Activation of PKA, p38 MAPK and ERK1/2 by gonadotropins in cumulus cells is critical for induction of EGF-like factor and TACE/ADAM17 gene expression during in vitro maturation of porcine COCs
© Yamashita et al; licensee BioMed Central Ltd. 2009
Received: 27 July 2009
Accepted: 24 December 2009
Published: 24 December 2009
During ovulation, it has been shown that LH stimulus induces the expression of numerous genes via PKA, p38 MAPK, PI3K and ERK1/2 in cumulus cells and granulosa cells. Our recent study showed that EGF-like factor and its protease (TACE/ADAM17) are required for the activation of EGF receptor (EGFR), cumulus expansion and oocyte maturation of porcine cumulus-oocyte complexes (COCs). In the present study, we investigated which signaling pathways are involved in the gene expression of EGF-like factor and in Tace/Adam17 expression in cumulus cells of porcine COC during in vitro maturation.
Areg, Ereg, Tace/Adam17, Has2, Tnfaip6 and Ptgs2 mRNA expressions were detected in cumulus cells of porcine COCs by RT-PCR. Protein level of ERK1/2 phosphorylation in cultured cumulus cells was analyzed by westernblotting. COCs were visualized using a phase-contrast microscope.
When COCs were cultured with FSH and LH up to 2.5 h, Areg, Ereg and Tace/Adam17 mRNA were expressed in cumulus cells of COCs. Areg, Ereg and Tace/Adam17 gene expressions were not suppressed by PI3K inhibitor (LY294002), whereas PKA inhibitor (H89), p38 MAPK inhibitor (SB203580) and MEK inhibitor (U0126) significantly suppressed these gene expressions. Phosphorylation of ERK1/2, and the gene expression of Has2, Tnfaip6 and Ptgs2 were also suppressed by H89, SB203580 and U0126, however, these negative effects were overcome by the addition of EGF to the medium, but not in the U0126 treatment group.
The results showed that PKA, p38 MAPK and ERK1/2 positively controlled the expression of EGF-like factor and TACE/ADMA17, the latter of which impacts the cumulus expansion and oocyte maturation of porcine COCs via the EGFR-ERK1/2 pathway in cumulus cells.
In mammals, luteinizing hormone (LH) stimulation induces morphological and physiological changes in granulosa cells and cumulus cells, causing them to progress to the ovulation process . During this period, cumulus cells expressed cumulus expansion-related genes, Hyaluronan synthase 2 (Has2) [2, 3], Tumor necrosis factor α-induced protein 6 (Tnfaip6) [4, 5], and Pentraxin 3 (Ptx3) [6, 7], which is necessary for the synthesis and stability of hyaluronan-rich extracellular matrix. In Tnfaip6 null mice  or Ptx3 null mice , number of ovulated oocytes decreased and in vivo fertilization was completely interrupted, suggesting that cumulus expansion was essential for both ovulation and fertilization processes.
It is known that since LH receptor (Lhcgr) is mainly expressed in granulosa cells, the EGF-like factor produced in granulosa cells by LH surge acts on cumulus cells to induce cumulus expansion. Some factors were introduced to transmit the LH signal from granulosa cells to cumulus cells. For example, prostagrandin E2 (PGE2) that produced from granulosa cells and cumulus cells by prostagradin synthase 2 (PTGS2) was required for induction of Has2 and Tnfaip6 gene, cumulus expansion and oocyte meiotic resumption . The EGF-like factors, Amphiregulin (AREG), Epiregulin (EREG) and β-cellulin (BTC) were also recently reported as potent factor. The EGF-like factor was induced by LH stimuli in granulusa cells, and EGF receptor (EGFR) was localized on cumulus cells [9–11]. When mouse COCs were cultured with AREG, Has2, Tnfaip6 and Ptgs2 were expressed in cumulus cells. TACE/ADAM17, the cleavage enzyme of EGF-like factor to soluble forms, was also expressed in porcine granulosa cells in vivo in response to hCG administration . Thus, in vivo during the ovulation process, LH induces EGF-like factor expression in granulosa cells and the release of the EGF domain by TACE/ADAM17 acts on cumulus cells, which induce cumulus expansion.
