caused by binding of calmodulin to the type I adenylyl cyclase enhance binding or coupling to activated Gs. These data illustrate that this adenylyl cyclase can couple Ca2+ and neurotransmitter signals to generate optimal cAMP levels, a property of the enzyme that may be important for its role in learning and memory in mammals (Wayman, 1994).

The second messengers cAMP and inositol-1,4,5-triphosphate have been implicated in the olfaction of various species. The odorant-induced cGMP response was investigated using cilia preparations and olfactory primary cultures. Odorants cause a delayed and sustained elevation of cGMP. A component of this cGMP response is attributable to the activation of one of two kinetically distinct cilial receptor guanylyl cyclases by calcium and a guanylyl cyclase-activating protein (GCAP). cGMP thus formed serves to augment the cAMP signal in a cGMP-dependent protein kinase (PKG) manner by direct activation of adenylate cyclase. cAMP, in turn, activates cAMP-dependent protein kinase (PKA) which can function in turn to negatively regulate guanylyl cyclase, thus limiting the cGMP signal. These data demonstrate the existence of a regulatory loop in which cGMP can augment a cAMP signal, and in turn cAMP negatively regulates cGMP production via PKA. Thus, a small, localized, odorant-induced cAMP response may be amplified to modulate downstream transduction enzymes or transcriptional events (Moon, 1998).

Cellular content of cAMP generated by activation of adenylylcyclase (AC) is a key determinant of functional responsiveness in the heart and other tissues. Two hypotheses have been tested regarding the relationship between AC content and beta-adrenergic receptor (betaAR)-mediated signal transduction in cardiac myocytes: (1) that AC content limits adrenergic signal transduction, and (2) that increased AC, independent of (betaAR) number and G-protein content, yields a proportional increase in betaAR-mediated transmembrane signaling. Recombinant adenovirus was used to increase AC isoform VI (ACVI) expression in neonatal cardiac myocytes. Cells that overexpress ACVI respond to agonist stimulation with marked increases in cAMP production in proportion to protein expressed. In parallel experiments performed on cells transfected with lacZ (control) or ACVI, [3H]forskolin binding (used to assess AC protein expression) is amplified 6-fold, while betaAR-stimulated cAMP production from these cells is increased 7-fold. No changes in betaAR number, or in the heterotrimeric GTP-binding proteins, Galphas or Galphai2, are observed. Previous studies indicate that increased cardiac expression of betaAR or Galphas does not yield proportional increases in transmembrane adrenergic signaling. In contrast, the current data demonstrate that increased ACVI expression provides a proportional increase in beta-adrenergic signal transduction. These results show that the amount of AC sets a limit on transmembrane beta-adrenergic signaling. It is speculated that similar functional responses are possible in other cell types in which AC plays an important physiological role (Gao, 1998).

The alpha subunit (Gsalpha) of the stimulatory heterotrimeric guanosine triphosphate binding protein [G protein) activates all isoforms of mammalian adenylyl cyclase. Adenylyl cyclase (Type V) and its subdomains, which interact with Gsalpha, promote inactivation of the G protein by increasing its guanosine triphosphatase (GTPase) activity. Adenylyl cyclase and its subdomains also augment the receptor-mediated activation of heterotrimeric Gs and thereby facilitate the rapid onset of signaling. These findings demonstrate that adenylyl cyclase functions as a GTPase activating protein (GAP) for the monomeric Gsalpha and enhances the GTP/GDP exchange factor (GEF) activity of receptors (Scholich, 1999).

