Protein kinase C interferes with Ni-mediated inhibition of human platelet adenylate cyclase

Addition of phorbol ester-activated, partially purified protein kinase C to membranes of human platelets had no effect on forskolin stimulation of the adenylate cyclase and increased stimulation by prostaglandin E1 only at high GTP concentrations by preventing inhibition by GTP. Hormonal inhibition of the platelet adenylate cyclase by epinephrine was eliminated or largely impaired. At low GTP concentrations, epinephrine even caused a small increase in cyclase activity. The data suggest that activated protein kinase C interferes with GTP- and hormone-induced adenylate cyclase inhibition probably by phosphorylating the inhibitory guanine nucleotide-binding regulatory component Ni.     

Activation of adipocyte adenylate cyclase by protein kinase C

Adenylate cyclase activity in purified rat adipocyte membranes is stimulated by the calcium- and phospholipid-dependent enzyme protein kinase C. Over the concentration range of 100-1000 milliunits/ml, both highly purified (approximately 3000 units/mg of protein) protein kinase C from rat brain and partially purified (14 units/mg of protein) protein kinase C from guinea pig pancreas stimulate cyclase activity. The actions of both protein kinase C preparations on adenylate cyclase activity are dependent on added calcium, which is effective at concentrations less than 10 microM. Exogenous phospholipids are not required for stimulation of adenylate cyclase by protein kinase C; but, under typical cyclase assay conditions, the adipocyte membranes satisfy the lipid requirement for protein kinase C phosphorylation of histone. The tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate enhances the kinase action on cyclase, and the phorbol ester is effective at concentrations equimolar with the kinase (less than 10 nM). With the brain protein kinase C, 12-O-tetradecanoylphorbol-13-acetate effects are especially evident at limiting calcium concentrations. Inhibitors of protein kinase C activity, such as chlorpromazine, palmitoylcarnitine, and polymyxin B, inhibit selectively that adenylate cyclase activity which is stimulated by protein kinase C plus calcium. It is concluded that protein kinase C acts directly on the adipocyte adenylate cyclase system.     

Protein kinase C and a viral K-RAS protein cooperatively enhance the response of adenylate cyclase to stimulators

The protein kinase C stimulator TPA (12-O-tetradecanoyl phorbol-13-acetate) enhanced the responsiveness of adenylate cyclase to IPR (isoproterenol) and PGE1 (prostaglandin E1) in quiescent tsKSV-NRK cells at the nonpermissive 41 degrees C. Reactivating the thermolabile mitogenic/oncogenic K-ras protein in tsKSV-NRK cells by dropping the temperature to 36 degrees C also enhanced the responsiveness of adenylate cyclase to IPR and PGE1. The enhancement was transient and peaked at 6 hours after the temperature shift. This enhanced responsiveness was specifically due to the reactivated viral K-ras protein rather than the temperature shift because the same temperature shift did not affect adenylate cyclase responsiveness in uninfected NRK cells, nor was it a result of the mitogenic stimulus since reacting the mitogenic pp60v-src protein in tsASV-NRK cells did not affect adenylate cyclase responsiveness. The increased responsiveness of adenylate cyclase at 6 hours after the temperature shift was not a result of elevated membrane-associated PKC activity. However, the reactivated viral K-ras protein strongly increased the stimulability of membrane-associated PKC by TPA and it further increased TPA's ability to enhance the responsiveness of adenylate cyclase to IPR and PGE1. Thus, a viral K-ras protein and membrane-associated protein kinase C can cooperate to increase the responsiveness of adenylate cyclase to agonists.     

