Defective synaptic plasticity in a model of Coffin-Lowry syndrome is rescued by simultaneously targeting PKA and MAPK pathways.

Empirical and computational methods were combined to examine whether individual or dual-drug treatments can restore the deficit in long-term synaptic facilitation (LTF) of the Aplysia sensorimotor synapse observed in a cellular model of Coffin-Lowry syndrome (CLS). The model was produced by pharmacological inhibition of p90 ribosomal S6 kinase (RSK) activity. In this model, coapplication of an activator of the mitogen-activated protein kinase (MAPK) isoform ERK and an activator of protein kinase A (PKA) resulted in enhanced phosphorylation of RSK and enhanced LTF to a greater extent than either drug alone and also greater than their additive effects, which is termed synergism. The extent of synergism appeared to depend on another MAPK isoform, p38 MAPK. Inhibition of p38 MAPK facilitated serotonin (5-HT)-induced RSK phosphorylation, indicating that p38 MAPK inhibits activation of RSK. Inhibition of p38 MAPK combined with activation of PKA synergistically activated both ERK and RSK. Our results suggest that cellular models of disorders that affect synaptic plasticity and learning, such as CLS, may constitute a useful strategy to identify candidate drug combinations, and that combining computational models with empirical tests of model predictions can help explain synergism of drug combinations.

The MAP-Kinase HOG1 Controls Cold Adaptation in Rhodosporidium kratochvilovae by Promoting Biosynthesis of Polyunsaturated Fatty Acids and Glycerol.

The aim of this study was to investigate the role of RKHog1 in the cold adaptation of Rhodosporidium kratochvilovae strain YM25235 and elucidate the correlation of biosynthesis of polyunsaturated fatty acids (PUFAs) and glycerol with its cold adaptation. The YM25235 strain was subjected to salt, osmotic, and cold stress tolerance analyses. mRNA levels of RKhog1, Δ12/15-fatty acid desaturase gene (RKD12), RKMsn4, HisK2301, and RKGPD1 in YM25235 were detected by reverse transcription quantitative real-time PCR. The contents of PUFAs, such as linoleic acid (LA) and linolenic acid (ALA) was measured using a gas chromatography-mass spectrometer, followed by determination of the growth rate of YM25235 and its glycerol content at low temperature. The RKHog1 overexpression, knockout, and remediation strains were constructed. Stress resistance analysis showed that overexpression of RKHog1 gene increased the biosynthesis of glycerol and enhanced the tolerance of YM25235 to cold, salt, and osmotic stresses, respectively. Inversely, the knockout of RKHog1 gene decreased the biosynthesis of glycerol and inhibited the tolerance of YM25235 to different stresses. Fatty acid analysis showed that the overexpression of RKHog1 gene in YM25235 significantly increased the content of LA and ALA, but RKHog1 gene knockout YM25235 strain had decreased content of LA and ALA. In addition, the mRNA expression level of RKD12, RKMsn4, RKHisK2301, and RKGPD1 showed an increase at 15 °C after RKHog1 gene overexpression but were unchanged at 30 °C. RKHog1 could regulate the growth adaptability and PUFA content of YM25235 at low temperature and this could be helpful for the cold adaptation of YM25235.

Chlorosis seedling lethality 1 encoding a MAP3K protein is essential for chloroplast development in rice.