Hormonal signals activate trimeric G proteins by substituting GTP for GDP bound to the G protein alpha subunit (Galpha), thereby generating two potential signaling molecules, Galpha-GTP and free Gbetagamma. A dominant negative Galpha mutation was created. A mutant alpha subunit is described that is designed to inhibit receptor-mediated hormonal activation of Gs, the stimulatory regulator of adenylyl cyclase. To construct this mutant, three separate mutations chosen because they impair alphas function in complementary ways were introduced into the alpha subunit (alphas) of Gs: the A366S mutant reduces affinity of alphas for binding GDP, whereas the G226A and E268A mutations impair the protein's ability to bind GTP and to assume an active conformation. The triple mutant robustly inhibits (by up to 80%) Gs-dependent hormonal stimulation of adenylyl cyclase in cultured cells. Inhibition is selective in that it does not affect cellular responses to expression of a constitutively active alphas mutant (alphas-R201C) or to agonists for receptors that activate Gq or Gi. This alphas triple mutant and cognate Galpha mutants should provide specific tools for dissection of G protein-mediated signals in cultured cells and transgenic animals (Iiri, 1999).

In the rat olfactory bulb, activation of opioid receptors enhances basal adenylyl cyclase activity and potentiates enzyme stimulation by Gs-coupled neurotransmitter receptors in a pertussis toxin-sensitive manner. The involvement of G protein betagamma subunits was studied by examining the effects of betagamma scavengers and exogenously added betagamma subunits of transducin [betagamma(t)]. The QEHA fragment of type II adenylyl cyclase (50 microM), a peptide that binds to and inactivates betagamma, inhibits the maximal stimulation of adenylyl cyclase activity elicited by Leu-enkephalin (Leu-enk) by about 50%. Similarly, the GDP-bound form of the alpha subunit of transducin (5 nM-1.5 microM), another betagamma scavenger, reduces both the opioid stimulation of basal adenylyl cyclase activity and the potentiation of vasoactive intestinal peptide-stimulated enzyme activity. Under the same experimental conditions, these agents fail to affect the stimulation of the enzyme activity elicited by activation of beta-adrenergic receptors with 1-isoproterenol. Moreover, the addition of betagamma(t)(400 nM) stimulates basal adenylyl cyclase by 80%, and this effect is not additive with that produced by Leu-enk. The data indicate that opioids enhance adenylyl cyclase activity in rat olfactory bulb by promoting the release of betagamma subunits from pertussis toxin-sensitive G proteins Gi/Go (Olianas, 1999).

Circadian functions of the suprachiasmatic nuclei (SCN) are influenced by cyclic AMP (cAMP). Adenylyl cyclase type II (AC-II) is a cAMP-generating enzyme that, in the context of activation by Gsalpha, is further stimulated by protein kinase C or G protein betagamma subunits. Using in situ hybridization a biphasic variation in AC-II mRNA was found within the rat SCN during the light-dark cycle (peaks at Zeitgeber time 6 and 18) and also in constant darkness (peaks at circadian time 2 and 14). The cingulate cortex shows no such variation. These findings suggest that circadian changes in AC-II expression may be pertinent to the rhythmic functions of the SCN (Cagampang, 1998).

The neurochemical basis was investigated for predominance of stimulatory mu-opioid signaling in guinea pig longitudinal muscle/myenteric plexus (LMMP) preparations after chronic in vivo morphine exposure. In a dose dependent manner, recombinant Gsalpha (rGsalpha) stimulates adenylyl cyclase (AC) activity in LMMP membranes obtained from opioid naive as well as tolerant LMMP tissue. However, the magnitude of the increase is significantly greater in the latter than in the former. The Gbetagamma blocking peptide QEHA essentially abolishes stimulation by rGsalpha in LMMP membranes obtained from both opioid naive and tolerant animals. Interestingly, after partial blockade by lower QEHA concentrations, the incremental AC stimulation by rGsalpha in tolerant LMMP membranes is no longer observed, indicating augmented Gbetagamma stimulatory responsiveness. Concomitant changes in the content of AC isoform protein are consistent with these biochemical observations. After chronic systemic morphine, AC protein is augmented significantly (56%). This increment is most likely to be composed of AC isoforms that are stimulated by Gbetagamma. This is the first demonstration in a complex mammalian tissue that persistent activation of opioid receptors results in augmented Gbetagamma/Gsalpha AC stimulatory interactiveness (Chakrabarti, 1998).