In in vitro maturation of oocytes, COCs were recovered from antral follicles and cultured with FSH and/or LH. We previously showed that FSH and LH up-regulate EGF-like factor and Tace/Adam17 mRNA expression, and gonadotropins-induced cumulus expansion and oocyte maturation of porcine COCs were suppressed by EGF receptor tyrosine kinase inhibitor or TACE/ADAM17 inhibitor . The results suggested that FSH- and LH-induced cumulus expansion was dependent on the expression and functions of EGF-like factors. Although the regulation of EGF-like factor expression in cancer cell lines has been reported [14, 15], the mechanisms of EGF-like factor and TACE/ADAM17 expression in cumulus cells cultured with FSH and/or LH have remained unclear during in vitro maturation of porcine COCs.
The binding of FSH and/or LH in granulosa cells to its own receptors led to rapidly and nongenomic activation of PKA, p38 MAPK, and PI3K in a cAMP-dependent manner  and of ERK1/2 via the SRC/RAS-dependent pathway . In mice, since each inhibitor of PKA, p38 MAPK, PI3K or ERK1/2 suppressed the expression of cumulus expansion-related gene [10, 18, 19], cumulus expansion [18, 19] or meiotic maturation of oocyte , we estimated that these signaling pathways induced by gonadotropin overlap the EGF-like factor-EGFR pathway, which induces full cumulus expansion and oocyte maturation.
In this study, to clarify the intracellular pathway involved in EGF-like factor and Tace/Adam17 expression in cumulus cells, we examined the effect of PKA inhibitor (H89), p38 MAPK inhibitor (SB203580), PI3K inhibitor (LY294002) and MEK inhibitor (U0126) on Areg, Ereg and Tace/Adam17 expression in cumulus cells during in vitro maturation of porcine COCs. Additionally, we investigated the effect of these drugs on ERK1/2 phosphorylation, cumulus expansion and oocyte meiotic resumption in pig.
High purified porcine FSH and porcine LH were gifts from the National Hormone and Pituitary Program (Rockville, MD, USA). Fetal calf serum (FCS) was obtained from Invitrogen (Carlsbad, CA, USA). Oligonucleotide poly- (dT) was purchased from Amersham Pharmacia Biotech (Newark, NJ, USA). Avian myeloblastosis virus reverse transcriptase and Taq DNA Polymerase were from Promega (Madison, WI). Routine chemicals and reagents were obtained from Nakarai Chemical Co. (Osaka, Japan) or Sigma (Sigma Chemical Co., St. Louis, MO, USA).
In vitro culture of porcine COCs
Isolation of porcine COCs was described previously . Briefly, porcine ovaries were recovered from 5- to 7-month-old prepubertal gilts at a local slaughterhouse. COCs were collected from the surfaces of intact healthy antral follicles measuring from 3 to 5 mm in diameter. The 20 COCs were cultured up to 40 h with both 20 ng/ml highly purified porcine FSH (NIDDK, Torrance, CA) and 500 ng/ml porcine LH (NIDDK). The maturation medium was modified NCSU37  supplemented with 10% (v/v) FCS (Gibco BRL, Grand Island, NY) and 7 mM Taurine (Sigma St Louis, MO). At selected time intervals, COCs were collected for RNA and protein isolation.
The assessment of cumulus expansion was observed using phase-contrast microscopy (Olympus IMT2, Olympus, Tokyo Japan) and a 10× objective.
Treatment with inhibitors
In the case of treatment with each specific inhibitor, namely PKA, H89 (10 uM) (Sigma), p38 MAPK, SB203580 (20 uM) (Sigma), PI3K, LY294002 (20 uM) (Sigma) or MEK, U0126 10 uM (Sigma), COCs were cultured for 0, 2.5, 5, 10, 20 or 40 h with each of these inhibitors. H89 was dissolved in maturation medium at 10 mM and stored at -20°C. SB203580, LY294002 and U0126 were dissolved in dimethylsulfoxide (DMSO) at 20 mM and 10 mM, respectively, and stored at -20°C. The final concentration of each compound (as described above) was obtained by dilution (1:1000) with the maturation medium. The final concentration of vehicles (DMSO) was 0.1% (vol/vol), which did not affect the function of cumulus cells during meiotic resumption of porcine oocytes .