BACKGROUND: Mitogen-activated protein kinase (MAPK) cascades are conserved signaling modules in eukaryotic organisms and play essential roles in immunity and stress responses. However, the role of MAPKs in chloroplast development remains to be evidently established. RESULTS: In this study, a rice chlorosis seedling lethality 1 (csl1) mutant with a Zhonghua11 (ZH11, japonica) background was isolated. Seedlings of the mutant were characterized by chlorotic leaves and death after the trefoil stage, and chloroplasts were observed to contain accumulated starch granules. Molecular cloning revealed that OsCSL1 encoded a MAPK kinase kinase22 (MKKK22) targeted to the endoplasmic reticulum (ER), and functional complementation of OsCSL1 was found to restore the normal phenotype in csl1 plants. The CRISPR/Cas9 technology was used for targeted disruption of OsCSL1, and the OsCSL1-Cas9 lines obtained therein exhibited yellow seedlings which phenocopied the csl1 mutant. CSL1/MKKK22 was observed to establish direct interaction with MKK4, and altered expression of MKK1 and MKK4 was detected in the csl1 mutant. Additionally, disruption of OsCSL1 led to reduced expression of chloroplast-associated genes, including chlorophyll biosynthetic genes, plastid-encoded RNA polymerases, nuclear-encoded RNA polymerase, and nuclear-encoded chloroplast genes. CONCLUSIONS: The findings of this study revealed that OsCSL1 played roles in regulating the expression of multiple chloroplast synthesis-related genes, thereby affecting their functions, and leading to wide-ranging defects, including chlorotic seedlings and severely disrupted chloroplasts containing accumulated starch granules.

Increased Death of Peripheral Blood Mononuclear Cells after TLR4 Inhibition in Sepsis Is Not via TNF/TNF Receptor-Mediated Apoptotic Pathway.

BACKGROUND: Apoptosis is one of the causes of immune depression in sepsis. Pyroptosis also occurs in sepsis. The toll-like receptor (TLR) 4 and receptor for advanced glycation end products (RAGE) have been shown to play important roles in apoptosis and pyroptosis. However, it is still unknown whether TLR4 inhibition decreases apoptosis in sepsis. METHODS: Stimulated peripheral blood mononuclear cells (PBMCs) with or without lipopolysaccharides (LPS) and high-mobility group box 1 (HMGB1) were cultured with or without TLR4 inhibition using monoclonal antibodies from 20 patients with sepsis. Caspase-3, caspase-8, and caspase-9 activities were measured. The expression of B cell lymphoma 2 (Bcl2) and Bcl2-associated X (Bax) was measured. The cell death of PBMCs was detected using a flow cytofluorimeter. RESULTS: After TLR4 inhibition, Bcl2 to Bax ratio elevated both in LPS and HMGB1-stimulated PBMCs. The activities of caspase-3, caspase-8, and caspase-9 did not change in LPS or HMGB1-stimulated PBMCs. The cell death of LPS and HMGB1-stimulated CD8 lymphocytes and monocytes increased after TLR4 inhibition. The cell death of CD4 lymphocytes was unchanged. CONCLUSION: The apoptosis did not decrease, while TLR4 was inhibited. After TLR4 inhibition, there was an unknown mechanism to keep cell death in stimulated PBMCs in patients with sepsis.

Let-7a targets Rsf-1 to modulate radiotherapy response of non-small cell lung cancer cells through Ras-MAPK pathway.

PURPOSE: Radiotherapy is the most commonly selective medical treatment for non-small cell lung cancer (NSCLC) and the multiple underlying mechanisms are considered as the effectively theoretical foundation. Herein, we investigated the effects of let-7a targets Rsf-1 on modulating the radiotherapy response in NSCLC cells by Ras-MAPK pathway. METHODS: A549 cells were divided into different groups to investigate the role of let-7a and Rsf-1 on the radiotherapy response. The expression of let-7a and Rsf-1 were detected by RT-PCR. Bioinformatic analysis indicated that Rsf-1 is the target of let-7a. The binding site of let-7a in the Rsf-1 3'UTR was detected based on double luciferase reporter assay and Western blot. The cell variability and proliferation were assessed by MTT and colony formation assay. The expression levels of Ras-MARK signaling pathway related proteins were assessed by RT-PCR. RESULTS: RT-PCR results showed that radiotherapy could up-regulate the expression of let-7a, thereby reducing the expression of Rsf-1, and the correlation between the two factors was negatively correlated. At the same time, let-7a overexpression and Rsf-1 silencing could further reduce the activity of A549 cells after radiotherapy, have an inhibitory effect on cell proliferation and inhibit the expression of related proteins in the Ras-MAPK pathway. CONCLUSIONS: Rsf-1 is the target of Let-7a. The present study provides evidence that let-7a targeting Rsf-1 can modulate radiotherapy response in NSCLC cells through Ras-MAPK pathway.

