The role of hydrogen sulfide (H2S) is addressed in Xenopuslaevis oocytes. Three enzymes involved in H2S metabolism, cystathionine β-synthase, cystathionine γ-lyase, and 3-mercaptopyruvate sulfurtransferase, were detected in prophase I and metaphase II-arrested oocytes and drove an acceleration of oocyte meiosis resumption when inhibited. Moreover, meiosis resumption is associated with a significant decrease in endogenous H2S. On another hand, a dose-dependent inhibition was obtained using the H2S donor, NaHS (1 and 5 mM). NaHS impaired translation. NaHS did not induce the dissociation of the components of the M-phase promoting factor (MPF), cyclin B and Cdk1, nor directly impacted the MPF activity. However, the M-phase entry induced by microinjection of metaphase II MPF-containing cytoplasm was diminished, suggesting upstream components of the MPF auto-amplification loop were sensitive to H2S. Superoxide dismutase and catalase hindered the effects of NaHS, and this sensitivity was partially dependent on the production of reactive oxygen species (ROS). In contrast to other species, no apoptosis was promoted. These results suggest a contribution of H2S signaling in the timing of amphibian oocytes meiosis resumption.

• Klíčová slova
• Xenopus laevis, cell cycle, hydrogen sulfide, meiosis, oocyte,
• MeSH
• apoptóza účinky léků   MeSH
• cyklin B metabolismus   MeSH
• cystathionin-beta-synthasa antagonisté a inhibitory   metabolismus   MeSH
• cystathionin-gama-lyasa antagonisté a inhibitory   metabolismus   MeSH
• cytoplazma metabolismus   MeSH
• faktor podporující zrání metabolismus   MeSH
• fosfatasy cdc25 metabolismus   MeSH
• katalasa metabolismus   MeSH
• meióza účinky léků   MeSH
• metafáze účinky léků   MeSH
• oocyty chemie   enzymologie   metabolismus   MeSH
• profáze meiózy I účinky léků   MeSH
• proteinkinasy metabolismus   MeSH
• proteiny buněčného cyklu metabolismus   MeSH
• proteiny Xenopus metabolismus   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• sulfan metabolismus   MeSH
• sulfidy metabolismus   farmakologie   MeSH
• sulfurtransferasy antagonisté a inhibitory   metabolismus   MeSH
• superoxiddismutasa metabolismus   MeSH
• viabilita buněk účinky léků   MeSH
• Xenopus laevis MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 3-mercaptopyruvate sulphurtransferase MeSH Prohlížeč
• CDC25C protein, human MeSH Prohlížeč
• CDK1 protein, Xenopus MeSH Prohlížeč
• cyklin B MeSH
• cystathionin-beta-synthasa MeSH
• cystathionin-gama-lyasa MeSH
• faktor podporující zrání MeSH
• fosfatasy cdc25 MeSH
• katalasa MeSH
• proteinkinasy MeSH
• proteiny buněčného cyklu MeSH
• proteiny Xenopus MeSH
• reaktivní formy kyslíku MeSH
• sodium bisulfide MeSH Prohlížeč
• sulfan MeSH
• sulfidy MeSH
• sulfurtransferasy MeSH
• superoxiddismutasa MeSH

Resumption of meiosis was inhibited in mouse oocyte cumulus complexes (OCC) co-cultured with pig membrane granulosa (PMG). After 3 and 6 h of co-culture these oocytes possessed an intact nuclear envelope and their nucleolar surface was associated with granules approximately 80 nm in diameter. Preincubation of OCCs for 30, 45, 60 or 90 min followed by co-culture with PMG for 2 h of either OCCs or denuded oocytes resulted in germinal vesicle breakdown (GVBD) in approximately 0, 30, 70 and 100% mouse OCCs and in approximately 30, 60, 80 and 100% denuded oocytes, respectively. It seems that the inhibitory contact between mouse oocytes and PMG was established during the first h of co-culture. After isolation from antral follicles the oocytes contained 2.75 fmol cyclic adenosine 3', 5'-monophosphate (cAMP). When OCCs were co-cultured for 1, 2 or 3 h with PMG, the amount of cAMP per oocyte was 1.34, 1.33 and 1.51 fmol, respectively. After culture of OCCs in control medium the amount of cAMP was 1.21, 1.39 and 2.16 fmol, respectively. The present results suggest that the inhibitory activity of PMG is not species-specific. Moreover, PMG prevented resumption of meiosis in mouse oocytes in spite of the cAMP drop in oocyte cytoplasm characteristic of the resumption of meiosis.