After cumulus cells were cultured, they were washed three times in PBS. Total RNA was extracted from cumulus cells using the SV Total RNA Isolation System (Promega) according to the instruction manual, and dissolved in nuclease-free water. The final RNA concentrations were determined by absorbance using a spectrophotometer.
List of primers employed for RT-PCR
Anneling temprature (X)
5'-CTA CAA TGA GCT GCG TGT GG-3'
5'-TAG CTC TTC TCC AGG GAG GA-3'
5'-CAC CAG AAC AAA AAG GTT CTG TC-3'
5'-AAG TCC ATG AAG ACT CAC ACC AT-3'
5'AAG ACA ATC CAC GTG TGG CTC AAG-3'
5'-CGA TTT TTG TAC CAT CTG CAG AAA-3'
5'-GAC ATG AAT GGC AAA TGT GAG AAA C-3'
5'-AGT CTG TGC TGG GGT CTT CCT GGA-3'
5'-GAA TTA CCC AGT CCT GGC TT-3'
5'-GGA TAA ACT GGT AGC CAA CA-3'
5'-TCA TAA CTC CAT ATG GCT TGA AC-3'
5'-TCT TCG TAC TCA TTT GGG AAG CC-3'
5'-CTG CCG TGT CGC TCT GCA CTG-3'
5'-TCA TAA CTC CAT ATG GCT TGA AC-3'
Western blot analysis
Cumulus cells were lysed in Laemmli sample buffer and protein extracts were stored at -80°C until use. After denaturing by boiling for 5 min, 10 ul of each samples containing equal amounts of protein (10 ug) was separated by SDS-PAGE on 10% polyacrylamide gel, then transferred onto PVDF membrane (GE Healthcare). The membrane was blocked with 5% (w/v) nonfat dry milk (GE Healthcare) in PBS. Primary antibodies were added in 2.5% (w/v) nonfat dry milk in 0.1% (v/v) Tween 20 (Sigma)/PBS (PBS-T), and incubated overnight at 4°C. Anti-phospho-ERK1/2 and β-ACTIN were purchased from Cell Signaling Technology, Inc (Beverly, MA) and diluted at 1:2,000 or 1:10,000, respectively. After four washes in PBS-T, the membranes were incubated for 1 h with a 1:2,000 dilution of goat anti-rabbit IgG HRP-linked antibody (Cell Signaling Technology, Inc) in 2.5% (w/v) nonfat dry milk in PBS-T at room temperature. After five washes of 10 min each with PBS-T, peroxidase activity was visualized using the ECL Western blotting detection system (GE Healthcare) according to the manufacturer's instructions.
Statistical analyses of all data from three or four replicates for comparison were carried out by one-way ANOVA followed by Duncan's multiple-range test (Statview; Abacus Concepts, Inc., Berkeley, CA). All percentage data were subjected to arcsine transformation before analysis.
Effect of each specific inhibitor of PKA, p38 MAPK, PI3K and MEK on the gonadotropin-induced Areg, Ereg and Tace/Adam17 mRNA expression during in vitro maturation of porcine COCs
Effect of each specific inhibitor of PKA, p38 MAPK and MEK on ERK1/2 phosphorylation in cumulus cells during in vitro maturation of porcine COCs
Effect of each specific inhibitor of PKA, p38 MAPK and MEK on meiotic resumption of oocytes and cumulus expansion during in vitro maturation of porcine COCs
Recently, it has been showed that the novel paracrine/autocrine factors expressed in granulosa cells and cumulus cells by LH stimuli acted on cumulus cells to induce cumulus expansion, meiotic maturation of oocytes and ovulation in mouse [9–11]. Since the suppression of the EGF receptor tyrosine kinase activity by specific inhibitor completely suppressed cumulus expansion and meiotic maturation of oocytes during in vitro maturation of mouse COCs , the EGF-like factor was quite important for ovulatory process. During in vitro maturation of porcine COCs, the addition of EGF to maturation medium significantly elevated developmental competence of oocytes to blastocyst stage , indicating that investigation of the transcriptional mechanism of the EGF-like factor and TACE/ADAM17 was essential for not only ovulatory process, but also cytoplasamic maturation of oocyte.
