RESUMEN
Decades ago, mitogen-promoted signaling duration and strength were observed to be sensed by the cell and to be critical for its decisions: to proliferate or differentiate. Landmark publications established the importance of mitogen signaling not only in the G1 cell cycle phase but also through the S and the G2/M transition. Despite these early milestones, how mitogen signal duration and strength, short and strong or weaker and sustained, control cell fate has been largely unheeded. Here, we center on cardinal signaling-related questions, including (i) how fluctuating mitogenic signals are converted into cell proliferation-differentiation decisions and (ii) why extended duration of weak signaling is associated with differentiation, while bursts of strong and short induce proliferation but, if too strong and long, induce irreversible senescence. Our innovative broad outlook harnesses cell biology and protein conformational ensembles, helping us to define signaling strength, clarify cell cycle decisions, and thus cell fate.
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Ciclo Celular , Diferenciación Celular , Transducción de Señal , Humanos , Animales , Mitógenos/metabolismo , Proliferación CelularRESUMEN
BACKGROUND: Mitogen-activated protein kinases (MAPKs) are ubiquitous signal transduction components in eukaryotes. In plants, MAPKs play an essential role in growth and development, phytohormone regulation, and abiotic stress responses. The typical recretohalophyte Limonium bicolor (Bunge) Kuntze has multicellular salt glands on its stems and leaves; these glands secrete excess salt ions from its cells to mitigate salt damage. The number, type, and biological function of L. bicolor MAPK genes are unknown. RESULTS: We identified 20 candidate L. bicolor MAPK genes, which can be divided into four groups. Of these 20 genes, 17 were anchored to 7 chromosomes, while LbMAPK18, LbMAPK19, and LbMAPK20 mapped to distinct scaffolds. Structure analysis showed that the predicted protein LbMAPK19 contains the special structural motif TNY in its activation loop, whereas the other LbMAPK members harbor the conserved TEY or TDY motif. The promoters of most LbMAPK genes carry cis-acting elements related to growth and development, phytohormones, and abiotic stress. LbMAPK1, LbMAPK2, LbMAPK16, and LbMAPK20 are highly expressed in the early stages of salt gland development, whereas LbMAPK4, LbMAPK5, LbMAPK6, LbMAPK7, LbMAPK11, LbMAPK14, and LbMAPK15 are highly expressed during the late stages. These 20 LbMAPK genes all responded to salt, drought and ABA stress. We explored the function of LbMAPK2 via virus-induced gene silencing: knocking down LbMAPK2 transcript levels in L. bicolor resulted in fewer salt glands, lower salt secretion ability from leaves, and decreased salt tolerance. The expression of several genes [LbTTG1 (TRANSPARENT TESTA OF GL1), LbCPC (CAPRICE), and LbGL2 (GLABRA2)] related to salt gland development was significantly upregulated in LbMAPK2 knockdown lines, while the expression of LbEGL3 (ENHANCER OF GL3) was significantly downregulated. CONCLUSION: These findings increase our understanding of the LbMAPK gene family and will be useful for in-depth studies of the molecular mechanisms behind salt gland development and salt secretion in L. bicolor. In addition, our analysis lays the foundation for exploring the biological functions of MAPKs in an extreme halophyte.
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Plumbaginaceae , Plumbaginaceae/metabolismo , Mitógenos/metabolismo , Estrés Salino/genética , Proteínas Quinasas Activadas por Mitógenos/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Estrés Fisiológico/genética , Reguladores del Crecimiento de las Plantas/metabolismo , Regulación de la Expresión Génica de las Plantas , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMEN
Mitogen-activated protein kinase (MAPK/MPK) cascades are key signaling modules that regulate plant immunity. ENHANCED DISEASE RESISTANCE1 (EDR1) encodes a Raf-like MAPK kinase kinase (MAPKKK) that negatively regulates plant defense in Arabidopsis (Arabidopsis thaliana). The enhanced resistance of edr1 requires MAPK KINASE4 (MKK4), MKK5, and MPK3. Although the edr1 mutant displays higher MPK3/6 activation, the mechanism by which plants increase MAPK cascade activation remains elusive. Our previous study showed that MAPKKK5 is phosphorylated at the Ser-90 residue in edr1 mutants. In this study, we demonstrated that the enhanced disease resistance of edr1 required MAPKKK5. Phospho-dead MAPKKK5S90A partially impaired the resistance of edr1, and the expression of phospho-mimetic MAPKKK5S90D in mapkkk5-2 resulted in enhanced resistance to the powdery mildew Golovinomyces cichoracearum strain UCSC1 and the bacterial pathogen Pseudomonas syringae pv. tomato (Pto) strain DC3000. Thus, Ser-90 phosphorylation in MAPKKK5 appears to play a crucial role in disease resistance. However, MAPKKK5-triggered cell death was not suppressed by EDR1. Furthermore, activated MPK3 phosphorylated the N terminus of MAPKKK5, and Ser-90 was one of the phosphorylated sites. Ser-90 phosphorylation increased MAPKKK5 stability, and EDR1 might negatively regulate MAPK cascade activation by suppressing the MPK3-mediated feedback regulation of MAPKKK5. Taken together, these results indicate that MPK3 phosphorylates MAPKKK5 to enhance MAPK cascade activation and disease resistance in edr1 mutants.