The Role of Intracellular Signaling Molecules in the Production of Granulocytic CSF Mononuclear Phagocytes in Stress and Cytostatic Influence.

Under conditions of steady-state hemopoiesis, nuclear factor NF-κB, in contrast to MAP kinase p38, plays an important role in the maintenance of the initial level of secretory activity of monocytes. The increase in the production of G-CSF under stress conditions (10-h immobilization) is mainly regulated by the alternative p38MARK signaling pathway via activation of p38 synthesis. It was shown that under conditions of cytostatic-induced myelosuppression, the production of protein kinase p38 in cells decreases, and it, like NF-κB, is not the main one in the production of hemopoietin by mononuclear phagocytes.

Contributions of Sodium-Hydrogen Exchanger 1 and Mitogen-Activated Protein Kinases to Enhanced Retinal Venular Constriction to Endothelin-1 in Diabetes.

Diabetes elevates endothelin-1 (ET-1) in the vitreous and enhances constriction of retinal venules to this peptide. However, mechanisms contributing to ET-1-induced constriction of retinal venules are incompletely understood. We examined roles of sodium-hydrogen exchanger 1 (NHE1), protein kinase C (PKC), mitogen-activated protein kinases (MAPKs), and extracellular calcium (Ca2+) in retinal venular constriction to ET-1 and the impact of diabetes on these signaling molecules. Retinal venules were isolated from control pigs and pigs with streptozocin-induced diabetes for in vitro studies. ET-1-induced vasoconstriction was abolished in the absence of extracellular Ca2+ and sensitive to c-Jun N-terminal kinase (JNK) inhibitor SP600125 but unaffected by extracellular signal-regulated kinase (ERK) inhibitor PD98059, p38 kinase inhibitor SB203580, or broad-spectrum PKC inhibitor Gö 6983. Diabetes (after 2 weeks) enhanced venular constriction to ET-1, which was insensitive to PD98059 and Gö 6983 but was prevented by NHE1 inhibitor cariporide, SB203580, and SP600125. In conclusion, extracellular Ca2+ entry and activation of JNK, independent of ERK and PKC, mediate constriction of retinal venules to ET-1. Diabetes activates p38 MAPK and NHE1, which cause enhanced venular constriction to ET-1. Treatments targeting these vascular molecules may lessen retinal complications in early diabetes.

S100A4 Is Involved in Stimulatory Effects Elicited by the FGF2/FGFR1 Signaling Pathway in Triple-Negative Breast Cancer (TNBC) Cells.

Triple-negative breast cancer (TNBC) is an aggressive breast tumor subtype characterized by poor clinical outcome. In recent years, numerous advancements have been made to better understand the biological landscape of TNBC, though appropriate targets still remain to be determined. In the present study, we have determined that the expression levels of FGF2 and S100A4 are higher in TNBC with respect to non-TNBC patients when analyzing "The Invasive Breast Cancer Cohort of The Cancer Genome Atlas" (TCGA) dataset. In addition, we have found that the gene expression of FGF2 is positively correlated with S100A4 in TNBC samples. Performing quantitative PCR, Western blot, CRISPR/Cas9 genome editing, promoter studies, immunofluorescence analysis, subcellular fractionation studies, and ChIP assays, we have also demonstrated that FGF2 induces in TNBC cells the upregulation and secretion of S100A4 via FGFR1, along with the ERK1/2-AKT-c-Rel transduction signaling. Using conditioned medium from TNBC cells stimulated with FGF2, we have also ascertained that the paracrine activation of the S100A4/RAGE pathway triggers angiogenic effects in vascular endothelial cells (HUVECs) and promotes the migration of cancer-associated fibroblasts (CAFs). Collectively, our data provide novel insights into the action of the FGF2/FGFR1 axis through S100A4 toward stimulatory effects elicited in TNBC cells.