The beta2-adrenoceptor (beta2AR) activates the G-protein Gsalpha to stimulate adenylate cyclase (AC). Fusion of the beta2AR C-terminus to the N-terminus of Gsalpha (producing beta2ARGsalpha) markedly increases the efficiency of receptor/G-protein coupling, as compared with the non-fused state. This increase in coupling efficiency can be attributed to the physical proximity of receptor and G-protein. To determine the optimal length for the tether between receptor and G-protein, fusion proteins were constructed from which 26 [beta2AR(Delta26)Gsalpha] or 70 [beta2AR(Delta70)Gsalpha] residues of the beta2AR C-terminus had been deleted and the properties of these fusion proteins were compared with the beta2ARGsalpha. Compared with beta2ARGsalpha, basal and agonist-stimulated GTP hydrolysis is markedly decreased in beta2AR(Delta70)Gsalpha, whereas the effect of the deletion on binding of guanosine 5'-[gamma-thio]triphosphate (GTP[S]) was relatively small. Surprisingly, deletions do not alter the efficiency of coupling of the beta2AR to Gsalpha as assessed by GTP[S]-sensitive high-affinity agonist binding. Moreover, basal and ligand-regulated AC activities in membranes expressing beta2AR(Delta70)Gsalpha and beta2AR(Delta26)Gsalpha are higher than in membranes expressing beta2ARGsalpha. These findings suggest that restricting the mobility of Gsalpha relative to the beta2AR results in a decrease in G-protein inactivation by GTP hydrolysis and thereby enhances activation of AC (Wenzel-Seifert, 1998).

Posttranslational modification of Ras protein has been shown to be critical for interaction with its effector molecules, including S. cerevisiae adenylyl cyclase. Lipid modification, specifically farnesylation, which accompanies the membrane attachment of Ras, stimulates the Ras-dependent activaion of yeast adenylyl cyclase. The leucine rich repeat in the middle repetitive domain of adenyl cyclase interacts with Ras. In this study, a reconstituted system was used with purified adenylyl cyclase and Ras proteins carrying various degrees of the modification to show that the posttranslational modification, especially the farnesylation step, is responsible for 5- to 10-fold increase in Ras-dependent activation of adenylyl cyclase activity even though it has no significant effect on binding of Ras and adenyl cyclase to each other. The stimulatory effect of farnesylation is found to depend on the association of adenylyl cyclase with 70-kDa adenylyl cyclase-associated protein (CAP), which is known to be required for proper in vivo response of adenylyl cyclase to Ras protein, by comparing the levels of Ras-dependent activation of purified adenylyl cyclase with and without bound CAP. The region of CAP required for this effect is mapped to CAP's N-terminal segment of 168 amino acid residues, which coincides with the region required for the in vivo effect. Furthermore, the stimulatory effect is successfully reconstituted by in vitro association of CAP with the purified adenylyl cyclase. These results indicate that the association of adenylyl cyclase with CAP is responsible for the stimulatory effect of posttranslational modification of Ras on its activity and that this may be the mechanism underlying its requirement for the proper in vivo cyclic AMP response (Shima, 1997).

A Saccharomyces cerevisiae gene encoding adenylate cyclase has been analyzed by deletion and insertion mutagenesis to localize regions required for activation by the S. cerevisiae RAS2 protein. The NH2-terminal 657 amino acids were found to be dispensable for the activation. However, almost all 2-amino acid insertions in the middle 600 residues (comprising leucine-rich repeats) and deletions in the COOH-terminal 66 residues completely abolish activation by the RAS2 protein, whereas insertion mutations in the other regions generally have no effect. Chimeric adenylate cyclases were constructed by swapping the upstream and downstream portions surrounding the catalytic domains between the S. cerevisiae and Schizosaccharomyces pombe adenylate cyclases and examined for activation by the RAS2 protein. The fusion containing both the NH2-terminal 1600 residues and the COOH-terminal 66 residues of the S. cerevisiae cyclase render the catalytic domain of the S. pombe cyclase, which otherwise did not respond to RAS proteins, activatable by the RAS2 protein. Thus the leucine-rich repeats and the COOH terminus of the Sa. cerevisiae adenylate cyclase appear to be required for interaction with RAS proteins (Suzuki, 1998).

