Mitogen-activated protein (MAP) kinases, also known as extracellular signal-regulated kinases (ERKs), are thought to act as an integration point for multiple biochemical signals because they are activated by a wide variety of extracellular signals, are rapidly phosphorylated on threonine and tyrosine residues, and are highly conserved in
evolution. Charest et al. (1993) cloned a cDNA encoding a member of a family of tyrosyl-phosphorylated and
mitogen-activated protein kinases (MAPKs) that participates in cell cycle progression. They referred to the protein as
p44(erk1) and the gene as ERK1.
NCBI Summary:
The protein encoded by this gene is a member of the MAP kinase family. MAP kinases, also known as extracellular signal-regulated kinases (ERKs), act in a signaling cascade that regulates various cellular processes such as proliferation, differentiation, and cell cycle progression in response to a variety of extracellular signals. This kinase is activated by upstream kinases, resulting in its translocation to the nucleus where it phosphorylates nuclear targets. Alternatively spliced transcript variants encoding different protein isoforms have been described. [provided by RefSeq, Jul 2008]
Expression and possible roles of extracellular signal-related kinases 1-2 (ERK1-2) in mouse primordial germ cell development. Sorrenti M et al. (2020) In the present work, we described the expression and activity of extracellular signal-related kinases 1-2 (ERK1-2) in mouse primordial germ cells (PGCs) from 8.5-14.5 days post coitum (dpc) and investigated whether these kinases play a role in regulating the various processes of PGC development. Using immunofluorescence and immunoblotting to detect the active phosphorylated form of ERK1-2 (p-ERK1-2), we found that the kinases were present in most proliferating 8.5-10.5 dpc PGCs, low in 11.5 dpc PGCs, and progressively increasing between 12.5-14.5 dpc both in female and male PGCs. In vitro culture experiments showed that inhibiting activation of ERK1-2 with the MEK-specific inhibitor U0126 significantly reduced the growth of 8.5 dpc PGCs in culture but had little effect on 11.5-12.5 dpc PGCs. Moreover, we found that the inhibitor did not affect the adhesion of 11.5 dpc PGCs, but it significantly reduced their motility features onto a cell monolayer. Further, while the ability of female PGCs to begin meiosis was not significantly affected by U0126, their progression through meiotic prophase I was slowed down. Notably, the activity of ERK1-2 was necessary for maintaining the correct expression of oocyte-specific genes crucial for germ cells survival and the formation of primordial follicles.//////////////////
MAPK3/1 Participates in the Activation of Primordial Follicles through mTORC1-KITL Signaling. Zhao Y et al. (2017) The majority of ovarian primordial follicles are preserved in a dormant state to maintain the female reproductive lifespan, and only a few primordial follicles are activated to enter the growing follicle pool in each wave. Recent studies have shown that primordial follicular activation depends on mammalian target of rapamycin complex 1 (mTORC1)-KIT ligand (KITL) signaling in pre-granulosa cells and its receptor (KIT)-phosphoinositol 3 kinase (PI3K) signaling in oocytes. However, the upstream regulator of mTORC1 signaling is unclear. The results of the present study showed that the phosphorylated mitogen-activated protein kinase3/1 (MAPK3/1) protein is expressed in some primordial follicles and all growing follicles. Culture of 3 days post-parturition (dpp) ovaries with the MAPK3/1 signaling inhibitor U0126 significantly reduced the number of activated follicles and was accompanied by dramatically reduced granulosa cell proliferation and increased oocyte apoptosis. Western blot and immunofluorescence analyses showed that U0126 significantly decreased the phosphorylation levels of Tsc2, S6K1, and rpS6 and the expression of KITL, indicating that U0126 inhibits mTORC1-KITL signaling. Furthermore, U0126 decreased the phosphorylation levels of Akt, resulting in a decreased number of oocytes with Foxo3 nuclear export. To further investigate MAPK3/1 signaling in primordial follicle activation, we used phosphatase and tensin homolog deleted on chromosome 10 (PTEN) inhibitor bpV(HOpic) to promote primordial follicle activation. In this model, U0126 also inhibited the activation of primordial follicles and mTORC1 signaling. Thus, these results suggest that MAPK3/1 participates in primordial follicle activation through mTORC1-KITL signaling. This article is protected by copyright. All rights reserved.//////////////////