It has been reported that during the ovulation process the PKA-dependent pathway involving the ligand activation of G protein-coupled receptors by their ligands in cumulus and/or granulosa cells induced the expression of ovulation-related genes, including Cyp11a1 , Star  and Ptgs2 . Other reports documented that, in rat granulosa cells, FSH and LH up-regulated the phosphorylation of p38 MAPK by a PKA-independent mechanism that might be involved in Epac . Furthermore, a recent report showed that FSH nongenomically activates ERK1/2 via SRC/RAS dependent pathway in rat granulosa cells , indicating that the activation of ERK1/2 was not directly activated by cAMP-activated PKA or p38 MAPK pathway. In this study, when porcine COCs were cultured for 2.5 h with PKA inhibitor, p38 MAPK inhibitor or MEK inhibitor, FSH- and LH-induced Areg, Ereg, and Tace/Adam17 mRNA expressions were significantly suppressed. In osteoblastic cells, the promoter region of the Areg gene had a putative CRE site, and the region was quite important for parathyroid hormone-induced Areg gene induction via CREB phosphorylation . In rat granulosa cells, it has been reported that the phosphorylation of CREB was induced by FSH within 1.5 h . Our recent study showed in rat granulosa cells that phosphorylation of CREB was induced by FSH dependent manner and the phosphorylation of CREB was essential for transcription of Areg mRNA via its promoter region of CRE site (Shitanaka et al., unpublished data). We also showed in granulose-specific Erk1/2 knockout mice that Areg expression level was significantly lower than that in wild-type mice . It is known that CREB has the sites phosphorylated by ERK1/2 and p38 MAPK [31, 32]. Thus, at early in the process, p38 MAPK and ERK1/2 might be involved in the phosphorylation of CREB, which would induce Areg gene expression in cumulus cells of porcine COCs.
Our previous study showed that EGFR tyrosine kinase inhibitor or TACE/ADAM17 inhibitor suppressed the phosphorylation of ERK1/2 and the meiotic resumption of oocytes . In this study, the addition of EGF to PKA or p38 MAPK inhibitor-containing medium overcame the negative effects or cumulus expansion and oocyte meiotic resumption. However, when COCs were cultured with U0126, the treatment with EGF did not overcome the U0126 effects. Thus, the MEK-ERK1/2 pathway played dual roles in cumulus cells. One role is the induction of EGF-like factor and TACE/ADAM17 expression. The other is direct induction of cumulus expansion of porcine COCs. In granulosa cell specific Erk1/2 knockout mice, the cumulus expansion was completely suppressed via the low induction of Ptgs2 expression. In this study, the Ptgs2 expression level was suppressed by U0126, suggesting that EGF-like factor was required for the induction of cumulus expansion and oocyte meiotic resumption in porcine COCs as well as in mice.
Herein, we showed that the expression of EGF-like factor and TACE/ADAM17 in cumulus cells was induced by a PKA-, p38 MAPK- and ERK1/2-dependent mechanism during in vitro maturation of porcine COCs. The intracellular mechanism of induction of the expression of EGF-like factor and TACE/ADAM17 further induce EGF-like factor and Tace/Adam17 mRNA expressions in cumulus cells via ERK1/2 activation. Thereby, the ERK1/2 maintained its activity via the EGF domain-EGFR-ERK1/2 pathway, which resulted in full cumulus expansion, and oocyte maturation during in vitro maturation of porcine COCs.
follicle stimulating hormone
- EGF-like factor:
epidermal growth factor-like factor
tumor necrosis factor α converting enzyme/a disinteglin and metalloptotease 17
hyaluronan synhase 2
tumor necrosis factor α-induced protein 6
prostagrandin synthase 2
human chorionic gonadotropin
protein kinase A
- p38 MAPK:
p38 mitogen-activated protein kinase
extracellular-signal regulated protein
rous sarcoma oncogene
rat sarcoma viral oncogene.
