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Endometriosis is a hormone-dependent, chronic inflammatory condition that affects 5-10% of reproductive-aged women. It is a complex disorder characterized by the growth of endometrial-like tissue outside the uterus, which can cause chronic pelvic pain and infertility. Despite its prevalence, the underlying molecular mechanisms of this disease remain poorly understood. Current treatment options are limited and focus mainly on suppressing lesion activity rather than eliminating it entirely. Although endometriosis is generally considered a benign condition, substantial evidence suggests that it increases the risk of developing specific subtypes of ovarian cancer. The discovery of cancer driver mutations in endometriotic lesions indicates that endometriosis may share molecular pathways with cancer. Moreover, the application of single-cell and spatial genomics, along with the development of organoid models, has started to illuminate the molecular mechanisms underlying disease etiology. This review aims to summarize the key genetic mutations and alterations that drive the development and progression of endometriosis to malignancy. We also review the significant recent advances in the understanding of the molecular basis of the disorder, as well as novel approaches and in vitro models that offer new avenues for improving our understanding of disease pathology and for developing new targeted therapies.
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Endometriosis , Endometriosis/patología , Endometriosis/genética , Endometriosis/etiología , Endometriosis/metabolismo , Humanos , Femenino , Neoplasias Ováricas/patología , Neoplasias Ováricas/etiología , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Mutación , AnimalesRESUMEN
Pulmonary fibrosis is a chronic, progressive, irreversible lung disease characterized by fibrotic scarring in the lung parenchyma. This condition involves the excessive accumulation of extracellular matrix (ECM) due to the aberrant activation of myofibroblasts in the alveolar environment. Transforming growth factor beta (TGF-ß) signaling is a crucial driver of fibrogenesis because it promotes excessive ECM deposition, thereby leading to scar formation and lung damage. A primary target of TGF-ß signaling in fibrosis is Collagen Triple Helix Repeat Containing 1 (CTHRC1), a secreted glycoprotein that plays a pivotal role in ECM deposition and wound repair. TGF-ß transcriptionally regulates CTHRC1 in response to tissue injury and controls the wound healing response through functional activity. CTHRC1 may also play an essential role in re-establishing and maintaining tissue homeostasis after wound closure by modulating both the TGF-ß and canonical Wnt signaling pathways. This dual function suggests that CTHRC1 regulates tissue remodeling and homeostasis. However, deregulated CTHRC1 expression in pathogenic fibroblasts has recently emerged as a hallmark of fibrosis in multiple organs and tissues. This review highlights recent studies suggesting that CTHRC1 can serve as a diagnostic and prognostic biomarker for fibrosis in idiopathic pulmonary fibrosis, systemic sclerosis, and post-COVID-19 lung fibrosis. Notably, CTHRC1 expression is responsive to antifibrotic drugs that target the TGF-ß pathway, such as pirfenidone and bexotegrast, indicating its potential as a biomarker of treatment success. These findings suggest that CTHRC1 may present new opportunities for diagnosing and treating patients with lung fibrosis.
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Proteínas de la Matriz Extracelular , Fibroblastos , Fibrosis Pulmonar , Humanos , Fibroblastos/metabolismo , Fibroblastos/patología , Proteínas de la Matriz Extracelular/metabolismo , Fibrosis Pulmonar/patología , Fibrosis Pulmonar/metabolismo , Animales , Factor de Crecimiento Transformador beta/metabolismo , Matriz Extracelular/metabolismo , Fibrosis Pulmonar Idiopática/patología , Fibrosis Pulmonar Idiopática/metabolismoRESUMEN
Background. The objective of this systematic review and meta-analysis was to estimate the proportions of individuals infected with Campylobacter, Escherichia, Salmonella, Shigella, or Yersinia who develop reactive arthritis. Methods. A systematic review was conducted, encompassing English-language articles published before January 2024, sourced from the Embase, PubMed, Scopus, and Web of Science databases. This review included observational studies that reported the occurrence of reactive arthritis (ReA) among patients with Campylobacter, Escherichia, Salmonella, Shigella, or Yersinia infections. Data extraction was carried out independently by two reviewers. Subsequently, a random-effects meta-analysis was performed, with heterogeneity assessed using the I2 value. Additionally, meta-regression was employed to investigate the potential influence of study-level variables on the observed heterogeneity. Results. A total of 87 studies were identified; 23 reported on ReA development after Campylobacter infection, 7 reported on ReA after Escherichia infection, 30 reported ReA onset after salmonellosis, 14 reported ReA after shigellosis, and 13 reported ReA after Yersinia infection. The proportion of Campylobacter patients who developed ReA was 0.03 (95% CI [0.01, 0.06], I2 = 97.62%); the proportion of Escherichia patients who developed ReA was 0.01 (95% CI [0.00, 0.06], I2 = 92.78%); the proportion of Salmonella patients was 0.04 (95% CI [0.02, 0.08], I2 = 97.67%); the proportion of Shigella patients was 0.01 (95% CI [0.01, 0.03], I2 = 90.64%); and the proportion of Yersinia patients who developed ReA was 0.05 (95% CI [0.02, 0.13], I2 = 96%). Conclusion. A significant proportion of Salmonella, Shigella, and Yersinia cases resulted in ReA. Nonetheless, it is important to interpret the findings cautiously due to the substantial heterogeneity observed between studies.