Short-term behavioral sensitization of the gill-withdrawal reflex after tail stimuli in Aplysia leads to an enhancement of the connections between sensory and motor neurons of this reflex. Both behavioral sensitization and enhancement of the connection between sensory and motor neurons are importantly mediated by serotonin. Serotonin activates two types of receptors in the sensory neurons, one of which is coupled to the cAMP/protein kinase A (PKA) pathway and the other to the inositol triphosphate/protein kinase C (PKC) pathway. A genetic approach to assessing the isolated contribution of the PKA pathway to short-term facilitation is described. An octopamine receptor gene, Ap oa1, that couples selectively to the cAMP/PKA pathway has been cloned from Aplysia. Ap oa1 contains an ORF of 394 amino acids. The amino acid sequences of Ap oa1 from two Aplysia species are nearly identical (94% homologous). Sequence identity with other OA/tyramine receptors is 36%-47% in transmembrane domains and 24%-30% in the entire amino acid sequences. Furthermore, the lengths of second extracellular and third intracellular loops and C-terminal tail are very different from those of other OA/tyramine receptors. Specifically, Ap oa1 has a rather short third intracellular loop region. Interestingly, however, these hypervariable loops of Ap oa1 are comparable in length to those of mammalian beta2-adrenergic receptors that are well known to couple to Gs protein. These features indicate that Ap oa1 may represent a new class of OA/tyramine receptors. This receptor has been ectopically expressed in Aplysia sensory neurons of the pleural ganglia, where it is not normally expressed. Activation of this receptor by octopamine stimulates all four presynaptic events involved in short-term synaptic facilitation that are normally produced by serotonin: (1) membrane depolarization; (2) increased membrane excitability; (3) increased spike duration; and (4) presynaptic facilitation. These results indicate that the cAMP/PKA pathway alone is sufficient to produce all the features of presynaptic facilitation (Chang, 2000).

Targets of adenylate cyclase signaling

The cAMP signaling system has been postulated to be involved in the embryogenesis of many animal species, however, little is known about its role in embryonic axis formation in vertebrates. In this study, the role of the cAMP signaling pathway in patterning the body plan of the Xenopus embryo was investigated by expressing and activating the exogenous human 5-hydroxytryptamine type 1a receptor (5-HT1a R), which inhibits adenylyl cyclase through inhibitory G-protein in embryos in a spatially- and temporally-controlled manner. In embryos, ventral, but not dorsal expression and stimulation of this receptor during blastula and gastrula stages induces a secondary axis. The secondary axis induced by ventral stimulation of the 5-HT1a R is usually incomplete, with no head structure or much reduced anterior structure. At the molecular level, 5-HT1a R stimulation induces expression of the dorsal mesoderm marker genes, and down-regulates expression of the ventral markers but has no effect on expression of the pan mesodermal marker gene in ventral marginal zone explants. Stimulation of 5-HT1a R at stage 8 induces the expressions of noggin and chordin, but not siamois in the gastrula (stage 11.5) VMZ explants. In contrast, the expression of ventral-specific marker, BMP-4, is significantly reduced in the stage 11.5 VMZs that had been stimulated with 5-HT. The expressions of Xvent-2 and PV.1 are also reduced but not as greatly as BMP-4. The expression of Xbra, a generic mesoderm marker, did not change in both DMZ and VMZ explants, indicating that the formation of mesoderm is not affected by the receptor activation. Ventral expression and stimulation of the receptor partially restores dorsal axis of UV-irradiated axis deficient embryos. Finally, the total mass of cAMP differs between dorsal and ventral regions of blastula and gastrula embryos and this is regulated in a temporally-specific manner. These results suggest that the cAMP signaling system may be involved in the transduction of ventral signals in the patterning of early embryos (Kim, 1999).