Supported, in part, by grant-in-Aid for Scientific Research (Y.Y., No. 19880020) and (M.S., No. 18688016) from the Japan Society for the Promotion of Science (JSPS). Porcine FSH and LH were kindly provided by Dr. A.F. Parlow, the National Hormone and Pituitary Program, the National Institute of Diabetes and Digestive and Kidney Disease, USA. We thank Mr. I Kawashima, Mr. T. Mizukami, Mr. T. Koike, and Ms. M. Okamoto for technical assistance. We also thank the staff of the Meat Inspection Office in Hiroshima City and Tottori prefecture for supplying the porcine ovaries.
- Richards JS: Ovulation: new factors that prepare the oocyte for fertilization. Mol Cell Endocrinol 2005, 234: 75–79. 10.1016/j.mce.2005.01.004PubMedView ArticleGoogle Scholar
- Camaioni A, Hascall VC, Yanagishita M, Salustri A: Effects of exogenous hyaluronic acid and serum on matrix organization and stability in the mouse cumulus cell-oocyte complex. J Biol Chem 1993, 268: 20473–20481.PubMedGoogle Scholar
- Chen L, Russell PT, Larsen WJ: Functional significance of cumulus expansion in the mouse: roles for the preovulatory synthesis of hyaluronic acid within the cumulus mass. Mol Reprod Dev 1993, 34: 87–93. 10.1002/mrd.1080340114PubMedView ArticleGoogle Scholar
- Ochsner SA, Day AJ, Rugg MS, Breyer RM, Gomer RH, Richards JS: Disrupted function of tumor necrosis factor-alpha-stimulated gene 6 blocks cumulus cell-oocyte complex expansion. Endocrinology 2003, 144: 4376–4384. 10.1210/en.2003-0487PubMedView ArticleGoogle Scholar
- Fulop C, Szanto S, Mukhopadhyay D, Bardos T, Kamath RV, Rugg MS, Day AJ, Salustri A, Hascall VC, Glant TT, Mikecz K: Impaired cumulus mucification and female sterility in tumor necrosis factor-induced protein-6 deficient mice. Development 2003, 130: 2253–2261. 10.1242/dev.00422PubMedView ArticleGoogle Scholar
- Varani S, Elvin JA, Yan C, DeMayo J, DeMayo FJ, Horton HF, Byrne MC, Matzuk MM: Knockout of pentraxin 3, a downstream target of growth differentiation factor-9, causes female subfertility. Mol Endocrinol 2003, 16: 1154–1167. 10.1210/me.16.6.1154View ArticleGoogle Scholar
- Salustri A, Garlanda C, Hirsch E, De Acetis M, Maccagno A, Bottazzi B, Doni A, Bastone A, Mantovani G, Beck Peccoz P, Salvatori G, Mahoney DJ, Day AJ, Siracusa G, Romani L, Mantovani A: PTX3 plays a key role in the organization of the cumulus oophorus extracellular matrix and in in vivo fertilization. Development 2004, 131: 1577–1586. 10.1242/dev.01056PubMedView ArticleGoogle Scholar
- Ochsner SA, Russell DL, Day AJ, Breyer RM, Richards JS: Decreased expression of tumor necrosis factor-alpha-stimulated gene 6 in cumulus cells of the cyclooxygenase-2 and EP2 null mice. Endocrinology 2003, 144: 1008–1019. 10.1210/en.2002-220435PubMedView ArticleGoogle Scholar
- Park JY, Su YQ, Ariga M, Law E, Jin SL, Conti M: EGF-like factor as mediators of LH action in the ovulatory follicle. Science 2004, 303: 682–684. 10.1126/science.1092463PubMedView ArticleGoogle Scholar
- Shimada M, Hernandez-Gonzalez I, Gonzalez-Robayna I, Richards JS: Paracrine and autocrine regulation of epidermal growth factor-like factors in cumulus oocyte complexes and granulosa cells: key roles for prostaglandin synthase 2 and progesterone receptor. Mol Endocrinol 2006, 20: 1352–1365. 10.1210/me.2005-0504PubMedView ArticleGoogle Scholar