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Proteínas de Arabidopsis , Arabidopsis , Humanos , Resistencia a la Enfermedad/genética , Proteínas de Arabidopsis/metabolismo , MAP Quinasa Quinasa Quinasa 5/metabolismo , Mitógenos/metabolismo , Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas , Enfermedades de las Plantas/microbiologíaRESUMEN
In mammalian cells, the decision to proliferate is thought to be irreversibly made at the restriction point of the cell cycle1,2, when mitogen signalling engages a positive feedback loop between cyclin A2/cyclin-dependent kinase 2 (CDK2) and the retinoblastoma protein3-5. Contrary to this textbook model, here we show that the decision to proliferate is actually fully reversible. Instead, we find that all cycling cells will exit the cell cycle in the absence of mitogens unless they make it to mitosis and divide first. This temporal competition between two fates, mitosis and cell cycle exit, arises because cyclin A2/CDK2 activity depends upon CDK4/6 activity throughout the cell cycle, not just in G1 phase. Without mitogens, mitosis is only observed when the half-life of cyclin A2 protein is long enough to sustain CDK2 activity throughout G2/M. Thus, cells are dependent on mitogens and CDK4/6 activity to maintain CDK2 activity and retinoblastoma protein phosphorylation throughout interphase. Consequently, even a 2-h delay in a cell's progression towards mitosis can induce cell cycle exit if mitogen signalling is lost. Our results uncover the molecular mechanism underlying the restriction point phenomenon, reveal an unexpected role for CDK4/6 activity in S and G2 phases and explain the behaviour of all cells following loss of mitogen signalling.
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Ciclo Celular , Quinasa 4 Dependiente de la Ciclina , Quinasa 6 Dependiente de la Ciclina , Fase G2 , Fase S , Animales , Ciclina A2/metabolismo , Quinasa 2 Dependiente de la Ciclina/metabolismo , Quinasa 4 Dependiente de la Ciclina/deficiencia , Quinasa 4 Dependiente de la Ciclina/metabolismo , Mitógenos/deficiencia , Mitógenos/metabolismo , Mitosis , Fosforilación , Proteína de Retinoblastoma/química , Proteína de Retinoblastoma/metabolismo , Quinasa 6 Dependiente de la Ciclina/deficiencia , Quinasa 6 Dependiente de la Ciclina/metabolismo , Fase G1RESUMEN
BACKGROUND: Chromosome instability (CIN) with recurrent copy number alterations is a feature of many solid tumors, including glioblastoma (GBM), yet the genes that regulate cell division are rarely mutated in cancers. Here, we show that the brain-abundant mitogen, platelet-derived growth factor-A (PDGFA) fails to induce the expression of kinetochore and spindle assembly checkpoint genes leading to defective mitosis in neural progenitor cells (NPCs). METHODS: Using a recently reported in vitro model of the initiation of high-grade gliomas from murine NPCs, we investigated the immediate effects of PDGFA exposure on the nuclear and mitotic phenotypes and patterns of gene and protein expression in NPCs, a putative GBM cell of origin. RESULTS: NPCs divided abnormally in defined media containing PDGFA with P53-dependent effects. In wild-type cells, defective mitosis was associated with P53 activation and cell death, but in some null cells, defective mitosis was tolerated. Surviving cells had unstable genomes and proliferated in the presence of PDGFA accumulating random and clonal chromosomal rearrangements. The outcome of this process was a population of tumorigenic NPCs with recurrent gains and losses of chromosomal regions that were syntenic to those recurrently gained and lost in human GBM. By stimulating proliferation without setting the stage for successful mitosis, PDGFA-transformed NPCs lacking P53 function. CONCLUSIONS: Our work describes a mechanism of transformation of NPCs by a brain-associated mitogen, raising the possibility that the unique genomic architecture of GBM is an adaptation to defective mitosis that ensures the survival of affected cells.
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Glioblastoma , Células-Madre Neurales , Humanos , Animales , Ratones , Mitógenos/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Mitosis , Células-Madre Neurales/patología , Glioblastoma/patologíaRESUMEN
Novel radioprotectors are strongly demanded due to their numerous applications in radiobiology and biomedicine, e.g., for facilitating the remedy after cancer radiotherapy. Currently, cerium-containing nanomaterials are regarded as promising inorganic radioprotectors due to their unrivaled antioxidant activity based on their ability to mimic the action of natural redox enzymes like catalase and superoxide dismutase and to neutralize reactive oxygen species (ROS), which are by far the main damaging factors of ionizing radiation. The freshwater planarian flatworms are considered a promising system for testing new radioprotectors, due to the high regenerative potential of these species and an excessive amount of proliferating stem cells (neoblasts) in their bodies. Using planarian Schmidtea mediterranea, we tested CeO2 nanoparticles, well known for their antioxidant activity, along with much less studied CeF3 nanoparticles, for their radioprotective potential. In addition, both CeO2 and CeF3 nanoparticles improve planarian head blastema regeneration after ionizing irradiation by enhancing blastema growth, increasing the number of mitoses and neoblasts' survival, and modulating the expression of genes responsible for the proliferation and differentiation of neoblasts. The CeO2 nanoparticles' action stems directly from their redox activity as ROS scavengers, while the CeF3 nanoparticles' action is mediated by overexpression of "wound-induced genes" and neoblast- and stem cell-regulating genes.