Modulation of adenylate cyclase in human keratinocytes by protein kinase C

Adenylate cyclase (ATP-pyrophosphate lyase (cyclizing); EC 4.6.1.1) in the human keratinocyte cell line SCC 12F was potentiated by 12-O-tetradecanoyl-phorbol-13-acetate (TPA), phorbol-12,13-diacetate, and 1,2-dioctanoylglycerol. Keratinocytes exposed to TPA showed a 2-fold enhancement of adenylate cyclase activity when assayed in the presence of isoproterenol or GTP. The half-maximal effective concentration (EC50) for both isoproterenol and GTP were unaltered by TPA treatment of the cells. Basal adenylate cyclase activity in membranes from TPA-treated cultures was also increased 2-fold relative to activity in control membranes. Potentiation of adenylate cyclase activity was dependent on the concentration of TPA to which the keratinocytes were exposed (EC50 for TPA = 3 nM). TPA actions on adenylate cyclase were maximal after 15 min of incubation of the cells with the compound, correlating well with the time course of translocation of protein kinase C (Ca2+/phospholipid-dependent enzyme) from cytosol to membrane. The action of cholera toxin on adenylate cyclase was additive with TPA. In contrast, pertussis toxin actions on adenylate cyclase were not additive with TPA. Treatment of control cells with pertussis toxin activated adenylate cyclase 1.5-fold, whereas cells exposed to pertussis toxin for 6 h followed by TPA for 15 min showed the same 2-fold increase in adenylate cyclase activity as observed in membranes from cells exposed to TPA without prior exposure to pertussis toxin. Pertussis toxin catalyzed ADP-ribosylation was increased 2-fold in membranes from SCC 12F cells exposed to TPA, indicating an increase in the alpha beta gamma form of Gi. These data suggest that exposure of human keratinocytes to phorbol esters increases adenylate cyclase activity by a protein kinase C-mediated increase in the heterotrimeric alpha beta gamma form of Gi resulting in decreased inhibition of basal adenylate cyclase activity.     

Protein kinase C activity can desensitize the gonadotropin-responsive adenylate cyclase in Leydig tumor cells. But hCG-induced desensitization does not involve protein kinase C activation

The murine Leydig tumor cell line, MLTC-1, contains a gonadotropin receptor-coupled adenylate cyclase. Although the binding of human choriogonadotropin (hCG) initially causes cells to accumulate cAMP, in time, the response to hCG is attenuated by desensitization. Treating intact cells with the tumor promoter, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), or with diacylglycerol also causes desensitization of the hCG response. These compounds are activators of calcium/phospholipid-dependent protein kinase (PKC). Treating MLTC-1 cells with TPA or dioctanoylglycerol increased the portion of PKC in the cell membrane fraction. This phenomenon is associated with activation of PKC. Treating isolated membranes with purified PKC desensitize the hCG response. Thus, desensitization caused by TPA or dioctanoylglycerol is probably mediated by PKC. PKC is normally activated when phosphoinositides are metabolized to diacylglycerol and inositol phosphates. There was no significant accumulation of inositol phosphates when cells were treated with hCG. hCG did not increase the portion of PKC in the cell membrane fraction. However, hCG could desensitize isolated membranes, but TPA could not. We conclude that although protein kinase C activity can desensitize the gonadotropin response, hCG does not cause desensitization by activating PKC. The implications of this observation are discussed.     

Protein kinase C and receptor kinase gene expression in olfactory receptor neurons

Recent biochemical evidence indicates that protein kinase C (PKC) and G-protein-coupled receptor kinases (GRKs) are involved in olfactory signal termination and desensitization. The polymerase chain reaction (PCR) was used to investigate the expression of PKC and GRK genes in olfactory tissue and in isolated olfactory receptor neurons from channel catfish (Ictalurus punctatus). Sequence analysis of cloned PKC PCR products showed that the α, β, δ, ϵ, and τ isotypes were expressed in olfactory tissue. Sequence analysis of PCR products obtained from isolated olfactory receptor neurons showed that PKCβ and PKCδ were expressed in the receptor cells. A 600-bp GRK PCR product was obtained from isolated olfactory neurons that shared 86% and 92% amino acid sequence identity to the mammalian β-adrenergic receptor kinase gene products βARK1 and βARK2, respectively. Go6976, a specific inhibitor of calcium-regulated PKC activity, completely inhibited odorant-stimulated PKC activity in isolated olfactory cilia. This result suggested that odorant-stimulated PKC activity is mediated by the calcium-sensitive PKCβ isotype. Taken together, these results are consistent with the conclusion that PKCβ and βARK mediate odorant receptor phosphorylation and olfactory signal termination. © 1997 John Wiley & Sons, Inc. J Neurobiol 33: 387–394, 1997     

Activation of protein kinase C modulates the adenylate cyclase effector system of B-lymphocytes