The Two Sides of the Same Coin-Influenza Virus and Intracellular Signal Transduction.

Cells respond to extracellular agents by activation of intracellular signaling pathways. Viruses can be regarded as such agents, leading to a firework of signaling inside the cell, primarily induced by pathogen-associated molecular patterns (PAMPs) that provoke safeguard mechanisms to defend from the invader. In the constant arms race between pathogen and cellular defense, viruses not only have evolved mechanisms to suppress or misuse supposedly antiviral signaling processes for their own benefit but also actively induce signaling to promote replication. This creates viral dependencies that may be exploited for novel strategies of antiviral intervention. Here, we will summarize the current knowledge of activation and function of influenza virus-induced signaling pathways with a focus on nuclear factor (NF)-κB signaling, mitogen-activated protein kinase cascades, and the phosphatidylinositol-3-kinase pathway. We will discuss the opportunities and drawbacks of targeting these signaling pathways for antiviral intervention.

Nongenomic Actions of Thyroid Hormone: The Integrin Component.

The extracellular domain of plasma membrane integrin αvß3 contains a cell surface receptor for thyroid hormone analogues. The receptor is largely expressed and activated in tumor cells and rapidly dividing endothelial cells. The principal ligand for this receptor is l-thyroxine (T4), usually regarded only as a prohormone for 3,5,3'-triiodo-l-thyronine (T3), the hormone analogue that expresses thyroid hormone in the cell nucleus via nuclear receptors that are unrelated structurally to integrin αvß3. At the integrin receptor for thyroid hormone, T4 regulates cancer and endothelial cell division, tumor cell defense pathways (such as anti-apoptosis), and angiogenesis and supports metastasis, radioresistance, and chemoresistance. The molecular mechanisms involve signal transduction via mitogen-activated protein kinase and phosphatidylinositol 3-kinase, differential expression of multiple genes related to the listed cell processes, and regulation of activities of other cell surface proteins, such as vascular growth factor receptors. Tetraiodothyroacetic acid (tetrac) is derived from T4 and competes with binding of T4 to the integrin. In the absence of T4, tetrac and chemically modified tetrac also have anticancer effects that culminate in altered gene transcription. Tumor xenografts are arrested by unmodified and chemically modified tetrac. The receptor requires further characterization in terms of contributions to nonmalignant cells, such as platelets and phagocytes. The integrin αvß3 receptor for thyroid hormone offers a large panel of cellular actions that are relevant to cancer biology and that may be regulated by tetrac derivatives.

The growth of Arabidopsis primary root is repressed by several and diverse amino acids through auxin-dependent and independent mechanisms and MPK6 kinase activity.

Amino acids serve as structural monomers for protein synthesis and are considered important biostimulants for plants. In this report, the effects of all 20-L amino acids in Arabidopsis primary root growth were evaluated. 15 amino acids inhibited growth, being l-leucine (l-Leu), l-lysine (l-Lys), l-tryptophan (l-Trp), and l-glutamate (l-Glu) the most active, which repressed both cell division and elongation in primary roots. Comparisons of DR5:GFP expression and growth of WT Arabidopsis seedlings and several auxin response mutants including slr, axr1 and axr2 single mutants, arf7/arf19 double mutant and tir1/afb2/afb3 triple mutant, treated with inhibitory concentrations of l-Glu, l-Leu, l-Lys and l-Trp revealed gene-dependent, specific changes in auxin response. In addition, l- isomers of Glu, Leu and Lys, but not l-Trp diminished the GFP fluorescence of pPIN1::PIN1:GFP, pPIN2::PIN2:GFP, pPIN3::PIN3:GFP and pPIN7::PIN7:GFP constructs in root tips. MPK6 activity in roots was enhanced by amino acid treatment, being greater in response to l-Trp while mpk6 mutants supported cell division and elongation at high doses of l-Glu, l-Leu, l-Lys and l-Trp. We conclude that independently of their auxin modulating properties, amino acids signals converge in MPK6 to alter the Arabidopsis primary root growth.