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Cerio , Nanopartículas , Planarias , Animales , Rayos X , Mitógenos/metabolismo , Mediterranea/metabolismo , Antioxidantes/farmacología , Antioxidantes/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Cerio/farmacología , Planarias/genéticaRESUMEN
Human hematopoietic stem cells (HSCs), like their counterparts in mice, comprise a functionally and molecularly heterogeneous population of cells throughout life that collectively maintain required outputs of mature blood cells under homeostatic conditions. In both species, an early developmental change in the HSC population involves a postnatal switch from a state in which most of these cells exist in a rapidly cycling state and maintain a high self-renewal potential to a state in which the majority of cells are in a quiescent state with an overall reduced self-renewal potential. However, despite the well-established growth factor dependence of HSC proliferation, whether and how this mechanism of HSC regulation might be affected by aging has remained poorly understood. To address this knowledge gap, we isolated highly HSC-enriched CD34+CD38-CD45RA-CD90+CD49f+ (CD49f+) cells from cord blood, adult bone marrow, and mobilized peripheral blood samples obtained from normal humans spanning 7 decades of age and then measured their functional and molecular responses to growth factor stimulation in vitro and their regenerative activity in vivo in mice that had undergone transplantation. Initial experiments revealed that advancing donor age was accompanied by a significant and progressively delayed proliferative response but not the altered mature cell outputs seen in normal older individuals. Importantly, subsequent dose-response analyses revealed an age-associated reduction in the growth factor-stimulated proliferation of CD49f+ cells mediated by reduced activation of AKT and altered cell cycle entry and progression. These findings identify a new intrinsic, pervasive, and progressive aging-related alteration in the biological and signaling mechanisms required to drive the proliferation of very primitive, normal human hematopoietic cells.
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Células Madre Hematopoyéticas , Mitógenos , Adulto , Humanos , Animales , Ratones , Integrina alfa6/metabolismo , Mitógenos/metabolismo , Células Madre Hematopoyéticas/metabolismo , Antígenos CD34/metabolismo , Proliferación Celular , Puntos de Control del Ciclo Celular , Ciclo Celular , Péptidos y Proteínas de Señalización Intercelular/metabolismoRESUMEN
PROBLEM: Amniochorion senescence generates mechanistic signals to initiate parturition. Activation of p38 mitogen-activated kinase (MAPK) in fetal amnion cells is a key mediator of senescence as well as epithelial-mesenchymal transition (EMT) of amnion cells. However, the impact of p38 MAPK in chorion trophoblast cells (CTCs) is unclear. We tested if eliminating p38 will reduce oxidative stress (OS) induced cell fates like cellular senescence, EMT, and inflammation induced by these processes in CTCs. METHODS: p38MAPK in CTCs was silenced using CRISPR/Cas9. OS was evoked by cigarette smoke extract (CSE) exposure. EMT was evoked by transforming growth factor (TGF)-ß treatment. Cell cycle, senescence, EMT, and inflammation were analyzed. RESULTS: CSE-induced changes in the cell cycle were not seen in p38KO CTCs compared to WT cells. OS induced by CSE evoked senescence and senescence-associated secretory phenotype (SASP as indicated by IL-6 and IL-8 increase) in WT but not in p38MAPK KO CTCs. No changes were noted in HLA-G expression regardless of the status of p38MAPK. Neither CSE nor TGF-ß evoked EMT in either WT or p38 KO CTCs. CONCLUSION: Senescence and senescence-associated inflammation in human fetal CTCs are mediated by p38MAPK. Compared to amnion epithelial cells, CTCs are resistant to EMT. This refractoriness may help them to maintain the barrier functions at the choriodecidual interface.