• MeSH
• AMP cyklický farmakologie   fyziologie   MeSH
• bazální membrána MeSH
• elektronová mikroskopie MeSH
• folikulární buňky fyziologie   MeSH
• kultivované buňky MeSH
• meióza účinky léků   fyziologie   MeSH
• myši MeSH
• oocyty účinky léků   fyziologie   ultrastruktura   MeSH
• prasata MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• AMP cyklický MeSH

BACKGROUND INFORMATION: In fully grown mouse oocytes, a decrease in cAMP concentration precedes and is linked to CDK1 (cyclin-dependent kinase 1) activation. The molecular mechanism for this coupling, however, is not defined. PKB (protein kinase B, also called AKT) is implicated in CDK1 activation in lower species. During resumption of meiosis in starfish oocytes, MYT1, a negative regulator of CDK1, is phosphorylated by PKB in an inhibitory manner. It can imply that PKB is also involved in CDK1 activation in mammalian oocytes. RESULTS: We monitored activation of PKB and CDK1 during maturation of mouse oocytes. PKB phosphorylation and activation preceded GVBD (germinal vesicle breakdown) in oocytes maturing either in vitro or in vivo. Activation was transient and PKB activity was markedly reduced when virtually all of the oocytes had undergone GVBD. PKB activation was independent of CDK1 activity, because although butyrolactone I prevented CDK1 activation and GVBD, PKB was nevertheless transiently phosphorylated and activated. LY-294002, an inhibitor of phosphoinositide 3-kinase-PKB signalling, suppressed activation of PKB and CDK1 as well as resumption of meiosis. OA (okadaic acid)-sensitive phosphatases are involved in PKB-activity regulation, because OA induced PKB hyperphosphorylation. During resumption of meiosis, PKB phosphorylated on Ser(473) is associated with nuclear membrane and centrosome, whereas PKB phosphorylated on Thr(308) is localized on centrosome only. CONCLUSIONS: The results of the present paper indicate that PKB is involved in CDK1 activation and resumption of meiosis in mouse oocytes. The presence of phosphorylated PKB on centrosome at the time of GVBD suggests its important role for an initial CDK1 activation.

• MeSH
• aktivace enzymů MeSH
• centrozom metabolismus   MeSH
• chromony farmakologie   MeSH
• fosforylace MeSH
• gama-butyrolakton analogy a deriváty   farmakologie   MeSH
• jaderný obal metabolismus   MeSH
• kyselina okadaová farmakologie   MeSH
• meióza * MeSH
• morfoliny farmakologie   MeSH
• myši MeSH
• oocyty účinky léků   fyziologie   MeSH
• proteinkinasa CDC2 metabolismus   MeSH
• protoonkogenní proteiny c-akt antagonisté a inhibitory   fyziologie   MeSH
• serin metabolismus   MeSH
• techniky in vitro MeSH
• threonin metabolismus   MeSH
• transport proteinů MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one MeSH Prohlížeč
• butyrolactone I MeSH Prohlížeč
• chromony MeSH
• gama-butyrolakton MeSH
• kyselina okadaová MeSH
• morfoliny MeSH
• proteinkinasa CDC2 MeSH
• protoonkogenní proteiny c-akt MeSH
• serin MeSH
• threonin MeSH

Membrana granulosa was isolated from healthy large antral follicles of prepubertal or cyclic gilts stimulated with PMSG or PMSG and hCG. Ultrastructural observations revealed that pieces of pig membrana granulosa were associated with the basement membrane. The cattle cumulus-enclosed oocytes (COC) were placed in the rolled pieces of the pig membrana granulosa (PMG). After 8 and 24 hr of coculture with PMG from prepubertal gilts, only 16% and 21% of oocytes underwent GVBD, respectively. PMG from PMSG-stimulated cyclic gilts blocked the resumption of meiosis in all COC. The inhibitory effect of heterologous granulosa cells was fully reversible. When COC were initially incubated for 2 and 4 hr, subsequent culture in PMG prevented GVBD in 100% and 36% of oocytes, respectively. This suggests that functional contact between COC and PMG was established during the first 2 hr of coculture. To follow metabolic cooperation between PMG and COC, PMG was prelabeled with 3H-uridine and cocultured with COC. Autoradiography on semithin sections revealed the intensive passage of 3H-uridine from PMG into the cumulus layer and an oocyte. COC placed in PMG after GVBD (8 and 12 hr of an initial incubation) did not extrude the first polar body. PMG isolated from cyclic gilts after PMSG and hCG stimulation also inhibited GVBD of COC. Since nearly all COC placed in PMG isolated 10 and 12 hr after hCG remained in the GV stage after 24 hr of coculture, the hCG stimulation did not substantially diminish the meiosis inhibiting activity of PMG.(ABSTRACT TRUNCATED AT 250 WORDS)