Antibodies to surface immunoglobulins activate inositol phospholipid hydrolysis in B-lymphocytes, but very little is known concerning their effects on cAMP levels. In other cells, products from the hydrolysis of phosphatidylinositol 4,5-bisphosphate can increase and/or potentiate cAMP accumulation. In this study we have examined whether goat anti-mouse IgM (mu-chain-specific) stimulates and/or potentiates increases in the cAMP levels of splenocytes from athymic nude mice. Goat anti-mouse IgM, by itself, stimulated a 60% increase in cAMP within 2 min. Pretreating the cell suspensions at 37 degrees C with anti-IgM produced opposite effects on the forskolin- and prostaglandin E1 (PGE1)-induced increase in cAMP. Anti-IgM (25 micrograms/ml) potentiated the rise in cAMP induced by 100 microM forskolin 76%, but it decreased the response to 50 nM PGE1 by 30%. Direct activation of protein kinase C (Ca2+/phospholipid-dependent enzyme) by 12-O-tetradecanoylphorbol 13-acetate and/or sn-1,2-dioctanoylglycerol resulted in a similar pattern of responses. A 3-min preincubation with 97 nM 12-O-tetradecanoylphorbol 13-acetate potentiated the forskolin-induced response from 1.7 +/- 0.1 to 4.3 +/- 0.6 pmol of cAMP/10(6) cells but reduced the PGE1 response from 0.98 +/- 0.06 to 0.51 +/- 0.03 pmol of cAMP/10(6) cells. Similarly, preincubating the cells for 3 min with 5 microM sn-1,2-dioctanoylglycerol increased the forskolin response from 1.7 +/- 0.1 to 5.1 +/- 0.2 pmol of cAMP/10(6) cells but reduced the response to PGE1 from 1.15 +/- 0.03 to 0.75 +/- 0.04 pmol of cAMP/10(6) cells. Thus, activation of protein kinase C by hydrolysis products of inositol phospholipids, 12-O-tetradecanoylphorbol 13-acetate, or exogenous diacylglycerols modified adenylate cyclase itself and sites upstream of adenylate cyclase such as the receptor or G proteins coupling the receptor to the cyclase. Furthermore, modification of the PGE1 response by anti-IgM provides a mechanism by which antigen can differentially regulate T- and B-cells responding to macrophage-produced prostaglandins during an immune response.     

Protein kinase C catalyzes phosphorylation of guanylate cyclase in vitro

Protein kinase C catalyzes phosphorylation of purified rat brain guanylate cyclase. The phosphorylation is marked by concomitant increase in guanylate cyclase activity. TPA further enhances both phosphorylation and activity of guanylate cyclase. Data seem to provide clues to the molecular mechanism of one of the transformation-like responses mimicked by 12-O-tetradecanoylphorbol-13-acetate, i.e. the elevation of cyclic GMP. It is envisaged that protein kinase C may have a central role in the understanding of molecular events triggering carcinogenesis.     

Interleukin 2 modulation of adenylate cyclase. Potential role of protein kinase C

Interleukin 2 (IL 2) stimulated DNA synthesis of murine T lymphocytes (CT6) in a concentration-dependent manner, over a range of 1-1000 units/ml. This proliferative effect of IL 2 was attenuated by simultaneous exposure to prostaglandin E2 (PGE)2. In intact cells, IL 2 inhibited both basal and PGE2-stimulated cAMP production; the amount of cAMP generated was dependent upon the relative concentrations of IL 2 and PGE2. The effect of IL 2 on CT6 cell proliferation and cAMP production was mimicked by the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), which, like IL 2, causes a translocation and activation of protein kinase C. While PGE2 stimulated adenylate cyclase activity in membrane preparations, neither IL 2 nor TPA inhibited either basal or stimulated membrane adenylate cyclase activity. However, when CT6 cells were pretreated with IL 2 or TPA and membranes incubated with calcium and ATP, both basal and PGE2-and NaF-stimulated membrane adenylate cyclase activity was inhibited. This inhibition of adenylate cyclase activity was also observed if membranes from untreated cells were incubated with protein kinase C purified from CT6 lymphocytes in the presence of calcium and ATP. The data suggest that the decreased cAMP production which accompanies CT6 cell proliferation results from an inhibition of adenylate cyclase activity mediated by protein kinase C and that these two distinct protein phosphorylating systems interact to modulate the physiological response to IL 2.     