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Mitógenos , Trofoblastos , Femenino , Humanos , Mitógenos/metabolismo , Trofoblastos/metabolismo , Células Epiteliales/fisiología , Senescencia Celular , Amnios/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Inflamación/metabolismoRESUMEN
Introduction: Insulin-like growth factor 2 (IGF2) mRNA has been found in human and mouse spermatozoa. It is currently unknown whether the IGF2 protein is expressed in human spermatozoa and, if so, its possible role in the cross-talk between germ and Sertoli cells (SCs) during spermatogenesis. Methods: To accomplish this, we analyzed sperm samples from four consecutive Caucasian men. Furthermore, to understand its role during the spermatogenetic process, porcine SCs were incubated with increasing concentrations (0.33, 3.33, and 10 ng/mL) of recombinant human IGF2 (rhIGF2) for 48 hours. Subsequently, the experiments were repeated by pre-incubating SCs with the non-competitive insulin-like growth factor 1 receptor (IGF1R) inhibitor NVP-AEW541. The following outcomes were evaluated: 1) Gene expression of the glial cell-line derived neurotrophic factor (GDNF), fibroblast growth factor 2 (FGF2), and stem cell factor (SCF) mitogens; 2) gene and protein expression of follicle-stimulating hormone receptor (FSHR), anti-Müllerian hormone (AMH), and inhibin B; 3) SC proliferation. Results: We found that the IGF2 protein was present in each of the sperm samples. IGF2 appeared as a cytoplasmic protein localized in the equatorial and post-acrosomal segment and with a varying degree of expression in each cell. In SCs, IGF2 significantly downregulated GDNF gene expression in a concentration-dependent manner. FGF2 and SCF were downregulated only by the highest concentration of IGF2. Similarly, IGF2 downregulated the FSHR gene and FSHR, AMH, and inhibin B protein expression. Finally, IGF2 significantly suppressed the SC proliferation rate. All these findings were reversed by pre-incubation with NVP-AEW541, suggesting an effect mediated by the interaction of IGF2 with the IGFR. Conclusion: In conclusion, sperm IGF2 seems to downregulate the expression of mitogens, which are known to be physiologically released by the SCs to promote gonocyte proliferation and spermatogonial fate adoption. These findings suggest the presence of paracrine regulatory mechanisms acting on the seminiferous epithelium during spermatogenesis, by which germ cells can influence the amount of mitogens released by the SCs, their sensitivity to FSH, and their rate of proliferation.
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Factor Neurotrófico Derivado de la Línea Celular Glial , Factor II del Crecimiento Similar a la Insulina , Células de Sertoli , Espermatogénesis , Animales , Humanos , Masculino , Hormona Antimülleriana/metabolismo , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Factor Neurotrófico Derivado de la Línea Celular Glial/genética , Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Factor II del Crecimiento Similar a la Insulina/metabolismo , Mitógenos/metabolismo , Semen , Células de Sertoli/metabolismo , Espermatogénesis/fisiología , Espermatogonias/metabolismo , Espermatozoides/metabolismo , PorcinosRESUMEN
The fission yeast mitogen-activated kinase (MAPK) Sty1 is essential for cell survival in response to different environmental insults. In unstimulated cells, Sty1 forms an inactive ternary cytoplasmatic complex with the MAPKK Wis1 and the MAPKAP kinase Srk1. Wis1 phosphorylates and activates Sty1, inducing the nuclear translocation of the complex. Once in the nucleus, Sty1 phosphorylates and activates Srk1, which in turns inhibits Cdc25 and cell cycle progression, before being degraded in a proteasome-dependent manner. In parallel, active nuclear Sty1 activates the transcription factor Atf1, which results in the expression of stress response genes including pyp2 (a MAPK phosphatase) and srk1. Despite its essentiality in response to stress, persistent activation of the MAPK pathway can be deleterious and induces cell death. Thus, timely pathway inactivation is essential to ensure an appropriate response and cell viability. Here, uncover a role for the MAPKAP kinase Srk1 as an essential component of a negative feedback loop regulating the Sty1 pathway through phosphorylation and inhibition of the Wis1 MAPKK. This feedback regulation by a downstream kinase in the pathway highlights an additional mechanism for fine-tuning of MAPK signaling. Thus, our results indicate that Srk1 not only facilitates the adaptation to stress conditions by preventing cell cycle progression, but also plays an instrumental role regulating the upstream kinases in the stress MAPK pathway.
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Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Retroalimentación , Regulación Fúngica de la Expresión Génica , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Mitógenos/metabolismo , Fosforilación , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/metabolismoRESUMEN
Liver regeneration is a well-known systemic homeostatic phenomenon. The N6-methyladenosine (m6A) modification pathway has been associated with liver regeneration and hepatocellular carcinoma. m6A methyltransferases, such as methyltransferase 3 (METTL3) and methyltransferase 14 (METTL14), are involved in the hepatocyte-specific-regenerative pathway. To illustrate the role of METTL14, secreted from non-parenchymal liver cells, in the initiation phase of liver regeneration, we performed 70% partial hepatectomy (PH) in Mettl14 heterozygous (HET) and wild-type (WT) mice. Next, we analyzed the ratio of liver weight to body weight and the expression of mitogenic stimulators derived from non-parenchymal liver cells. Furthermore, we evaluated the expression of cell cycle-related genes and the hepatocyte proliferation rate via MKI67-immunostaining. During regeneration after PH, the weight ratio was lower in Mettl14 HET mice compared to WT mice. The expressions of hepatocyte growth factor (HGF) and tumor necrosis factor (TNF)-α, mitogens derived from non-parenchymal liver cells that stimulate the cell cycle, as well as the expressions of cyclin B1 and D1, which regulate the cell cycle, and the number of MKI67-positive cells, which indicate proliferative hepatocyte in the late G1-M phase, were significantly reduced in Mettl14 HET mice 72 h after PH. Our findings demonstrate that global Mettl14 mutation may interrupt the homeostasis of liver regeneration after an acute injury like PH by restraining certain mitogens, such as HGF and TNF-α, derived from sinusoidal endothelial cells, stellate cells, and Kupffer cells. These results provide new insights into the role of METTL14 in the clinical treatment strategies of liver disease. [BMB Reports 2022; 55(12): 633-638].