Effect of the transient pharmacological inhibition of mapk3/1 pathway on ovulation in mice. Siddappa D et al. (2015) Mitogen-activated protein kinase 3/1 (Mapk3/1) pathway is critical for LH signal transduction during ovulation. However, the mechanisms remain incompletely understood. We hypothesized that Mapk pathway regulates ovulation through transcriptional regulation of ovulatory genes. To test this hypothesis we used immature mice superovulated with equine and human chorionic gonadotropins (eCG and hCG) and PD0325901, to inhibit hCG-induced Mapk3/1 activity. Mice received either the inhibitor PD0325901 (25 μg/g, i.p.) or vehicle at 2h before hCG stimulation. Administration of the inhibitor abolished Mapk3/1 phosphorylation in granulosa cells. While vehicle-treated mice ovulated normally, there were no ovulations in inhibitor-treated mice. First, we analyzed gene expression in granulosa cells at 0h, 1h and 4h post-hCG. There was expected hCG-driven increase in mRNA abundance of many ovulation-related genes including Ptgs2 in vehicle-treated granulosa cells, but not (P<0.05) in inhibitor-treated group. There was also reduced mRNA and protein abundance of the transcription factor, early growth response 1 (Egr1) in inhibitor-treated granulosa cells. We then used GRMO2 cell-line to test if Egr1 is recruited to promoter of Ptgs2 followed by chromatin immunoprecipitation with either Egr1 or control antibody. Enrichment of the promoter regions in immunoprecipitants of Egr1 antibody indicated that Egr1 binds to the Ptgs2 promoter. We then knocked down Egr1 expression in mouse primary granulosa cells using siRNA technology. Treatment with Egr1-siRNA inhibited Egr1 transcript accumulation, which was associated with reduced expression of Ptgs2 when compared to control-siRNA treated granulosa cells. These data demonstrate that transient inhibition of LH-stimulated MAPK3/1 activity abrogates ovulation in mice. We conclude that Mapk3/1 regulates ovulation, at least in part, through Egr1 and its target gene, Ptgs2 in granulosa cells of ovulating follicles in mice.//////////////////
ERK1/2 is involved in luteal cell autophagy regulation during corpus luteum regression via an mTOR-independent pathway. Choi J 2014 et al.
Autophagy is known to be regulated by the PI3K-AKT and/or MEK1/2-ERK1/2 pathways, leading to activation of mammalian target of rapamycin (mTOR), a major negative regulator of autophagy. However, some reports have also suggested that autophagic regulation by the PI3K-AKT and/or MEK1/2-ERK1/2 pathways may not be mediated by mTOR activity, and there is no direct evidence of the involvement of these pathways in luteal cell autophagy regulation. To elucidate the luteal cell-specific regulatory mechanisms of autophagy induction during corpus luteum (CL) regression, we evaluated whether luteal cell autophagy is regulated by the PI3K-AKT pathway and/or MEK1/2-ERK1/2 pathway and if this regulation is mediated by mTOR. We found that autophagy induction increased despite mTOR activation in luteal cells cultured with prostaglandin F2a (PGF2a), an important mediator of CL regression, suggesting that PGF2a-induced autophagy is independent of mTOR regulation. We also found that PGF2a-induced autophagy was not mediated by AKT activity, because AKT inhibition using a PI3K inhibitor (wortmannin) did not change autophagy induction or mTOR activity. In contrast, ERK1/2 activity increased in PGF2a-treated luteal cells, as did the levels of autophagy induction despite increased mTOR activity. Furthermore, PGF2a-mediated up-regulation of luteal cell autophagy was reversed by addition of ERK1/2 inhibitors, despite a decrease in mTOR activity. These in-vitro results suggest that luteal cell autophagy is induced by increased ERK1/2 activity during CL regression, and is independent of mTOR activity. This finding was further supported by in-vivo experiments in a pseudopregnant rat model, which showed that induction of luteal cell autophagy increased during luteal stage progression and that this was accompanied by increased ERK1/2 and mTOR activity. Taken together, our findings indicate that activation of ERK1/2 is a key event in the induction of luteal cell autophagy during CL regression which is not associated with mTOR regulation.