Cytochemical localization of adenylate cyclase activity in rat olfactory cells

Summary Adenylate cyclase activity was demonstrated in the cilia, dendritic knob and axon of rat olfactory cells by using a strontium-based cytochemical method. The activity in the cilia and the dendritic knob was enhanced by non-hydrolyzable GTP (guanosine triphosphate) analogues and forskolin, and inhibited by Ca²⁺, all in agreement with biochemical reports of the odorant-sensitive adenylate cyclase. The results support the hypothesis of cyclic AMP working as a second messenger in olfactory transduction and imply that the transduction sites exist not only in the olfactory cilia but also in the dendritic knob. Enzymatic activity was also observed in the olfactory dendritic shaft by treating the tissue with 0.0002% Triton X-100, although the properties and role of the enzyme in this region are uncertain. The detergent inhibited the enzymatic activity in the cilia and the dendritic knob.     

Protein kinase C phosphorylates the inhibitory guanine-nucleotide-binding regulatory component and apparently suppresses its function in hormonal inhibition of adenylate cyclase

Human platelet membrane proteins were phosphorylated by exogenous, partially purified Ca2+-activated phospholipid-dependent protein kinase (protein kinase C). The phosphorylation of one of the major substrates for protein kinase C (Mr = 41 000) was specifically suppressed by the beta subunit of the inhibitory guanine-nucleotide-binding regulatory component (Gi, Ni) of adenylate cyclase. The free alpha subunit of Gi (Mr = 41 000) also served as an excellent substrate for the kinase (greater than 0.5 mol phosphate incorporated per mol of subunit), but the Gi oligomer (alpha X beta X gamma) did not. Treatment of cyc- S49 lymphoma cells, which are deficient in Gs/Ns (the stimulatory component) but contain functional Gi/Ni, with the phorbol ester, 12-O-tetradecanoylphorbol 13-acetate, a potent activator of protein kinase C, did not alter stimulation of adenylate cyclase catalytic activity by forskolin, whereas the Gi/Ni-mediated inhibition of the cyclase by the hormone, somatostatin, was impaired in these membranes. The results suggest that the alpha subunit of the inhibitory guanine-nucleotide-binding regulatory component of adenylate cyclase may be a physiological substrate for protein kinase C and that the function of the component in transducing inhibitory hormonal signals to adenylate cyclase is altered by its phosphorylation.     

Functional reconstitution of the olfactory membrane: incorporation of the olfactory adenylate cyclase in liposomes

Chemosensory cilia isolated from the olfactory epithelium of Rana catesbeiana were solubilized with Lubrol PX in the presence of supplementary lipid, forskolin, and sodium fluoride. Subsequent removal of the detergent by adsorption onto Biobeads SM2 results in the formation of proteoliposomes that display forskolin- and GTP gamma S-sensitive adenylate cyclase activity. Sucrose gradient centrifugation of liposomes formed in the presence of fluorescently labeled phosphatidylcholine demonstrates association between the olfactory adenylate cyclase and the exogenously added lipid. Forskolin stimulates the enzyme in reconstituted membranes with the same potency as in native membranes (EC50 = 1-2 microM). However, GTP gamma S is 350-fold more potent in native membranes (EC50 = 4.0 +/- 0.5 nM) than in reconstituted membranes (EC50 = 1.4 +/- 0.3 microM). These studies represent a first step toward the functional reconstitution and molecular dissection of the olfactory membrane.     

Parathyroid hormone stimulates protein kinase C but not adenylate cyclase in mouse epidermal keratinocytes.

Intact human parathyroid hormone, hPTH [1-84], and the hPTH [1-34] fragment stimulated membrane-associated protein kinase C (PKC) activity in immortalized (but still differentiation-competent) murine BALB/MK-2 skin keratinocytes. Unexpectedly, the hormone and its fragment did not stimulate adenylate cyclase. The failure of PTH to stimulate adenylate cyclase activity was not due to the lack of a functioning receptor-cyclase coupling mechanism because the cells were stimulated to synthesize cyclic adenosine monophosphate (cyclic AMP) by the beta-adrenergic drug isoproterenol. Thus, skin keratinocytes seem to have an unconventional PTH receptor that is coupled to a PKC-activating mechanism but not to adenylate cyclase. These observations suggest that normal and neoplastic skin keratinocytes respond to the PTH-related peptide that they make and secrete.     