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Regeneración Hepática , Metiltransferasas , Mitógenos , Animales , Ratones , Células Endoteliales , Hepatectomía , Hígado/metabolismo , Regeneración Hepática/fisiología , Mitógenos/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Metiltransferasas/genéticaRESUMEN
Paxillin is a focal adhesion-associated protein that functions as an adaptor to recruit diverse cytoskeleton and signaling molecules into a complex and plays a crucial role in several signaling pathways in mammal cells. However, paxillin-mediated signal pathways are largely unknown in phytopathogenic fungi. Previously, Pax1 of Magnaporthe oryzae (MoPax1), a paxillin-like protein, has been identified as a crucial pathogenicity determinant. Here, we report the identification of a mitogen-activated protein (MAP) kinase (MAPK) activator, Mka1 of M. oryzae (MoMka1), that physically interacts with MoPax1. Targeted gene deletion of MoMKA1 resulted in pleiotropic defects in aerial hyphal growth, conidiation, appressorium formation, and pathogenicity in M. oryzae. MoMka1 interacts with Mst50, an adaptor protein of the Mst11-Mst7-Pmk1 and Mck1-Mkk2-Mps1 cascades. Moreover, the phosphorylation levels of both Pmk1 and Mps1 in aerial hyphae of the ΔMomka1 mutant were significantly reduced, indicating that MoMka1 acts upstream from the MAPK pathways. Interestingly, we found that MoMka1 interacts with MoAtg6 and MoAtg13. Deletion of MoMKA1 led to impaired MoAtg13 phosphorylation and enhanced autophagic flux under nutrient-rich conditions, indicating that MoMka1 is required for regulation of autophagy in M. oryzae. Taken together, the paxillin MoPax1 may activate MAP kinase signaling pathways and autophagy through MAP kinase activator MoMka1 and play important roles during appressorium-mediated plant infection by the rice blast fungus. IMPORTANCE Paxillin, as an adaptor recruiting diverse cytoskeleton and signaling molecules into a complex, plays a crucial role in several signaling pathways in mammal cells. However, paxillin-mediated signal pathways are largely unknown in phytopathogenic fungi. Here, we identified that MoMka1 physically interacts with MoPax1. Furthermore, MoMka1 acts upstream from the MAPK pathways through interacting with Mst50, a key protein of the Mst11-Mst7-Pmk1 and Mck1-Mkk2-Mps1 cascades. Meanwhile, MoMka1 interacts with both MoAtg6 and MoAtg13 and controls autophagy initiation by influencing the phosphorylation level of MoAtg13. In summary, we describe a model in which MoPax1 activates MAP kinase signaling pathways and autophagy through MoMka1 during appressorium-mediated plant infection by M. oryzae.
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Sistema de Señalización de MAP Quinasas , Magnaporthe , Animales , Mitógenos/metabolismo , Paxillin/genética , Proteínas Fúngicas/genética , Magnaporthe/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Transducción de Señal , Autofagia , Mamíferos/metabolismo , Enfermedades de las Plantas/microbiología , Esporas Fúngicas , Regulación Fúngica de la Expresión GénicaRESUMEN
Cell wall integrity (CWI) maintenance is central for plant cells. Mechanical and chemical distortions, pH changes, and breakdown products of cell wall polysaccharides activate plasma membrane-localized receptors and induce appropriate downstream responses. Microbial interactions alter or destroy the structure of the plant cell wall, connecting CWI maintenance to immune responses. Cellulose is the major polysaccharide in the primary and secondary cell wall. Its breakdown generates short-chain cellooligomers that induce Ca2+-dependent CWI responses. We show that these responses require the malectin domain-containing CELLOOLIGOMER-RECEPTOR KINASE 1 (CORK1) in Arabidopsis and are preferentially activated by cellotriose (CT). CORK1 is required for cellooligomer-induced cytoplasmic Ca2+ elevation, reactive oxygen species (ROS) production, mitogen-associated protein kinase (MAPK) activation, cellulose synthase phosphorylation, and the regulation of CWI-related genes, including those involved in biosynthesis of cell wall material, secondary metabolites and tryptophan. Phosphoproteome analyses identified early targets involved in signaling, cellulose synthesis, the endoplasmic reticulum/Golgi secretory pathway, cell wall repair and immune responses. Two conserved phenylalanine residues in the malectin domain are crucial for CORK1 function. We propose that CORK1 is required for CWI and immune responses activated by cellulose breakdown products.