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Using porcine granulosa cells (PGC),
Cameron MR et al showed that physiological concentrations of LH and FSH
increase enzymatic activity of p44MAPK extracellular regulated kinase 1 (ERK1) but not that of p42MAPK (ERK2) in the
cytosol and of both ERK1 and ERK2 in the nucleus. Cytosolic ERK1 was activated by LH more rapidly than by FSH.
Cyclic AMP increased kinase activities of both ERK1 and ERK2 in the cytoplasm as well as in the nucleus. Activation of
ERK1 by gonadotropins and cAMP was accompanied by increased tyrosine phosphorylation of the kinase.
Immunohistochemical studies demonstrated predominantly cytoplasmic staining for MAPK in untreated PGC cultures
whereas treatment with gonadotropins led to increased nuclear immunoreactivity indicating translocation of MAPK to the
nucleus. The translocation as well as increase in nuclear ERK1 and ERK2 was delayed and coincided with a decrease in
cytosolic ERK1 activity. Epidermal growth factor (EGF) increased ERK1 and ERK2-associated kinase activity 7-8-fold in
the cytoplasm of PGC, while kinase activity of cytoplasmic ERK1 was enhanced 3-4-fold by LH, FSH, or cAMP.
Expression regulated by
FSH, LH, Growth Factors/ cytokines, GH, mir
Comment
MAPK3/1 is conducive to LH-mediated CNP decrease in bovine granulosa cells. Yang L et al. (2015) C-type natriuretic peptide (CNP) plays a role as an oocyte maturation inhibitor (OMI) in many species, including the bovine. However, the effects of luteinizing hormone (LH) on CNP expression and its potential mechanisms have not reported in the bovine. In the present study, we aimed to study the effects of LH on CNP expression and to illuminate the potential molecular mechanism in this process. Our results showed that LH induced epidermal growth factor receptor (EGFR) phosphorylation, mitogen-activated protein kinase3/1 (MAPK3/1) activation and CNP mRNA decrease in cultured bovine granulosa cells. Further study revealed that LH suppressed CNP expression via the MAPK3/1 signaling pathway, which was activated by the EGFR pathway. In conclusion, our research suggested that MAPK3/1 is involved in LH-mediated decrease of CNP and that this process is related to the EGFR and MAPK3/1 signal pathways.//////////////////
Characterization of microRNA profile in human
cumulus granulosa cells: identification of microRNAs that regulate Notch signaling and are associated with PCOS. Xu B et al. (2015) Polycystic ovary syndrome (PCOS), a complex and heterogeneous endocrine condition, is characterized by polycystic ovaries, hyperandrogenism, insulin resistance and chronic anovulation. Cumulus granulosa cells surrounding the oocyte are involved in different aspects of PCOS pathology. Several studies suggested that miRNAs play an important regulatory role at the post-transcriptional level in cumulus granulosa cells. Our objective was to describe the altered miRNA expression profiles and miRNA targeted signaling pathways in PCOS. Case-control study that involved 21 women with PCOS and 20 women without the disease (controls).The miRNA expression profiles of human cumulus granulosa cells were determined using next generation sequencing by Illumina Hiseq 2000. The differentially expressed miRNAs and novel miRNAs were validated by quantitative real-time PCR. The Notch3 and MAPK3 were demonstrated to be targeted by miR-483-5p based on quantitative real-time PCR, western blot and luciferase activity assay. Compared to controls, a total of 59 known miRNA were identified that differentially