• MeSH
• choriogonadotropin farmakologie   MeSH
• elektronová mikroskopie MeSH
• folikulární buňky cytologie   účinky léků   MeSH
• gonadotropiny koňské farmakologie   MeSH
• meióza * MeSH
• mezibuněčná komunikace MeSH
• oocyty cytologie   MeSH
• prasata MeSH
• skot MeSH
• techniky in vitro MeSH
• zvířata MeSH
• Check Tag
• skot MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• choriogonadotropin MeSH
• gonadotropiny koňské MeSH

In vivo, resumption of oocyte meiosis occurs in large ovarian follicles after the preovulatory surge of luteinizing hormone (LH). The LH surge leads to the activation of a broad signaling network in mural granulosa cells equipped with LH receptors. The signals generated in the mural granulosa cells are further augmented by locally produced peptides or steroids and transferred to the cumulus cell compartment and the oocyte itself. Over the last decade, essential progress has been made in the identification of molecular events associated with the final maturation and ovulation of mammalian oocytes. All new evidence argues for a multiple roles of mitogen-activated protein kinase 3/1 (MAPK3/1) in the gonadotropin-induced ovulation processes. However, the knowledge of gonadotropin-induced signaling pathways leading to MAPK3/1 activation in follicular cells seems limited. To date, only the LH-induced transactivation of the epidermal growth factor receptor/MAPK3/1 pathway has been described in granulosa/cumulus cells even though other mechanisms of MAPK3/1 activation have been detected in other types of cells. In this review, we aimed to summarize recent advances in the elucidation of gonadotropin-induced mechanisms leading to the activation of MAPK3/1 in preovulatory follicles and cultured cumulus-oocyte complexes and to point out a specific role of this kinase in the processes accompanying final maturation of the mammalian oocyte.

• MeSH
• folikulární buňky metabolismus   MeSH
• MAP kinasový signální systém genetika   MeSH
• meióza fyziologie   MeSH
• mitogenem aktivovaná proteinkinasa 1 metabolismus   MeSH
• mitogenem aktivovaná proteinkinasa 3 metabolismus   MeSH
• mutace MeSH
• myši MeSH
• oocyty fyziologie   MeSH
• steroidy biosyntéza   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• mitogenem aktivovaná proteinkinasa 1 MeSH
• mitogenem aktivovaná proteinkinasa 3 MeSH
• steroidy MeSH

In vitro maturation of rabbit cumulus-enclosed oocytes was fully inhibited in alpha-amanitin- (100 micrograms/ml) and cycloheximide- (5 micrograms/ml) supplemented media. The inhibition was reversible and substantially reduced by delaying the addition of alpha-amanitin (2h) or cycloheximide (3 h). In contrast, both drugs did not inhibit germinal vesicle breakdown in denuded oocytes. Co-culture of granulosa cells (1 x 10(6)/ml) with denuded oocytes did not substitute for an intact cumulus. The data presented here suggest that the resumption of meiosis in rabbit cumulus-enclosed oocytes is dependent upon early transcriptional and translational events which probably occur within the cumulus cells.

• MeSH
• amanitiny farmakologie   MeSH
• cykloheximid farmakologie   MeSH
• genetická transkripce MeSH
• králíci * MeSH
• meióza * účinky léků   MeSH
• oocyty cytologie   účinky léků   fyziologie   MeSH
• ovariální folikul cytologie   fyziologie   MeSH
• proteosyntéza * MeSH
• RNA biosyntéza   MeSH
• zvířata MeSH
• Check Tag
• králíci * MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• amanitiny MeSH
• cykloheximid MeSH
• RNA MeSH