- Hsieh M, Lee D, Panigone S, Horner K, Chen R, Theologis A, Lee DC, Threadgill DW, Conti M: Luteinizing hormone-dependent activation of the epidermal growth factor network is essential for ovulation. Mol Cells Biol 2007, 27: 1914–1924. 10.1128/MCB.01919-06View ArticleGoogle Scholar
- Kawashima I, Okazaki T, Noma N, Nishibori M, Yamashita Y, Shimada M: Sequential exposure of porcine cumulus cells to FSH and/or LH is critical for appropriate expression of steroidogenic and ovulation-related genes that impact oocyte maturation in vivo and in vitro . Reproduction 2008, 136: 9–12. 10.1530/REP-08-0074PubMedView ArticleGoogle Scholar
- Yamashita Y, Kawashima I, Yanai Y, Nishibori M, Richards JS, Shimada M: Hormone-induced expression of tumor necrosis factor alpha-converting enzyme/A disintegrin ad metalloptotease-17 impacts porcine cumulus cell oocyte complex expression and meiotic maturation via activation of the epidermal growth factor receptor. Endocrinology 2007, 148: 6164–6175. 10.1210/en.2007-0195PubMedView ArticleGoogle Scholar
- Zhang Q, Thomas SM, Lui VW, Xi S, Siegfried JM, Fan H, Smithgall TE, Mills GB, Grandis JR: Phosphorylation of TNF-alpha converting enzyme by gastrin-releasing peptide induces amphiregulin release and EGF receptor activation. Proc Natl Acad Sci USA 2006, 103: 6901–6906. 10.1073/pnas.0509719103PubMed CentralPubMedView ArticleGoogle Scholar
- Thomas SM, Bhola NE, Zhang Q, Contrucci SC, Wentzel AL, Freilino ML, Gooding WE, Siegfriend JM, Chan DC, Grandis JR: Cross-talk between G protein-cuopled receptor and epidermal growth factor receptor signailig pathways contributes to growth and invation of head and neck squamous cell carcinoma. Cancer Res 2006, 66: 11831–11839. 10.1158/0008-5472.CAN-06-2876PubMedView ArticleGoogle Scholar
- Richards JS: New signaling pathways for hormones and cyclic adenisine 3', 5'-monophosphate action in endocrine cells. Mol Endocrinol 2001, 15: 2009–2018. 10.1210/me.15.2.209Google Scholar
- Wayne CM, Fan HY, Cheng X, Richards JS: Follicle-stimulating hormone induces multiple signaling cascades: evidence that activation of Rous sarcoma oncogene, RAS, and the epidermal growth factor receptor are critical for granulosa cell differentiation. Mol Endocrinol 2007, 21: 1940–1957. 10.1210/me.2007-0020PubMedView ArticleGoogle Scholar
- Downs SM, Hunzicker-Dunn M: Differential regulation of oocyte maturation and cumulus expansion in the mouse oocyte-cumulus cell complex by site-selective analogs of cyclic adenosine monophosphate. Dev Biol 1995, 172: 72–85. 10.1006/dbio.1995.0006PubMedView ArticleGoogle Scholar
- Su YQ, Denegre JM, Wigglesworth K, Pendola FL, O'Brien MJ, Eppig JJ: Oocyte-dependent activation of mitogen-activated protein kinase (ERK1/2) in cumulus cells is required for the maturation of the mouse oocyte-cumulus cell complex. Dev Biol 2003, 263: 126–138. 10.1016/S0012-1606(03)00437-8PubMedView ArticleGoogle Scholar
- Hoshino Y, Yokoo M, Yoshida N, Sasada H, Matsumoto H, Sato E: Phosphatidylinositol 3-kinase and Akt participate in the FSH-induced meiotic maturation of mouse oocytes. Mol Reprod Dev 69: 77–86. 10.1002/mrd.20150
- Yamashita Y, Shimada M, Okazaki T, Maeda T, Terada T: Production of progesterone from de novo-synthesized cholesterol in cumulus cells and its physiological role during meiotic resumption of porcine oocytes. Biol Reprod 2003, 68: 1193–1198. 10.1095/biolreprod.102.010934PubMedView ArticleGoogle Scholar