Protein kinase C stimulates adenylate cyclase activity in prolactin-secreting rat adenoma (GH4C1) pituicytes by inactivating the inhibitory GTP-binding protein Gi

The phorbol ester 12-O-tetradecanoyl-phorbol 13-acetate (TPA) and thyroliberin exerted additive stimulatory effects on prolactin release and synthesis in rat adenoma GH4C1 pituicytes in culture. Both TPA and thyroliberin activated the adenylate cyclase in broken cell membranes. When combined, the secretagogues displayed additive effects. TPA did not alter the time course (time lag) of adenylate cyclase activation by hormones, guanosine 5'-[beta,gamma-imino]triphosphate or forskolin, nor did it affect the enzyme's apparent affinity (basal, 7.2 mM; thyroliberin-enhanced, 2.2 mM) for free Mg2+. The TPA-mediated adenylate cyclase activation was entirely dependent on exogenously added guanosine triphosphate. ED50 (dose yielding half-maximal activation) was 60 microM. Access to free Ca2+ was necessary to express TPA activation of the enzyme, however, the presence of calmodulin was not mandatory. TPA-stimulated adenylate cyclase activity was abolished by the biologically inactive phorbol ester, 4 alpha-phorbol didecanoate, by the protein kinase C inhibitor polymyxin B and by pertussis toxin, while thyroliberin-sensitive adenylate cyclase remained unaffected. Experimental conditions known to translocate protein kinase C to the plasma membrane and without inducing adenylate cyclase desensitization, increased both basal and thyroliberin-stimulated enzyme activities, while absolute TPA-enhanced adenylate cyclase was maintained. Association of extracted GTP-binding inhibitory protein, Gi, from S49 cyc- murine lymphoma cells with GH4C1 cell membranes yielded a reduction of basal and hormone-stimulated adenylate cyclase activities, while net inhibition of the cyclase of somatostatin was dramatically enhanced. However, TPA restored completely basal and hormone-elicited adenylate cyclase activities in the Gi-enriched membranes. Finally, TPA completely abolished the somatostatin-induced inhibition of adenylate cyclase in both hybrid and non-hybrid membranes. These data suggest that, in GH4C1 cells, protein kinase C stimulation by phorbol esters completely inactivates the n alpha i subunit of the inhibitory GTP-binding protein, leaving the n beta subunit functionally intact. It can also be inferred that thyroliberin conveys its main effect on the adenylate cyclase through activation of the stimulatory GTP-binding protein, Gs.     

Parathyroid hormone uses both adenylate cyclase and protein kinase C to regulate acid production in osteoclasts

Osteoclasts, isolated from the endosteum of 2.5- to 3-week-old chickens, were treated with acridine orange, a hydrogen ion concentration-sensitive fluorescent dye, in order to monitor changes in acid production. The adenylate cyclase inhibitor, alloxan, blocked parathyroid hormone (PTH)-stimulated acid production. Dibutyryl cyclic adenosine monophosphate, a membrane-permeant form of cyclic adenosine monophosphate, mimicked the PTH effect. Bisindolylmaleimide, a specific inhibitor of protein kinase C (PKC), blocked the initial stimulation (15, 30, and 60 min) of acid production by PTH but had no effect on long-term stimulation (120 min). Confocal microscopy of osteoclasts stained with fluorescein-conjugated bisindolylmaleimide revealed a shift in location of PKC from the cytoplasm to the plasma membrane region after treatment with parathyroid hormone. The results of these studies support the hypothesis that PTH regulation of acid production in osteoclasts involves both adenylate cyclase and PKC as effectors. J. Cell. Biochem. 65:565–573. © 1997 Wiley-Liss Inc.     

Regulation of GH3 pituitary tumour-cell adenylate cyclase activity by activators of protein kinase C.