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Arabidopsis , Arabidopsis/metabolismo , Celulosa/metabolismo , Mitógenos/metabolismo , Fenilalanina/metabolismo , Proteínas Quinasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Triptófano/metabolismoRESUMEN
Forkhead box O transcription factors (FoxOs) play an important role in maintaining normal cell physiology by regulating survival, apoptosis, autophagy, oxidative stress, the development and maturation of T and B lymphocytes, and the secretion of inflammatory cytokines. Cell types whose functions are regulated by FoxOs include keratinocytes, mucosal dermis, neutrophils, macrophages, dendritic cells, tumor-infiltrating activated regulatory T (Tregs) cells, B cells, and natural killer (NK) cells. FoxOs plays a crucial role in physiological and pathological immune responses. FoxOs control the development and function of Foxp3+ Tregs. Treg cells and Th17 cells are subsets of CD4+ T cells, which play an essential role in immune homeostasis and infection. Dysregulation of the Th17/Treg cell balance has been implicated in the development and progression of several disorders, such as autoimmune diseases, inflammatory diseases, and cancer. In addition, FoxOs are stimulated by the mitogen-activated protein (MAP) kinase pathway and inhibited by the PI3 kinase/AKT pathway. Downstream target genes of FoxOs include pro-inflammatory signaling molecules (toll-like receptor (TLR) 2, TLR4, interleukin (IL)-1ß, and tumor necrosis factor (TNF)-α), chemokine receptors (CCR7 and CXCR2), B-cell regulators (APRIL and BLYS), T-regulatory modulators (Foxp3 and CTLA-4), and DNA repair enzymes (GADD45α). Here, we review the recent progress in our understanding of FoxOs as the key molecules involved in immune cell differentiation and its role in the initiation of autoimmune diseases caused by dysregulation of immune cell balance. Additionally, in various diseases, FoxOs act as a cancer repressor, and reviving the activity of FoxOs forces Tregs to egress from various tissues. However, FoxOs regulate the cytotoxicity of both CD8+ T and NK cells against tumor cells, aiding in the restoration of redox and inflammatory homeostasis, repair of the damaged tissue, and activation of immune cells. A better understanding of FoxOs regulation may help develop novel potential therapeutics for treating immune/oxidative stress-related diseases.
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Enfermedades Autoinmunes , Neoplasias , Enfermedades Autoinmunes/metabolismo , Antígeno CTLA-4/metabolismo , Citocinas/metabolismo , Factores de Transcripción Forkhead/metabolismo , Humanos , Interleucinas/metabolismo , Mitógenos/metabolismo , Neoplasias/metabolismo , Oxidación-Reducción , Estrés Oxidativo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores CCR7/metabolismo , Linfocitos T Reguladores , Receptor Toll-Like 2/metabolismo , Receptor Toll-Like 4/metabolismo , Factores de Necrosis Tumoral/metabolismoRESUMEN
BACKGROUND: Endogenous Jaagsiekte sheep retrovirus envelope protein (enJSRV-Env) plays an important role in trophoblast cell fusion in sheep. However, the underlying mechanism remains unclear. METHODS: Primary endometrial luminal epithelial cells (LECs) were isolated from the sheep uterus and cocultured with sheep trophoblast cells (STCs). Giemsa staining was conducted to count multinucleated cells in the coculture system. Gain- and loss-of-function assays were performed to explore the role of enJSRV-Env in trophoblast cell fusion in the coculture system. Co-immunoprecipitation and mass spectrometry were carried out to identify the interacting partner of enJSRV-Env in the cocultures. Western blot analysis were conducted to determine the activation of protein kinase A (PKA)/mitogen-activated extracellular signal-regulated kinase (MEK)/extracellular signal-regulated kinase 1/2 (ERK1/2) signaling. RESULTS: Primary LECs were identified by the expression of epithelial marker cytokeratin 18. Overexpression of enJSRV-Env promoted the formation of multinucleated cells in the coculture system. enJSRV-Env activated and physically interacted with PKA, along with the activation of MEK/ERK1/2 signaling. PKA inhibition completely reversed enJSRV-Env-induced MEK/ERK1/2 activation, and ERK1/2 inhibition abolished enJSRV-Env-induced formation of multinucleated cells in the coculture system. CONCLUSION: enJSRV-Env promotes trophoblast cell fusion in the sheep placenta by activating PKA/MEK/ERK1/2 signaling. This finding reveals a novel mechanism underlying the contribution of enJSRV-Env to trophoblast cell fusion during placental morphogenesis.