Adenylyl cyclase (AC) modulation of vesicular cycling was visualized at cultured cerebellar granule cell synapses using the sequential uptake of antibodies directed against the intraluminal domain of synaptotagmin I. Vesicle recycling due to spontaneous transmitter release in the absence of action potentials is increased by the AC/protein kinase A (PKA) activators forskolin and CPT-cAMP. These effects are blocked by the PKA inhibitor Rp-cAMPs. Cyclic AMP elevation also induces new cycling at previously silent sites. Activation of L-AP4-sensitive mGluR reduces the cAMP/PKA enhancement at preexisting synapses downstream of both AC and calcium channels. Modulation of the turnover and the number of vesicular release sites provide one mechanism that may underlie cAMP-dependent cerebellar long-term potentiation (Chavis, 1998).

Co-opting the unfolded protein response to elicit olfactory receptor feedback: Adenylyl cyclase 3 relieves the UPR

Olfactory receptor (OR) expression in mammals requires the transcriptional activation of 1 out of 1,000s of OR alleles and a feedback signal that preserves this transcriptional choice. The mechanism by which olfactory sensory neurons (OSNs) detect ORs to signal to the nucleus remains elusive. This study shows that OR proteins generate this feedback by activating the unfolded protein response (UPR). OR expression induces Perk-mediated phosphorylation of the translation initiation factor eif2α causing selective translation of activating transcription factor 5 (ATF5, a member of the ATF/CREB family of transcription factors). ATF5 induces the transcription of adenylyl cyclase 3 (Adcy3), which relieves the UPR. These data provide a role for the UPR in defining neuronal identity and cell fate commitment and support a two-step model for the feedback signal: (1) OR protein, as a stress stimulus, alters the translational landscape of the OSN and induces Adcy3 expression; (2), Adcy3 relieves that stress, restores global translation, and makes OR choice permanent (Dalton, 2014).

The mammalian main olfactory epithelium (MOE) is characterized by extreme diversity of olfactory sensory neurons (OSNs), each defined by the expression of a single olfactory receptor (OR) allele. In the mouse, the expressed OR is selected, in a monogenic, monoallelic and seemingly stochastic fashion from a repertoire of more than 1,000 genes. Heterochromatic silencing of all ORs, at a developmental stage that precedes their transcriptional activation and aggregation of the silent OR genes in distinct, heterochromatic nuclear foci assure their efficient repression and set the stage for the transcriptional activation of a single OR allele. Indeed, the active allele in each OSN is spatially separated from the repressed OR loci, interacts with the H enhancer, and carries activating histone marks, suggesting that selective desilencing of a single allele and relocation to a transcriptionally competent nuclear territory is the basis of OR activation. Lysine demethylase 1 (LSD1) plays a key role in this epigenetic switch because it catalyzes the removal of repressive lysine 9 methyl marks from histone H3 on the chosen OR allele). Importantly, the subsequent downregulation of LSD1 in response to OR expression prevents the desilencing of additional ORs and stabilizes the expression of the activated allele revealing that LSD1 is the target of an OR-elicited feedback that locks OR choice for the life of the neuron (Dalton, 2014).

The observation that the expression of OR protein causes the downregulation of LSD1 and, therefore, the stabilization of OR choice poses significant questions regarding the cellular mechanisms that elicit this feedback. OR gene activation induces expression of Adenylyl Cyclase 3 (Adcy3), which then signals for the downregulation of LSD1, providing a link between OR and LSD1 expression. However, these results do not explain how an OR is detected by the neuron in the first place; Adcy3 plays a central role in the stabilization of OR choice, however, it is unlikely to be a 'first responder' or initiator of the feedback because its expression relies upon OR expression. Therefore, a central question toward the understanding of the OR feedback signal is how ORs are detected by the OSN and how this detection leads to the stable expression of Adcy3 protein. Because stabilization of OR choice requires the timely downregulation of LSD1, detecting and vetting the OR protein after targeting to the cell membrane may be too slow because GPCR targeting requires an elaborate series of posttranslational modifications and trafficking through the endoplasmatic reticulum (ER) and Golgi. Thus, protein quality control pathways placed in the first relay station of OR translation and processing, the ER, would rapidly link the onset of OR expression to Adcy3 transcription and, consequently, could provide a kinetic advantage for the stabilization of OR choice (Dalton, 2014).