The influence of protein kinase C (PKC) activation on cyclic AMP production in GH3 cells has been studied. The stimulation of cyclic AMP accumulation induced by forskolin and cholera toxin was potentiated by 4 beta-phorbol 12,13-dibutyrate (PDBu). Moreover, PDBu, which causes attenuation of the maximal response to vasoactive intestinal polypeptide (VIP), also induced a small right shift in the dose-response curve for VIP-induced cyclic AMP accumulation. PDBu-stimulated cyclic AMP accumulation was unaffected by pretreatment of cells with pertussis toxin or the inhibitory muscarinic agonist, oxotremorine. PDBu stimulation of adenylate cyclase activity required the presence of a cytosolic factor which appeared to translocate to the plasma membrane in response to the phorbol ester. The diacylglycerol-generating agents thyroliberin, bombesin and bacterial phospholipase C each stimulated cyclic AMP accumulation, but, unlike PDBu, did not attenuate the stimulation induced by VIP. These results suggest that PKC affects at least two components of the adenylate cyclase complex. Stimulation of cyclic AMP accumulation is probably due to modification of the catalytic subunit, whereas attenuation of VIP-stimulated cyclic AMP accumulation appears to be due to the phosphorylation of a different site, which may be the VIP receptor.     

Down-regulation of protein kinase C in rat glioma C6 cells: effects on the beta-adrenergic receptor-coupled adenylate cyclase

Continuous exposure of rat glioma C6 cells to 12-O-tetradecanoylphorbol-13-acetate (TPA) resulted in a time and dose dependent loss of [3H]phorbol dibutyrate binding sites and protein kinase C activity. Thus, by 24 h, the cells were essentially depleted of protein kinase C activity. In agreement with previous studies, TPA treatment caused a reduction in isoproterenol-stimulated adenylate cyclase activity and a sequestration of beta-adrenergic receptors. Cells were treated with TPA for 24-48 h to completely down-regulate protein kinase C and then exposed to isoproterenol. Agonist-mediated desensitization of adenylate cyclase and sequestration of beta-adrenergic receptors occurred at similar rates in control and TPA-treated cells. In addition, agonist-mediated down-regulation of beta-adrenergic receptors was not impaired by the absence of protein kinase C activity. Although both agonists and phorbol esters cause desensitization of the beta-adrenergic receptor-coupled adenylate cyclase, agonist-mediated events can occur independently of protein kinase C.     

Microtubule-associated adenylate cyclase

Twice-cycled bovine brain or rat brain microtubule protein contains an adenylate cyclase activity that passes 0.2 micron filters, is activated 2-7-fold by 30 microM forskolin, shows modest stimulation by fluoride (especially in the presence of added AI3+), but is virtually insensitive to added guanine nucleotides. The activity is insensitive to various hormones or Ca2+/calmodulin. The adenylate cyclase is active with both Mg2+ and Mn2+ but activity is less in the presence of Mg2+ than with Mn2+. The cyclase is inhibited by agonists of the adenosine P site. It is proposed that the catalytic unit of adenylate cyclase and probably small quantities of the guanine nucleotide regulatory protein, Ns, are cycled along with microtubules.     

Particulate adenylate cyclase plays a key role in human sperm olfactory receptor-mediated chemotaxis

Human sperm chemotaxis is a critical component of the fertilization process, but the molecular basis for this behavior remains unclear. Recent evidence shows that chemotactic responses depend on activation of the sperm olfactory receptor, hOR17-4. Certain floral scents, including bourgeonal, activate hOR17-4, trigger pronounced Ca(2+) fluxes, and evoke chemotaxis. Here, we provide evidence that hOR17-4 activation is coupled to a cAMP-mediated signaling cascade. Multidimensional protein identification technology was used to identify potential components of a G-protein-coupled cAMP transduction pathway in human sperm. These products included various membrane-associated adenylate cyclase (mAC) isoforms and the G(olf)-subunit. Using immunocytochemistry, specific mAC isoforms were localized to particular cell regions. Whereas mAC III occurred in the sperm head and midpiece, mAC VIII was distributed predominantly in the flagellum. In contrast, G(olf) was found mostly in the flagellum and midpiece. The observed spatial distribution patterns largely correspond to the spatiotemporal character of hOR17-4-induced Ca(2+) changes. Behavioral and Ca(2+) signaling responses of human sperm to bourgeonal were bioassayed in the presence, or absence, of the adenylate cyclase antagonist SQ22536. This specific agent inhibits particulate AC, but not soluble AC, activation. Upon incubation with SQ22536, cells ceased to exhibit Ca(2+) signaling, chemotaxis, and hyperactivation (faster swim speed and flagellar beat rate) in response to bourgeonal. Particulate AC is therefore required for induction of hOR17-4-mediated human sperm behavior and represents a promising target for future design of contraceptive drugs.