- Yamashita Y, Nishibori M, Terada T, Isobe N, Shimada M: Gonadotropin-induced delta14-reductase and delta7-reductase gene expression in cumulus cells during meiotic resumption of porcine oocytes. Endocrinology 2005, 146: 186–194. 10.1210/en.2004-0617PubMedView ArticleGoogle Scholar
- Shimada M, Anas MKI, Terada T: Effects of phosphotidylinositol 3-kinase inhibitors, wortmannin and LY29 on germinal vesicle breakdown (GVBD) in porcine oocytes. J Reprod Dev 4002, 44: 281–288. 10.1262/jrd.44.281View ArticleGoogle Scholar
- Sher N, Yivgi-Ohana N, Orly J: Transcriptional regulation of the cholesterol side chain cleavage cytochrome P450 gene (CYP11A1) revisited: binding of GATA, cyclic adenosine 3', 5'-monophosphate response element-binding protein and activation protein (AP)-1 proteins to a distal novel cluster of cis-regulatory element potentiates AP-2 and steroidogenic factor-1-dependent gene expression in the rodent placenta. Mol Endocrinol 2007, 21: 948–962. 10.1210/me.2006-0226PubMedView ArticleGoogle Scholar
- Yivgi-Ohana N, Sher N, Melamed-Book N, Eimerl S, Koler M, Manna PR, Stocco DM, Orly J: Transcriptional of steroidogenic acute regulatory protein in the rodent ovary and placenta: alternative modes of cyclic adenosine 3', 5'-monohposphate dependent and independent regulation. Endocrinology 2009, 150: 977–989. 10.1210/en.2008-0541PubMed CentralPubMedView ArticleGoogle Scholar
- Wu YL, Wiltbank MC: Transcriotional regulation of the cyclooxygenase-2 gene changes form protein kinase (PK) A to PKC-dependent after luteinazation of granulosa cells. Biol Reprod 2002, 66: 1505–1514. 10.1095/biolreprod66.5.1505PubMedView ArticleGoogle Scholar
- Gonzalez-Robayna IJ, Fakender AE, Ochsner S, Firestone GL, Richards JS: Follicle-Stimulating hormone (FSH) stimulates phosphorylation and activation of protein kunase B (PKB/Akt) and serum and glucocorticoid-induced kinase (Sgk): evidence for A kinase-independent signaling by FSH in granulosa cells. Mol Endocrinol 2000, 14: 1283–1300. 10.1210/me.14.8.1283PubMedView ArticleGoogle Scholar
- Qin L, Partridge NC: Stimulation of amphiregulin expression in osteoblastic cells by parathyroid hormone requires the protein kinase A and cAMP response element-binding protein signaling pathway. J Cell Biochem 2005, 96: 632–640. 10.1002/jcb.20550PubMedView ArticleGoogle Scholar
- Carlone DL, Richards JS: Functional interactions, phosphorylation, and levels of 3', 5'-cyclic adenosine monophosphate-regulatory element binding protein and steroidogenic factor-1 mediate hormone-regulated and constitutive expression of aromatase in gonadal cells. Mol Endocrinol 1997, 11: 292–304. 10.1210/me.11.3.292PubMedGoogle Scholar
- Fan HY, Liu Z, Shimada M, Sterneck E, Johnson PF, Hedrick SM, Richards JS: MAPK3/1 (ERK1/2) in ovarian granulosa cells are essential for female fertility. Science 2009, 324: 938–941. 10.1126/science.1171396PubMed CentralPubMedView ArticleGoogle Scholar
- Lochner A, Marais E, Genade S, Huisamen B, du Toit EF, Moolman JA: Protection of the ischaemic heart: investigations into the phenomenon of ischaemic preconditioning. Cardiovasc J Afr 2009, 20: 43–51.PubMed CentralPubMedGoogle Scholar
- Xing J: Coupling of RAS-MAPK pathway to gene activation by RSK2, a growth factor-regulated CREB kinase. Science 1996, 273: 959–963. 10.1126/science.273.5277.959PubMedView ArticleGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.