HOG-Independent Osmoprotection by Erythritol in Yeast Yarrowia lipolytica.

Erythritol is a polyol produced by Yarrowia lipolytica under hyperosmotic stress. In this study, the osmo-sensitive strain Y. lipolytica yl-hog1Δ was subjected to stress, triggered by a high concentration of carbon sources. The strain thrived on 0.75 M erythritol medium, while the same concentrations of glucose and glycerol proved to be lethal. The addition of 0.1 M erythritol to the medium containing 0.75 M glucose or glycerol allowed the growth of yl-hog1Δ. Supplementation with other potential osmolytes such as mannitol or L-proline did not have a similar effect. To examine whether the osmoprotective effect might be related to erythritol accumulation, we deleted two genes involved in erythritol utilization, the transcription factor Euf1 and the enzyme erythritol dehydrogenase Eyd1. The strain eyd1Δ yl hog1Δ, which lacked the erythritol utilization enzyme, reacted to the erythritol supplementation significantly better than yl-hog1Δ. On the other hand, the strain euf1Δ yl-hog1Δ became insensitive to supplementation, and the addition of erythritol could no longer improve the growth of this strain in hyperosmotic conditions. This indicates that Euf1 regulates additional, still unknown genes involved in erythritol metabolism.

A Kinase-Phosphatase-Transcription Factor Module Regulates Adventitious Root Emergence in Arabidopsis Root-Hypocotyl Junctions.

Adventitious roots form from non-root tissues as part of normal development or in response to stress or wounding. The root primordia form in the source tissue, and during emergence the adventitious roots penetrate the inner cell layers and the epidermis; however, the mechanisms underlying this emergence remain largely unexplored. Here, we report that a regulatory module composed of the AP2/ERF transcription factor ABSCISIC ACID INSENSITIVE 4 (ABI4), the MAP kinases MPK3 and MPK6, and the phosphatase PP2C12 plays an important role in the emergence of junction adventitious roots (J-ARs) from the root-hypocotyl junctions in Arabidopsis thaliana. ABI4 negatively regulates J-AR emergence, preventing the accumulation of reactive oxygen species and death of epidermal cells, which would otherwise facilitate J-AR emergence. Phosphorylation by MPK3/MPK6 activates ABI4 and dephosphorylation by PP2C12 inactivates ABI4. MPK3/MPK6 also directly phosphorylate and inactivate PP2C12 during J-AR emergence. We propose that this "double-check" mechanism increases the robustness of MAP kinase signaling and finely regulates the local programmed cell death required for J-AR emergence.

GOLVEN peptide signalling through RGI receptors and MPK6 restricts asymmetric cell division during lateral root initiation.

During lateral root initiation, lateral root founder cells undergo asymmetric cell divisions that generate daughter cells with different sizes and fates, a prerequisite for correct primordium organogenesis. An excess of the GLV6/RGF8 peptide disrupts these initial asymmetric cell divisions, resulting in more symmetric divisions and the failure to achieve lateral root organogenesis. Here, we show that loss-of-function GLV6 and its homologue GLV10 increase asymmetric cell divisions during lateral root initiation, and we identified three members of the RGF1 INSENSITIVE/RGF1 receptor subfamily as likely GLV receptors in this process. Through a suppressor screen, we found that MITOGEN-ACTIVATED PROTEIN KINASE6 is a downstream regulator of the GLV pathway. Our data indicate that GLV6 and GLV10 act as inhibitors of asymmetric cell divisions and signal through RGF1 INSENSITIVE receptors and MITOGEN-ACTIVATED PROTEIN KINASE6 to restrict the number of initial asymmetric cell divisions that take place during lateral root initiation.

