J. Anim Sci. 2007. 85:E4-E6. doi:10.2527/jas.2006-475
© 2007 American Society of Animal Science
TRIENNIAL REPRODUCTION SYMPOSIUM |
Regulation of meiotic maturation1
F. J. Richard2
Centre de Recherche en Biologie de la Reproduction, Département des Sciences Animales, Université Laval, Québec, Canada G1K 7P4
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Abstract
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Mammalian oocytes are arrested at prophase of the first meiotic division before induction of maturation by the preovulatory LH surge. In vitro, oocyte maturation occurs spontaneously. The first meiotic arrest is characterized by a large nucleus called the germinal vesicle. One important signaling molecule for resumption of meiosis is cyclic AMP (cAMP). High levels of cAMP block spontaneous meiotic resumption. Research investigating the regulation of oocyte cAMP has led to the discovery of new receptors, guanosine 5'-triphosphate-binding (G) proteins, cyclases, and phosphodiesterases. Leydig insulin-like 3, a polypeptide growth factor of the insulin family, is expressed in thecal cells. Leydig insulin-like 3 activates the Leu-rich, repeat-containing, G protein-coupled receptor 8, which is expressed in the oocyte. Coupled to the inhibitory GTP binding protein, this receptor leads to a decrease in cAMP production. Treatment with Leydig insulin-like 3 polypeptide initiates meiotic progression of oocytes in preovulatory follicles, demonstrating the importance of cAMP management for meiotic resumption. Furthermore, microinjection of an antibody against stimulatory G protein (Gs) into mouse oocytes results in meiotic resumption, suggesting that meiotic arrest of the oocyte is dependent on Gs activity. The orphan Gs-linked receptor, GPR3, is expressed in the oocyte. The oocytes of GPR3-null mice resume meiosis when still in their follicles, suggesting that GPR3 is involved in the control of cAMP production and thus meiotic arrest. Cyclic nucleotides are synthesized by cyclases and degraded by phosphodiesterases. Mouse and rat oocytes express isoform 3 of adenylyl cyclase. In the mouse, the null mutation results in approximately 50% of the oocytes resuming meiosis, demonstrating the importance of the synthesis of cAMP in controlling nuclear maturation. The null mutation of the major phosphodiesterase expressed in mouse oocytes results in female sterility due to ovulation of meiotically arrested oocytes that cannot be fertilized. Maintenance of meiotic arrest is explained by constitutive cAMP signaling associated with undetectable cAMP-phosphodiesterase activity. Collectively, these results are beginning to illuminate the key signaling molecules involved in the control of intraoocyte cAMP levels, thus regulating the arrest and resumption of meiosis.
Key Words: mammal • meiosis • oocyte
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INTRODUCTION
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Oogonia entering meiosis progress to the dictyate stage of the first meiotic prophase, at which meiosis is arrested and the oocytes display a characteristic nucleus commonly known as the germinal vesicle (GV). Mammalian oocytes arrested at the GV stage acquire the capacity to resume meiosis during growth of the ovarian follicle from the primary to the preovulatory stage. The ovulatory surge of gonadotropins triggers oocyte maturation and rupture of the ovarian follicle. The resumption of nuclear maturation in the oocyte after the prophase stage is characterized by GV breakdown and occurs spontaneously in vitro (Pincus and Enzmann, 1935
; Edwards, 1965). Competence for the resumption of meiosis is acquired during oocyte growth at the stage of follicular development when the antrum is formed (Sorensen and Wassarman, 1976).
Meiotic arrest relies on cyclic AMP (cAMP), high levels of which in the oocyte prevent premature resumption (Cho et al., 1974). Cyclic AMP also blocks spontaneous resumption of meiosis efficiently in numerous species, such as starfish (Taieb et al., 1997), Xenopus (Ferrell, 1999), pigs (Racowsky, 1985), cows (Aktas et al., 1995), rats (Dekel et al., 1981), and mice (Vivarelli et al., 1983). The purpose of this brief review is to describe some recent results suggesting that the oocyte has all of the machinery required for the synthesis and degradation of cyclic nucleotides. Recent efforts to understand the management of cAMP in the oocyte have revealed new signaling machinery involving receptors, guanosine 5'-triphosphate-binding (G) proteins, cyclases, and phosphodiesterases.
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THE LEYDIG INSULIN-LIKE 3 SYSTEM
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Leydig insulin-like 3 (INSL3) is a polypeptide growth factor of the insulin family. Female INSL3-null mice have impaired fertility associated with deregulation of the estrous cycle (Nef and Parada, 1999). This insulin-related growth factor is expressed in the thecal cells of rodent ovarian follicles and upregulated by LH (Kawamura et al., 2004). It binds the Leu-rich, repeat-containing, G protein-coupled receptor 8 (LGR8) that is expressed in the oocyte (Kawamura et al., 2004). Coupling to the inhibitory G protein, LGR8 leads to a decrease in the intracellular cAMP concentration (Kawamura et al., 2004). Because INSL3 initiates meiotic progression in follicle-enclosed oocytes (Kawamura et al., 2004) and decreases the intracellular level of cAMP, and its transcription in ovarian thecal cells is upregulated by LH, it is suggested that the INSL3-LGR8 system plays a paracrine role in mediating preovulatory LH stimulation. This system seems to be entirely driven by cAMP as a conventional second messenger.