Aurora kinase A (AURKA) is an important mitotic kinase involved in the G2/M transition, centrosome maturation and separation, and spindle formation in somatic cells. We used transgenic models that specifically overexpress in mouse oocytes either wild-type (WT-AURKA) or a catalytically inactive (kinase-dead) (KD-AURKA) AURKA to gain new insights regarding the role of AURKA during oocyte maturation. AURKA activation occurs shortly after hCG administration that initiates maturation in vivo. Although AURKA activity is increased in WT-AURKA oocytes, resumption of meiosis is not observed in the absence of hCG administration. Control oocytes contain one to three microtubule organizing centers (MTOCs; centrosome equivalent) at prophase I. At the time of germinal vesicle breakdown (GVBD), the first visible marker of resumption of meiosis, the MTOC number increases. In WT-AURKA oocytes, the increase in MTOC number occurs prematurely but transiently without GVBD, whereas the increase in MTOC number does not occur in control and KD-AURKA oocytes. AURKA activation is biphasic with the initial activation not requiring CDC25B-CDK1 activity, whereas full activation, which is essential for the increase in MTOCs number, depends on CDK1 activity. AURKA activity also influences spindle length and regulates, independent of its protein kinase activity, the amount of MTOC associated with gamma-tubulin. Both WT-AURKA and KD-AURKA transgenic mice have normal fertility during first 6 mo of life. These results suggest that although AURKA is not a trigger kinase for G2/M transition in mouse oocytes, it regulates MTOC number and spindle length, and, independent of its protein kinase activity, gamma-tubulin recruitment to MTOCs.

• MeSH
• aktivace enzymů účinky léků   MeSH
• Aurora kinasa A MeSH
• choriogonadotropin farmakologie   MeSH
• HeLa buňky MeSH
• kinasy Aurora MeSH
• kultivované buňky MeSH
• lidé MeSH
• meióza účinky léků   genetika   fyziologie   MeSH
• myši inbrední C57BL MeSH
• myši knockoutované MeSH
• myši MeSH
• oocyty enzymologie   metabolismus   fyziologie   MeSH
• oogeneze účinky léků   genetika   fyziologie   MeSH
• organizační centrum mikrotubulů účinky léků   metabolismus   MeSH
• protein-serin-threoninkinasy genetika   metabolismus   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• AURKA protein, human MeSH Prohlížeč
• Aurka protein, mouse MeSH Prohlížeč
• Aurora kinasa A MeSH
• choriogonadotropin MeSH
• kinasy Aurora MeSH
• protein-serin-threoninkinasy MeSH

Mammalian oocytes are arrested at prophase I until puberty when luteinizing hormone (LH) induces resumption of meiosis of follicle-enclosed oocytes. Resumption of meiosis is tightly coupled with regulating cyclin-dependent kinase 1 (CDK1) activity. Prophase I arrest depends on inhibitory phosphorylation of CDK1 and anaphase-promoting complex-(APC-CDH1)-mediated regulation of cyclin B levels. Prophase I arrest is maintained by endogenously produced cyclic adenosine monophosphate (cAMP), which activates protein kinase A (PKA) that in turn phosphorylates (and activates) the nuclear kinase WEE2. In addition, PKA-mediated phosphorylation of the phosphatase CDC25B results in its cytoplasmic retention. The combined effect maintains low levels of CDK1 activity that are not sufficient to initiate resumption of meiosis. LH triggers synthesis of epidermal growth factor-like factors in mural granulosa cells and leads to reduced cGMP transfer from cumulus cells to oocytes via gap junctions that couple the two cell types. cGMP inhibits oocyte phosphodiesterase 3A (PDE3A) and a decline in oocyte cGMP results in increased PDE3A activity. The ensuing decrease in oocyte cAMP triggers maturation by alleviating the aforementioned phosphorylations of WEE2 and CDC25B. As a direct consequence CDC25B translocates into the nucleus. The resulting activation of CDK1 also promotes extrusion of WEE2 from the nucleus thereby providing a positive amplification mechanism for CDK1 activation. Other kinases, e.g. protein kinase B, Aurora kinase A and polo-like kinase 1, also participate in resumption of meiosis. Mechanisms governing meiotic prophase I arrest and resumption of meiosis share common features with DNA damage-induced mitotic G2-checkpoint arrest and checkpoint recovery, respectively. These common features include CDC14B-dependent activation of APC-CDH1 in prophase I arrested oocytes or G2-arrested somatic cells, and CDC25B-dependent cell cycle resumption in both oocytes and somatic cells.