Upregulated-gene expression of pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) via TLRs following NF-κB and MAPKs in bovine mastitis.

Mastitis is the inflammation of mammary glands which causes huge economic loss in dairy cows. Inflammation, any tissue injury and pathogens in cow udder activate Toll-like Receptors (TLRs). Staphylococcus aureus (S. aureus) is the major cause of mastitis. In mastitis, activated TLRs initiate the NF-κB/MAPKs pathways which further trigger the gene expression associated with mastitis followed by innate immune response. In this study, pathogenic-induced gene expression profile of pro-inflammatory cytokines in mammary gland tissues, was investigated in mastitis. The Hematoxylin and Eosin (H & E) results indicated severe histopathological changes in infected tissues. Western blot results suggested the over expressions of TLR2/TLR4 with NF-κB/MAPKs pathways activation in infected tissues. qRT-PCR results revealed the gene expression associated with TLR2/TLR4-mediated NF-κB/MAPKs pathways in infected tissues in comparison with non-infected. Statistical analysis of mRNA and relative protein expression levels indicated the up-regulation of pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) in infected tissues rather than non-infected tissues. These results suggested that the up-regulation of gene expression levels implicated the underlying regulatory pathways for proper immune function in mammary glands. In conclusion, our study might give new insights for investigation and better understanding of mammary gland pathophysiology and TLRs and NF-κB/MAPKs-mediated gene expression of pro-inflammatory cytokines.

[Function and molecular mechanism of mitogen-activated protein kinase (MAPK) in regulating oocyte meiotic maturation and ovulation].

The mitogen-activated protein kinase (MAPK) signaling pathway is a highly conserved signal transduction pathway from yeast to human species, and is widely distributed in various eukaryotic cells. In almost all of the species studied over the past three decades, this signaling pathway plays a crucial role in the development of female germ cells and meiotic maturation. Especially in a variety of mammalian species including primates, rodents, and domestic animals, the MAPK signaling pathway is activated during the resumption of first oocyte meiosis and plays an indispensable role in meiotic spindle assembly and cell cycle progression. In granulosa cells of fully grown ovarian follicles, the MAPK pathway also mediates the physiological action of gonadotropins, including cumulus expansion, ovulation, and corpus luteum formation. Although the MAPK signaling pathway plays a wide range of physiological functions during the female reproduction process, and these functions are highly conserved in evolution, their underlying mechanisms, especially their direct and physiological target molecules, have not been sufficiently studied for a long time. In recent years, based on some new gene-editing mouse models and theoretical findings, as well as the wide application of various omics techniques, it has been further revealed that MAPK directly phosphorylates and activates the RNA binding protein cytoplasmic polyadenylation element-binding protein-1 (CPEB1), promoting poly(A) tail extension of maternal mRNA to regulate protein translation during meiotic recovery. These findings not only constitute the current basic mechanism of mammalian oocyte maturation and ovulation, but also provide useful research ideas for other related research in this field. In this review, we summarize the research findings in our laboratory and from other groups regarding the role of MAPK cascade in regulating oocyte maturation and ovulation. We also discuss the latest research progress on MAPK regulation of mRNA translation and degradation by directly activating the translation initiation complex and mRNA poly(A) polymerase by phosphorylation in the granulosa cells.

Two protein kinases UvPmk1 and UvCDC2 with significant functions in conidiation, stress response and pathogenicity of rice false smut fungus Ustilaginoidea virens.