expressed in PCOS cumulus granulosa cells, including 21 miRNAs increase and 38 miRNAs decrease. Moreover, the novel miRNAs were predicted in PCOS and control cumulus granulosa cells. The potential regulating roles of miRNA in pathophysiology of PCOS were analyzed by GO and KEGG pathway annotation, and several important processes were identified to be targeted by the differentially expressed miRNAs, such as Notch signaling, regulation of hormone, and energy metabolism. Furthermore, Notch3 and MAPK3, the members of Notch signalling and ERK-MAPK pathway, were demonstrated to be regulated by miR-483-5p based on negative expression correlation validation and detection of Notch3/MAPK3 expression after miR-483-5p mimics transfection. Dual luciferase activity assay suggested that Notch3 and MAPK3 were directly targeted by miR-483-5p. Our data suggested that miRNAs and their targeted pathways (e.g. Notch signaling pathway) play important roles in the etiology and pathophysiology of PCOS, and provides novel candidates for molecular biomarkers or treatment targets in the research of female infertility associated to PCOS.//////////////////
Keel BA et al demonstrate that porcine GCs contain
immunodetectable MAP kinases. EGF, in a concentration- and time-dependent manner, increases tyrosine phosphorylation
and MBP kinase activity (i.e. activation) of ERK2, and to a lesser degree ERK1, suggesting that the activation of MAP
kinase may mediate the mitogenic action of EGF in GCs. Sirotkin AV,et al reported the involvement of MAP kinase in the mediation of GH action on ovarian
granulosa cells.
Growth hormone (GH), prostaglandins F (PGF) and prostaglandins E (PGE) are important
regulators of ovarian function. Therefore, interrelationships between GH and these substances and
their intracellular mechanisms might be of physiological significance in the ovary. The aims of this
study on cultured porcine ovarian granulosa cells were to determine the effect of GH on the
secretion of oxytocin (OT), PGF and PGE and whether MAP kinase could be involved in the
mediation of GH action. Experiments were carried out with cultured porcine granulosa cells to
investigate the effects of exogenous pGH (1-100 ng/ml) on the expression of MAP kinase (ERK-1,
-2) and of PGH (1-100 ng/ml) and the MAP kinase blocker PD 98059 (1 microg/ml) on the
secretion of PGF, PGE and OT. The cellular content of ERK-1 and -2 was analyzed by Western
immunoblotting and immunocytochemistry, whilst PGF, PGE and OT accumulation in the medium
was measured by RIA. Addition of GH to culture medium significantly altered the pattern of ovarian
ERK MAP kinase on SDS-PA gels: the 44 and 42 kDa bands were reduced and additional 50 and
48 kDa bands appeared. Moreover, there was an increase in the percentage of cells containing ERK
MAP kinase. GH stimulated the secretion of PGF (at a concentration of 1 ng GH per ml medium)
and OT (100 ng GH per ml), but not PGE. The MAP kinase blocker alone did not affect PGF, PGE
and OT secretion but did prevent the stimulatory effects of GH on PGF and induced stimulatory
action of GH (10 ng/ml) on PGE. GH-stimulated OT secretion was unaffected. These observations
confirm the role of GH in regulating porcine ovarian PGF, PGE and OT secretion and the presence
of ERK MAP kinase in porcine granulosa cells. Furthermore, our studies demonstrate that MAP
kinase-dependent intracellular mechanisms are dependent on GH, and that these mechanisms are
involved in the mediation of GH action on ovarian PGF and PGE but not OT secretion.