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Retrovirus Ovino Jaagsiekte , Animales , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Femenino , Retrovirus Ovino Jaagsiekte/metabolismo , Queratina-18/metabolismo , Sistema de Señalización de MAP Quinasas , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Mitógenos/metabolismo , Placenta/metabolismo , Embarazo , Ovinos , Trofoblastos/metabolismoRESUMEN
BACKGROUND: The migration, proliferation and apoptosis of vascular smooth muscle cells (VSMCs) are critical for plaque stability. WNT-inducible signalling pathway protein-1 (WISP1), a member of the CCN family of extracellular matrix proteins, can expedite the migration and proliferation of VSMCs. However, its underlying mechanism and relationship with atherosclerosis remain elusive. The relationship between WISP1 and apoptosis of VSMCs has not been determined previously. METHOD: In the study, we aimed to investigate the relationship between WISP1 and plaque stability and its related mechanism.ApoE-/- mice were divided following groups: the null lentivirus (NC), lentivirus WISP1 (IvWISP1) and WISP1-shRNA (shWISP1) groups. Immunofluorescence, Oil Red O and Masson's staining of the carotid arteries were performed. Transwell wound healing assay, CCK8 assay, and TdT-mediated dUTP nick-end labeling (TUNEL) staining were performed using VSMCs. The levels of WISP1, P38, C-Jun N-terminal kinase, extracellular signal-regulated kinase (ERK), mitogen-activated extracellular signal-regulated kinase (MEK), focal adhesion kinase (FAK), phosphatidylinositol 3-kinase (PI3K), Akt (also known as PKB, protein kinase B), mammalian target of rapamycin (mTOR), cleaved caspase3, Bcl2 and Bax were detected by western blotting. RESULTS: The relative area of lipids and monocytes/macrophages in the shWISP1 group increased compared with that of the NC group. However, the relative area of smooth muscle cell and collagen in the IvWISP1 group increased compared with that in the NC group. Therefore, WISP1 could stabilize atherosclerotic plaques. Besides, WISP1 accelerate the migration and proliferation of VSMCs via integrin α5ß1 and FAK/MEK/ERK signalling pathways. In addition, WISP1 can inhibit the apoptosis of VSMCs via the PI3K/Akt/mTOR pathway. CONCLUSION: WISP1 not only inhibits the apoptosis of VSMCs via the PI3K/Akt/mTOR pathway but also enhances the migration and proliferation of VSMCs via the integrin α5ß1 and FAK/MEK/ERK pathways. Therefore, WISP1 could enhance the stability of atherosclerotic plaques.
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Proteínas CCN de Señalización Intercelular , Quinasa 1 de Adhesión Focal , Placa Aterosclerótica , Proteínas Proto-Oncogénicas c-akt , Proteínas Proto-Oncogénicas , Animales , Apolipoproteínas E/genética , Proteínas CCN de Señalización Intercelular/genética , Proteínas CCN de Señalización Intercelular/metabolismo , Proliferación Celular , Células Cultivadas , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Quinasa 1 de Adhesión Focal/genética , Quinasa 1 de Adhesión Focal/metabolismo , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Integrina alfa5beta1/metabolismo , Sistema de Señalización de MAP Quinasas , Mamíferos/genética , Mamíferos/metabolismo , Ratones , Ratones Noqueados para ApoE , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Mitógenos/metabolismo , Miocitos del Músculo Liso/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Placa Aterosclerótica/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Neutrophils are the most prevalent immune cells in circulation, but the repertoire of canonical inflammasomes in neutrophils and their respective involvement in neutrophil IL-1ß secretion and neutrophil cell death remain unclear. Here, we show that neutrophil-targeted expression of the disease-associated gain-of-function Nlrp3A350V mutant suffices for systemic autoinflammatory disease and tissue pathology in vivo. We confirm the activity of the canonical NLRP3 and NLRC4 inflammasomes in neutrophils, and further show that the NLRP1b, Pyrin and AIM2 inflammasomes also promote maturation and secretion of interleukin (IL)-1ß in cultured bone marrow neutrophils. Notably, all tested canonical inflammasomes promote GSDMD cleavage in neutrophils, and canonical inflammasome-induced pyroptosis and secretion of mature IL-1ß are blunted in GSDMD-knockout neutrophils. In contrast, GSDMD is dispensable for PMA-induced NETosis. We also show that Salmonella Typhimurium-induced pyroptosis is markedly increased in Nox2/Gp91Phox -deficient neutrophils that lack NADPH oxidase activity and are defective in PMA-induced NETosis. In conclusion, we establish the canonical inflammasome repertoire in neutrophils and identify differential roles for GSDMD and the NADPH complex in canonical inflammasome-induced neutrophil pyroptosis and mitogen-induced NETosis, respectively.
