Amphibian oocyte maturation and protein synthesis: Related inhibition by cyclic AMP, theophylline, and papaverine

Abstract

Two inhibitors of cyclic AMP phosphodiesterase (3':5'-cyclic-AMP 5'-nucleotidohydrolase, EC 3.1.4.17), theophylline and papaverine, inhibit the maturation of Xenopus laevis oocytes induced by four different stimuli: human chorionic gonadotropin, progesterone, testosterone, and lanthanum ions. Addition of 1 mM cyclic AMP to the medium delays maturation by approximately 2 hr. Papaverine, theophylline, and cyclic AMP inhibit amino acid incorporation into oocyte proteins by 50% or more but do not inhibit amino acid uptake. The capacity of theophylline to block maturation and protein synthesis is reversed in a parallel fashion by addition of 1-5 mM calcium ion to the medium. Addition of papaverine, theophylline, and cycloheximide to oocytes at different times after hormonal treatment shows that the step sensitive to blockage by the three drugs is coincident and precedes germinal vesicle breakdown by about 1.5 hr. The maturation of full-grown oocytes is a complex process that involves meiotic cell division and prepares the cell for fertilization. Much attention has been devoted to the study of the maturation of amphibian oocytes because these cells can be obtained in large amounts from a single ovary, they are big enough to be microinjected, and they can be induced to mature in vitro.One of the puzzling aspects of the maturation process is that its onset can be triggered by such different stimuli as the hormones human chorionic gonadotropin (hCG), progesterone, and testosterone and by nonphysiological reagents such as ionophore A 23187, organomercurial compounds, and 10 mM lanthanum ions (1-5).Although we are still far from understanding the molecular mechanism that is responsive to these various stimuli, the work of several research groups has provided evidence that de novo protein synthesis, protein phosphorylation, and calcium ions play important roles in the early steps of the maturation process (6-10). Recent evidence has also pointed to the participation of cyclic AMP (cAMP) In this report we show that theophylline and papaverine, two well-known cAMP phosphodiesterase (3':5'-cyclic-AMP 5'-nucleotidohydrolase, EC 3.1.4.17) inhibitors, are potent inhibitors of oocyte maturation induced by four different stimuli: hCG, progesterone, testosterone, and lanthanum ions. These drugs, which inhibit endogenous oocyte cAMP phosphodiesterase (13), do not cause any measurable increase in the overall cAMP concentration of these cells. However, theophylline or papaverine treatment of oocytes eliminates the increase of Ca2+ uptake induced by progesterone and hCG and results in a very significant inhibition of protein synthesis in these cells. cAMP itself, added to the oocyte incubation medium, also causes an inhibition of protein synthesis and delays the onset of hormone-induced maturation. Several characteristics of the effect of theophylline, papaverine, and cAMP in blocking oocyte maturation seem to indicate that this effect is due to their inhibition of cellular protein synthesis.MATERIALS AND METHODS X. laevis wer...

G 2 arrest in Xenopus oocytes depends on phosphorylation of cdc25 by protein kinase A

“…Although cAMP levels show a partial, transient drop soon after progesterone addition and another drop before GVBD (15,18,54,55), total cellular PKA activity shows only minor fluctuations in activity between progesterone addition and GVBD (data not shown). Taken together, these results suggest that the phosphorylation status of cdc25 on Ser-287 could be controlled by selective, compartmentalized binding of cdc25, PKA, and the Ser-287 phosphatase to one of the substrate-specific PKA scaffolding proteins known as AKAPs (56,57).…”

Progesterone inhibits protein kinase A (PKA) inXenopusoocytes: demonstration of endogenous PKA activities using an expressed substrate

“…Overall, there appears to be a consensus that progesterone causes a rapid (within minutes) but modest (~20%) reduction of cAMP (Smith, 1989). However, some have also reported that this reduction is transient, with levels of cAMP recovered quickly (within 10-20 minutes) , whereas others have reported more persistently reduced levels of cAMP throughout the maturation process (Bravo et al, 1978). The basal cAMP concentration in G2 oocytes is estimated as 1 pmol/oocyte ) (V. Montplaisir and X. J. Liu, unpublished data) or 1 µM (assuming 1 µl as the average volume of a sphere of 1.2-1.3 mm diameter).…”

Vesicular traffic at the cell membrane regulates oocyte meiotic arrest

“…In Xenopus, progesterone-induced oocyte maturation is associated with a rapid transient decline (20-60%) in cAMP (Maller et al, 1979;Cicirelli and Smith, 1985), due to inhibition of adenylate cyclase (AC) (Sadler and Maller, 1981;Sadler and Maller, 1985;Finidori-Lepicard et al, 1981). Furthermore, interventions that increase cAMP, either through activation of AC (Schorderet-Slatkine and Baulieu, 1982), increasing protein kinase A (PKA) activity (Maller and Krebs, 1977) or inhibiting cAMP phosphodiesterase (Bravo et al, 1978;Sadler and Maller, 1987), block progesteroneinduced oocyte maturation. Supporting these results, inhibition of PKA induces oocyte maturation in the absence of progesterone (Maller and Krebs, 1977;Huchon et al, 1981;Sun and Machaca, 2004;Daar et al, 1993).…”