Oliver RH, et al. reported the induction of apoptosis in luteinized granulosa cells by the MAP kinase kinase (MEK) inhibitor PD98059.Motlik J, et al. reported that co-culture with pig membrana granulosa cells modulates the activity of cdc2 and MAP kinase in maturing cattle oocytes.
The phosphorylation state of ERK 1,2 proteins was determined by Western blotting.
Participation of Mitogen-activated Protein Kinase in Luteinizing Hormone-induced Differential Regulation of Steroidogenesis and Steroidogenic Gene Expression in Mural and Cumulus Granulosa Cells of Mouse Preovulatory Follicles. Su YQ et al. The LH surge induces the terminal differentiation and onset of luteinization in granulosa cells of preovulatory follicles, a process involving the differential expression of genes essential for steroidogenesis, and appearing to be mediated by complex signaling pathways. The objective of this study was to investigate whether these processes that commonly occur in mural granulosa cells (MGCs) also occur in cumulus cells, and whether they are mediated by the mitogen-activated protein kinase (MAPK), specifically MAPK3/1 (also commonly known as extracellular signal-regulated kinase 1&2, ERK1/2). The standard superovulation model for premature female mice was used to obtain MGCs and cumulus-oocyte complexes (COCs), and sensitive real-time reverse transcription polymerase chain reaction was used to simultaneously detect the expression levels of transcripts encoding key steroidogenic enzymes in the same sample. We observed significant downregulation of Cyp19a1 and upregulation of Star and Cyp11a1 mRNA expression in both COCs and MGCs after in vivo administration of hCG or in vitro treatment with gonadotropins or 8-Br-cAMP. This differential pattern of steroidogenic gene expression was correlated with the ultimate changes of circulating estradiol (E2) and progesterone (P4) levels after administration of hCG. In vitro, when MGCs and COCs were treated with U0126, a specific inhibitor of MAPK3/1 activation, gonadotropin- or 8-Br-cAMP-induced P4 production, as well as expression of Star and Cyp11a1 mRNA, were significantly downregulated; whereas the levels of E2 and Cyp19a1 mRNA in the same samples were significantly upregulated. We conclude that the surge of preovulatory LH induces the differential expression of transcripts encoding key steroidogenic enzymes essential for E2 and P4 synthesis in both cumulus and MGCs and this process is mediated by the MAPK3/1-dependent pathway.
Transcriptional profiling of granulosa cells from a patient with recurrent empty follicle syndrome. Inan MS et al. Empty follicle syndrome (EFS) is characterized by the absence of oocytes after apparently normal follicular development and the pathogenesis of this syndrome is not well characterized. The aim of this study was to analyse whole gene expression of granulosa cells (GC) from a patient with recurrent EFS by using Affymetrix GeneChip. A total of 160 genes were identified as being differentially expressed (by at least two-fold) between EFS GC and the control GC. Most of the differentially expressed genes were involved in cell growth and death. Among these were MAPK3, which plays an important role in the inhibition of apoptosis, was down-regulated 2.3-fold in EFS GC. Moreover, secretory phospholipase A2 and transforming growth factor receptor II, key regulators of cell death pathway, were down-regulated 3.54- and 2.82-fold respectively in EFS. Gene expression of granulosa cells from the EFS patient was significantly altered. The absence of the oocytes might be due to the increased apoptotic gene expression and the reduction of transcripts whose products are responsible for healthy follicular growth. Gene expression analyses might be a useful technique in identifying markers to follow a healthy follicular maturation and understanding the events that lead to EFS.
Follicle stages
Antral, Corpus luteum
Comment
Luteinizing hormone-induced extracellular-signal regulated kinase activation differently modulates progesterone and androstenedione production in bovine theca cells Tajima K, et al .