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Trampas Extracelulares , Inflamasomas , Neutrófilos , Proteínas de Unión a Fosfato , Proteínas Citotóxicas Formadoras de Poros , Piroptosis , Animales , Inflamasomas/metabolismo , Interleucina-1beta/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones , Ratones Endogámicos C57BL , Mitógenos/metabolismo , NADP/metabolismo , NADPH Oxidasas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR , Neutrófilos/metabolismo , Proteínas de Unión a Fosfato/metabolismo , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Pirina/metabolismoRESUMEN
The mechanisms by which insulin activates the insulin receptor to promote metabolic processes and cellular growth are still not clear. Significant advances have been gained from recent structural studies in understanding how insulin binds to its receptor. However, the way in which specific interactions lead to either metabolic or mitogenic signalling remains unknown. Currently there are only a few examples of insulin receptor agonists that have biased signalling properties. Here we use novel insulin analogues that differ only in the chemical composition at the A6-A11 bond, as it has been changed to a rigid, non-reducible C=C linkage (dicarba bond), to reveal mechanisms underlying signaling bias. We show that introduction of an A6-A11 cis-dicarba bond into either native insulin or the basal/long acting insulin glargine results in biased signalling analogues with low mitogenic potency. This can be attributed to reduced insulin receptor activation that prevents effective receptor internalization and mitogenic signalling. Insight gained into the receptor interactions affected by insertion of an A6-A11 cis-dicarba bond will ultimately assist in the development of new insulin analogues for the treatment of diabetes that confer low mitogenic activity and therefore pose minimal risk of promoting cancer with long term use.
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Insulina , Receptor de Insulina , Disulfuros , Insulina/metabolismo , Péptidos y Proteínas de Señalización Intercelular , Mitógenos/metabolismo , Mitógenos/farmacología , Receptor IGF Tipo 1/metabolismo , Receptor de Insulina/metabolismoRESUMEN
BACKGROUND: In lipopolysaccharide-induced RAW264.7 cells, Aster tataricus (AT) inhibits the nuclear factor kappa-light-chain-enhancer of activated B cells and MAPKs pathways and critical pathways of osteoclast development and bone resorption. OBJECTIVES: This study examined how aster saponin A2 (AS-A2) isolated from AT affects the processes and function of osteoclastogenesis induced by receptor activator of nuclear factor kappa-B ligand (RANKL) in RAW264.7 cells and bone marrow macrophages (BMMs). METHODS: The cell viability, tartrate-resistant acid phosphatase staining, pit formation assay, polymerase chain reaction, and western blot were carried out to determine the effects of AS-A2 on osteoclastogenesis. RESULTS: In RAW264.7 and BMMs, AS-A2 decreased RANKL-initiated osteoclast differentiation in a concentration-dependent manner. In AS-A2-treated cells, the phosphorylation of ERK1/2, JNK, and p38 protein expression were reduced considerably compared to the control cells. In RAW264.7 cells, AS-A2 suppressed the RANKL-induced activation of osteoclast-related genes. During osteoclast differentiation, AS-A2 suppressed the transcriptional and translational expression of NFATc1 and c-Fos. AS-A2 inhibited osteoclast development, reducing the size of the bone resorption pit area. CONCLUSION: AS-A2 isolated from AT appears to be a viable therapeutic therapy for osteolytic illnesses, such as osteoporosis, Paget's disease, and osteogenesis imperfecta.
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Resorción Ósea , Saponinas , Animales , Células de la Médula Ósea/metabolismo , Resorción Ósea/genética , Resorción Ósea/metabolismo , Resorción Ósea/veterinaria , Diferenciación Celular , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/farmacología , Mitógenos/metabolismo , Mitógenos/farmacología , FN-kappa B/metabolismo , Factores de Transcripción NFATC/genética , Factores de Transcripción NFATC/metabolismo , Factores de Transcripción NFATC/farmacología , Osteoclastos/metabolismo , Osteogénesis/fisiología , Ligando RANK/metabolismo , Ligando RANK/farmacología , Saponinas/farmacología , Transducción de SeñalRESUMEN
Microglia are immune cells in the central nervous system (CNS) that participate in response to pathological process after ischemic injury. Non-mitogenic fibroblast growth factor 1 (nmFGF1) is an effective neuroprotective factor that is also known as a metabolic regulator. The present study aimed to investigate the effects and mechanism of the neuroprotective ability of nmFGF1 on microglia in mice after photothrombosis (PT) stroke model, to determine whether it could ameliorate ischemic injury in stroke experiment. We discovered that the intranasal administration of nmFGF1 reduced infarct size and ameliorated neurological deficits in behavioral assessment by regulating the secretion of proinflammatory and anti-inflammatory cytokines. Furthermore, in the in vitro experiments, we found that nmFGF1 regulated the expression levels of proinflammatory and anti-inflammatory cytokines in oxygen-glucose deprivation (OGD) and lipopolysaccharide (LPS) stimulation. Evidence have shown that when nuclear factor erythroid 2-related factor 2 (Nfr2) is activated, it inhibits nuclear factor-kappa B (NF-κB) activation to alleviate inflammation. Interestingly, nmFGF1 treatment in vivo remarkably inhibited NF-κB pathway activation and activated Nrf2 pathway. In addition, nmFGF1 and NF-κB inhibitor (BAY11-7082) inhibited NF-κB pathway in LPS-stimulated BV2 microglia. Moreover, in LPS-stimulated BV2 microglia, the anti-inflammatory effect produced by nmFGF1 was knocked down by Nrf2 siRNA. These results indicate that nmFGF1 promoted functional recovery in experimental stroke by modulating microglia/macrophage-mediated neuroinflammation via Nrf2 and NF-κB signaling pathways, making nmFGF1 a potential agent against ischemic stroke.