In the ER, a highly conserved protein quality control pathway, the unfolded protein response (UPR), acts to homeostatically adjust the ER environment upon detection of unfolded proteins. These adjustments include transcriptional induction of chaperones, acting to increase ER protein folding capacity, and inhibition of translation initiation, aiming to decrease ER load. The inhibition of translation initiation occurs downstream of the ER-resident kinase Perk, which in response to detection of unfolded proteins phosphorylates the translation initiation factor eif2a. This serves to limit the availability of tRNAmet, resulting in a general inability of ribosomes to initiate translation. Paradoxically, a small number of mostly stress-responsive mRNAs are preferentially translated under these conditions. This can be explained by the presence of inhibitory upstream open reading frames in their 5' UTRs, which are selectively bypassed when tRNAmet becomes limiting, slowing ribosome assembly. Activating transcription factor 4 (ATF4), which is selectively translated under these conditions in many cell types, induces transcriptional changes that contribute to the clearance from the ER of misfolded proteins or to the adaptation of the ER to increased protein load (Dalton, 2014).

Seeking to reveal the mechanistic outline of the OR feedback process, this study tests the hypothesis that UPR components detect OR proteins in the ER and transmit this information to the nucleus. The experiments show that OR expression activates Perk in the neuronal ER, which phosphorylates the translation initiation factor eif2a, leading to selective and transient translation of Activating Transcription Factor 5 (ATF5), a paralogue to ATF4 that is highly transcribed in the MOE. Translation of the nuclear form of Atf5 induces the transcription of Adcy3, which relieves the UPR, restores global translation, promotes OSN differentiation, and stabilizes the expression of the chosen OR. PERK and ATF5 KO mice, as well as eif2a phosphorylation mutants, exhibit unstable OR expression and OSN maturation deficits, whereas pharmacological induction of the UPR or transgenic expression of nuclear ATF5 can bypass the lack of OR expression or the blockage of this signaling pathway. These data solve a long-lasting puzzle in OR regulation and provide a use for the UPR in neuronal differentiation and cell fate commitment that is likely applicable to other neurodevelopmental processes (Dalton, 2014).

Adenylate cyclase and cell cycle

The intracellular levels of cAMP play a critical role in the meiotic arrest of mammalian oocytes. However, it is debated whether this second messenger is produced endogenously by the oocytes or is maintained at levels inhibitory to meiotic resumption via diffusion from somatic cells. Adenylyl cyclase genes and corresponding proteins are expressed in rodent oocytes. The mRNA coding for the AC3 isoform of adenylyl cyclase was detected in rat and mouse oocytes by RT-PCR and by in situ hybridization. The expression of AC3 protein was confirmed by immunocytochemistry and immunofluorescence analysis in oocytes in situ. Cyclic AMP accumulation in denuded oocytes is increased by incubation with forskolin, and this stimulation is abolished by increasing intraoocyte Ca2+ with the ionophore A23187. The Ca2+ effects are reversed by an inhibitor of Ca2+, calmodulin-dependent kinase II. These regulations of cAMP levels indicate that the major cyclase that produces cAMP in the rat oocyte has properties identical to those of recombinant or endogenous AC3 expressed in somatic cells. Furthermore, mouse oocytes deficient in AC3 show signs of a defect in meiotic arrest in vivo and accelerated spontaneous maturation in vitro. Collectively, these data provide evidence that an adenylyl cyclase is functional in rodent oocytes and that its activity is involved in the control of oocyte meiotic arrest (Horner, 2003).

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rutabaga: Biological Overview | Regulation | Developmental Biology | Effects of Mutation | References

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