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ORPHAN GUANINE NUCLEOTIDE BINDING PROTEIN-COUPLED STIMULATORY G PROTEIN-LINKED RECEPTOR
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Meiotic arrest in mammalian oocytes is maintained by an unknown signal from the surrounding somatic cells of the ovarian follicle (Tsafriri and Channing, 1975; Richard and Sirard, 1996; Downs, 1997). One important signaling molecule for cAMP synthesis is the heterotrimeric G protein. Microinjection of an antibody against the stimulatory G protein (Gs) into the mouse oocyte results in the resumption of meiosis (Mehlmann et al., 2002). Consistent with these results, the orphan G-protein-coupled, Gs-linked receptor, GPR3, is expressed in the oocyte (Mehlmann et al., 2004).
In female GPR3-null mice, the oocytes resume meiosis within the antral follicles (Mehlmann et al., 2004). When mouse oocytes are injected with small interfering double-stranded RNA targeting GPR3, meiosis is resumed (Mehlmann, 2005). In mouse oocytes, GPR3 activates Gs constitutively (Freudzon et al., 2005), consistent with its functional role. Recently, an additional G-protein-coupled Gs-linked receptor, GPR12, has been shown to be expressed specifically in rat oocytes (Hinckley et al., 2005), suggesting that GPR3 or GPR12, or both, are involved in cAMP synthesis. Thus, activation of signaling molecules in the cAMP synthesis pathway in the oocyte is involved in meiotic arrest.
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CYCLASE AND PHOSPHODIESTERASE
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Cyclic nucleotides are directly synthesized by cyclases. Adenylyl cyclase has been demonstrated immunocytochemically in bovine cumulus oocyte complexes (Kuyt et al., 1988). The adenylyl cyclase activator, forskolin, is known to increase the intracellular level of cAMP in bovine oocytes (Bilodeau et al., 1993). Mouse and rat oocytes express isoform 3 of adenylyl cyclase (Horner et al., 2003). A null mutation of isoform 3 in the mouse leads to the resumption of meiosis in approximately 50% of the oocytes (Horner et al., 2003), again indicating the importance of intraoocyte cAMP synthesis in the resumption process.
Once cyclic nucleotides have been synthesized, they are degraded by phosphodiesterases (PDE). Null mutation of the major PDE expressed in mouse oocytes (PDE3A) causes female sterility, owing to ovulation of GV stage-arrested oocytes that cannot be fertilized (Masciarelli et al., 2004). Oocytes deficient in PDE3A do not undergo meiotic maturation, suggesting that the meiotic block is maintained by constitutive cAMP signaling associated with a lack of cAMP-PDE activity (Masciarelli et al., 2004). In the rat (Richard et al., 2001), cow (Mayes and Sirard, 2002; Thomas et al., 2002), and pig (Laforest et al., 2005), the use of specific PDE3 inhibitors has demonstrated that cAMP degradation is important in maintaining meiotic arrest. Interestingly, PDE3 activity seems to be finely regulated in rodent oocytes during in vitro maturation; it is upregulated, decreasing the intracellular cAMP level a few minutes before meiosis is resumed (Richard et al., 2001).
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EPIDERMAL GROWTH FACTOR-LIKE PEPTIDES
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It is well known that epidermal growth factor (EGF) induces the resumption of meiosis in oocytes (Dekel and Sherizly, 1985; Downs et al., 1988). The EGF-like peptide, epiregulin, has been reported to be expressed in bovine granulosa cells, in follicles with oocytes competent to develop into blastocysts (Robert et al., 2001). In rodents, 3 EGF-like peptides, epiregulin, amphiregulin, and ß-cellulin, are expressed in the granulosa cells in response to LH stimulation (Park et al., 2004). All 3 stimulate the resumption of meiosis in cultures of follicle-enclosed oocytes (Park et al., 2004). In the rat, the same 3 EGF-like peptides seem to mediate the stimulation of oocyte maturation by LH (Ashkenazi et al., 2005). In a recent model of EGF-like peptide action, it has been proposed that after the EGF-like peptides are released by the granulosa cells upon LH stimulation, they bind to their receptors and promote oocyte maturation either by steroid production (Jamnongjit et al., 2005) or in association with the intracellular level of cAMP (Conti et al., 2005).
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SUMMARY AND CONLCUSIONS
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Collectively, these results are beginning to illuminate the key signaling molecules involved in the control of intraoocyte cAMP levels, thus regulating the arrest and resumption of meiosis. The new evidence for a role of EGF-like growth factors in the resumption of oocyte meiosis are promising but need to be studied further in nonrodent species.
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Footnotes
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1 Presented at the ADSA-ASAS Joint Annual Meeting, Triennial Reproduction Symposium: The Follicle and Oocyte, Minneapolis, MN, July 2006. 
2 Corresponding author: Francois.Richard{at}crbr.ulaval.ca
Received for publication July 18, 2006.
Accepted for publication September 28, 2006.
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Abstract
INTRODUCTION