Ustilaginoidea virens is an important fungus causing rice false smut, a devastating disease on spikelets of rice. In this study, we identified and characterized two CMGC (CDK/MAPK/GSK3/CLK) kinase genes, UvPmk1 and UvCDC2, in U. virens. Although UvPmk1 and UvCDC2 are, respectively, homologous to Fus3/Kss1 mitogen-activated protein kinases (MAPKs) and cyclin-dependent kinases (CDKs), they all have a conserved serine/threonine protein kinase domain. The qRT-PCR analysis of the relative expression of UvPmk1 and UvCDC2 during the infection of U. virens showed that these two genes were highly expressed during infection. UvPmk1 and UvCDC2 knockout mutants exhibited no significant changes in mycelial vegetative growth but decreases in conidiation. In addition, both UvPmk1 and UvCDC2 knockout mutants showed increases in tolerance to hyperosmotic and cell wall stresses, but they, respectively, exhibited decreases and increases in tolerance to oxidative stress compared with the wild-type strain HWD-2. Pathogenicity and infection assays demonstrated the defective growth of infection hyphae and significant loss of virulence in UvPmk1 and UvCDC2 knockout mutants. Taken together, our results demonstrate that UvPmk1 and UvCDC2 play important roles in the conidiation, stress response, and pathogenicity of U. virens.

AflSte20 Regulates Morphogenesis, Stress Response, and Aflatoxin Biosynthesis of Aspergillus flavus.

Various signaling pathways in filamentous fungi help cells receive and respond to environmental information. Previous studies have shown that the mitogen-activated protein kinase (MAPK) pathway is phosphorylation-dependent and activated by different kinase proteins. Serine/threonine kinase plays a very important role in the MAPK pathway. In this study, we selected the serine/threonine kinase AflSte20 in Aspergillus flavus for functional study. By constructing Aflste20 knockout mutants and complemented strains, it was proven that the Aflste20 knockout mutant (ΔAflste20) showed a significant decrease in growth, sporogenesis, sclerotinogenesis, virulence, and infection compared to the WT (wild type) and complemented strain (ΔAflste20C). Further research indicated that ΔAflste20 has more sensitivity characteristics than WT and ΔAflste20C under various stimuli such as osmotic stress and other types of environmental stresses. Above all, our study showed that the mitogen-activated kinase AflSte20 plays an important role in the growth, conidia production, stress response and sclerotia formation, as well as aflatoxin biosynthesis, in A. flavus.

EGF suppresses the expression of miR-124a in pancreatic ß cell lines via ETS2 activation through the MEK and PI3K signaling pathways.

Diabetes mellitus is characterized by pancreatic ß cell dysfunction. Previous studies have indicated that epidermal growth factor (EGF) and microRNA-124a (miR-124a) play opposite roles in insulin biosynthesis and secretion by beta cells. However, the underlying mechanisms remain poorly understood. In the present study, we demonstrated that EGF could inhibit miR-124a expression in beta cell lines through downstream signaling pathways, including mitogen-activated protein kinase kinase (MEK) and phosphatidylinositol 3-kinase (PI3K) cascades. Further, the transcription factor ETS2, a member of the ETS (E26 transformation-specific) family, was identified to be responsible for the EGF-mediated suppression of miR-124a expression, which was dependent on ETS2 phosphorylation at threonine 72. Activation of ETS2 decreased miR-124a promoter transcriptional activity through the putative conserved binding sites AGGAANA/TN in three miR-124a promoters located in different chromosomes. Of note, ETS2 played a positive role in regulating beta cell function-related genes, including miR-124a targets, Forkhead box a2 (FOXA2) and Neurogenic differentiation 1 (NEUROD1), which may have partly been through the inhibition of miR-124 expression. Knockdown and overexpression of ETS2 led to the prevention and promotion of insulin biosynthesis respectively, while barely affecting the secretion ability. These results suggest that EGF may induce the activation of ETS2 to inhibit miR-124a expression to maintain proper beta cell functions and that ETS2, as a novel regulator of insulin production, is a potential therapeutic target for diabetes mellitus treatment.