It has been reported that gonadotropins promoted phosphorylation of ERK/MAPK in granulosa cells. However, little is known about the effects of gonadotropin on ERK activity in theca cells. This study explores how LH/forskolin controls ERK phosphorylation in cultured bovine theca cells. Effects of ERK on steroidogenesis were also investigated. Phosphorylation of ERK in bovine theca cells was augmented by LH and forskolin in 5 min; it decreased thereafter below basal levels in 20 min. Nevertheless, phosphorylation of the ERK kinase, MEK, was unaffected. Addition of H89 (a protein kinase A inhibitor) significantly reduced the effect of LH/forskolin on ERK phosphorylation. A potent MEK inhibitor PD98059 eliminated ERK phosphorylation and augmented progesterone production concomitantly with the elevation of intracellular steroidogenic acute regulatory protein mRNA in LH/forskolin-stimulated theca cells. In contrast to progesterone production, androgen production was diminished significantly by inhibition of ERK with decreased intracellular P450c17 mRNA levels. Taking these results together, we conclude that LH/cAMP leads to phosphorylation of ERK in a biphasic manner through MEK-independent pathway in bovine theca cells. Protein kinase A-induced phosphatase could possibly contribute to the phosphorylation process. Furthermore, modulation of ERK phosphorylation involves control of thecal steroidogenesis via modulation of the expression of steroidogenic acute regulatory protein and P450c17.
Phenotypes
Mutations
2 mutations
Species: mouse
Mutation name: None
type: null mutation fertility: infertile - ovarian defect Comment: MAPK3/1 (ERK1/2) in ovarian granulosa cells are essential for female fertility. Fan HY et al. A surge of luteinizing hormone (LH) from the pituitary gland triggers ovulation, oocyte maturation, and luteinization for successful reproduction in mammals. Because the signaling molecules RAS and ERK1/2 (extracellular signal-regulated kinases 1 and 2) are activated by an LH surge in granulosa cells of preovulatory follicles, we disrupted Erk1/2 in mouse granulosa cells and provide in vivo evidence that these kinases are necessary for LH-induced oocyte resumption of meiosis, ovulation, and luteinization. In addition, biochemical analyses and selected disruption of the Cebpb gene in granulosa cells demonstrate that C/EBPbeta (CCAAT/Enhancer-binding protein-beta) is a critical downstream mediator of ERK1/2 activation. Thus, ERK1/2 and C/EBPbeta constitute an in vivo LH-regulated signaling pathway that controls ovulation- and luteinization-related events.
Species: mouse
Mutation name: type: null mutation fertility: infertile - ovarian defect Comment: ERK1/2 Activities Are Dispensable for Oocyte Growth but Are Required for Meiotic Maturation and Pronuclear Formation in Mouse. Zhang YL et al. (2015) Previous studies revealed that extracellular regulated kinase-1 and -2 (ERK1/2) cascade plays pivotal roles in regulating oocyte meiotic cell cycle progression. However, most knowledge about the in vivo function of ERK1/2 in mammalian oocytes was indirectly obtained from analyzing the phenotypes of Mos knockout mice. In this study, we knocked out Erk1 and Erk2 in mouse oocytes as early as the primordial follicle stage using the well-characterized Gdf9-Cre mouse model, and for the first time directly investigated the in vivo function of ERK1/2 in mouse oocytes. In this novel mouse model, we observed that ERK1/2 activities in oocyte are dispensable for primordial follicle maintenance, activation and follicle growth. Different from the Mos null oocytes, the ERK1/2-deleted oocytes had well-assembled spindles at metaphase I (MI), extruded polar body-1 (PB1) with normal sizes, and did not undergo a full parthenogenetic activation characterized for pronuclear formation. However, the ovulated ERK1/2-deleted oocytes had poorly-assembled metaphase II (MII) spindles, spontaneously released polar body-2 (PB2), and were arrested at another metaphase called metaphase III (MIII). In addition, ERK1/2 deletion prevented male pronuclear formation after fertilization, and caused female infertility. In conclusion, these results indicate that ERK1/2 activities are required for not only MII-arrest maintenance, but also efficient pronuclear formation in mouse oocytes.//////////////////