• MeSH
• AMP cyklický metabolismus   MeSH
• anafázi podporující komplex MeSH
• Cdh1 proteiny MeSH
• cyklin B1 metabolismus   MeSH
• epidermální růstový faktor metabolismus   MeSH
• G2 fáze * MeSH
• inhibiční proteiny cyklin-dependentních kinas metabolismus   MeSH
• komplexy ubikvitinligas metabolismus   MeSH
• meióza * MeSH
• metafáze MeSH
• myši MeSH
• oocyty metabolismus   MeSH
• oogeneze * MeSH
• profáze meiózy I MeSH
• proliferace buněk * MeSH
• proteiny buněčného cyklu metabolismus   MeSH
• signální transdukce * MeSH
• věkové faktory MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• AMP cyklický MeSH
• anafázi podporující komplex MeSH
• Ccnb1 protein, mouse MeSH Prohlížeč
• Cdh1 proteiny MeSH
• cyklin B1 MeSH
• epidermální růstový faktor MeSH
• Fzr1 protein, mouse MeSH Prohlížeč
• inhibiční proteiny cyklin-dependentních kinas MeSH
• komplexy ubikvitinligas MeSH
• proteiny buněčného cyklu MeSH

In mammalian females, oocytes are stored in the ovary and meiosis is arrested at the diplotene stage of prophase I. When females reach puberty oocytes are selectively recruited in cycles to grow, overcome the meiotic arrest, complete the first meiotic division and become mature (ready for fertilization). At a molecular level, the master regulator of prophase I arrest and meiotic resumption is the maturation-promoting factor (MPF) complex, formed by the active form of cyclin dependent kinase 1 (CDK1) and Cyclin B1. However, we still do not have complete information regarding the factors implicated in MPF activation. In this study we document that out of three mammalian serum-glucocorticoid kinase proteins (SGK1, SGK2, SGK3), mouse oocytes express only SGK1 with a phosphorylated (active) form dominantly localized in the nucleoplasm. Further, suppression of SGK1 activity in oocytes results in decreased CDK1 activation via the phosphatase cell division cycle 25B (CDC25B), consequently delaying or inhibiting nuclear envelope breakdown. Expression of exogenous constitutively active CDK1 can rescue the phenotype induced by SGK1 inhibition. These findings bring new insights into the molecular pathways acting upstream of MPF and a better understanding of meiotic resumption control by presenting a new key player SGK1 in mammalian oocytes.

• Klíčová slova
• CDK1, MPF, Meiosis, Nuclear envelope breakdown, Oocyte, SGK1,
• MeSH
• faktor podporující zrání * metabolismus   MeSH
• kontrolní body buněčného cyklu MeSH
• meióza MeSH
• myši MeSH
• oocyty metabolismus   MeSH
• profáze meiózy I MeSH
• protein-serin-threoninkinasy genetika   MeSH
• proteiny bezprostředně časné * genetika   metabolismus   MeSH
• savci metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• faktor podporující zrání * MeSH
• protein-serin-threoninkinasy MeSH
• proteiny bezprostředně časné * MeSH
• serum-glucocorticoid regulated kinase MeSH Prohlížeč

In denuded and cumulus-enclosed pig oocytes, puromycin at concentrations 5, 10, and 25 micrograms/ml did not lower the rate of germinal vesicle breakdown (GVBD) after 24 h of culture. GVBD was prevented in 50, 75, and 100 micrograms/ml of puromycin. After 40 h of culture, 5 and 10 micrograms puromycin/ml impaired significantly incidence of metaphase II (42 and 30%), respectively. Concentrations of 25 and 50 micrograms puromycin/ml absolutely prevented the first polar body (I PB) expulsion. The results indicated that GVBD in pig oocytes is far less sensitive to puromycin than I PB expulsion. Culture of cumulus-enclosed pig oocytes isolated with a piece of membrana granulosa (C + P oocytes) did not allow GVBD after 24 and 32 h in control medium. After 24 h of culture, GVBD occurred in 43 and 56% of C + P oocytes in the medium supplemented with 17 and 25 micrograms puromycin/ml. GV was broken down in 80 and 68% of C + P oocytes cultured in 17 and 25 micrograms puromycin/ml for 32 h. It is concluded that inhibition of protein synthesis by puromycin released pig oocytes from the block exerted by granulosa cells.

• MeSH
• folikulární buňky cytologie   účinky léků   MeSH
• inhibitory syntézy proteinů farmakologie   MeSH
• kultivované buňky účinky léků   MeSH
• meióza účinky léků   MeSH
• oocyty cytologie   účinky léků   MeSH
• ovariální folikul cytologie   účinky léků   MeSH
• prasata MeSH
• puromycin farmakologie   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• inhibitory syntézy proteinů MeSH
• puromycin MeSH