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Introduction: Rheumatoid arthritis (RA) is an autoimmune disease of unknown etiology that leads to disability due to articular and extra-articular damage. RA prevalence is variable. The disease is most common among females with a 3 : 1 ratio. The interaction of environmental and host factors contributes to RA development. Currently, the genome-wide association studies (GWAS) give the opportunity to uncover the RA genetic background. Anticitrullinated peptide antibody (ACPA) is a highly specific RA antibody, associated with poor prognosis and severe course of RA, and regulated by numerous genes. Our study is aimed at investigating whether there are any clinical and genetic aspects correlate with ACPA presence in Kazakhstani patients with RA. Indeed, the available studies on this subject are focused on Caucasian and East Asian populations (mainly Japanese and Chinese), and there are scarce data from Central Asia. Methods: Our study included 70 RA patients. Patients' blood samples were collected and genotyped for 14 SNPs by real-time polymerase chain reaction (RT-PCR). General examination, anamnestic, and clinical and laboratory data collection were carried out. Statistical analysis was performed using R statistics. Results and Conclusion. Our study revealed a significant association of ACPA positivity with Fc receptor-like 3 (FCRL3) and ACPA negativity with signal transducer and activator of transcription 4 (STAT4) genes, but not with T cell activation Rho GTPase activating protein (TAGAP). In addition, ACPA positivity was associated with radiographic progression, rheumatoid factor (RF), erythrocyte sedimentation rate (ESR), age of RA onset, the patient global assessment, body mass index (BMI), and Gamma globulin. Conclusion: Remained 11 earlier identified significantly associated in Caucasian and Asian population SNPs were not replicated in our cohort. Further studies on larger cohorts are needed to confirm our findings with higher confidence levels and stronger statistical power.
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Artritis Reumatoide , Polimorfismo de Nucleótido Simple , Autoanticuerpos , Femenino , Estudio de Asociación del Genoma Completo , Humanos , Péptidos , Proyectos PilotoRESUMEN
The human endometrium is a unique tissue undergoing important changes through the menstrual cycle. Under the exposure of different risk factors in a woman's lifetime, normal endometrial tissue can give rise to multiple pathologic conditions, including endometriosis and endometrial cancer. Etiology and pathophysiologic changes behind such conditions remain largely unclear. This review summarizes the current knowledge of the pathophysiology of endometriosis and its potential role in the development of endometrial cancer from a molecular perspective. A better understanding of the molecular basis of endometriosis and its role in the development of endometrial pathology will improve the approach to clinical management.
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Neoplasias Endometriales/fisiopatología , Endometriosis/fisiopatología , Neoplasias Endometriales/diagnóstico , Neoplasias Endometriales/epidemiología , Neoplasias Endometriales/etiología , Endometriosis/complicaciones , Endometriosis/diagnóstico , Endometriosis/epidemiología , Femenino , Estudios de Asociación Genética , Humanos , Neoplasias Ováricas , Análisis de Secuencia de ADNRESUMEN
Rheumatoid arthritis (RA) is a chronic autoimmune disease causing inflammation of joints, cartilage destruction and bone erosion. Biomarkers and new drug targets are actively sought and progressed to improve available options for patient treatment. The Collagen Triple Helix Repeat Containing 1 protein (CTHRC1) may have an important role as a biomarker for rheumatoid arthritis, as CTHRC1 protein concentration is significantly elevated in the peripheral blood of rheumatoid arthritis patients compared to osteoarthritis (OA) patients and healthy individuals. CTHRC1 is a secreted glycoprotein that promotes cell migration and has been implicated in arterial tissue-repair processes. Furthermore, high CTHRC1 expression is observed in many types of cancer and is associated with cancer metastasis to the bone and poor patient prognosis. However, the function of CTHRC1 in RA is still largely undefined. The aim of this review is to summarize recent findings on the role of CTHRC1 as a potential biomarker and pathogenic driver of RA progression. We will discuss emerging evidence linking CTHRC1 to the pathogenic behavior of fibroblast-like synoviocytes and to cartilage and bone erosion through modulation of the balance between bone resorption and repair.
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Artritis Reumatoide/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Animales , Artritis Reumatoide/fisiopatología , Biomarcadores , Proteínas de la Matriz Extracelular/fisiología , Humanos , Sinoviocitos/metabolismo , Sinoviocitos/fisiología , Vía de Señalización WntRESUMEN
Spondyloarthritis comprises a group of inflammatory diseases of the joints and spine, with various clinical manifestations. The group includes ankylosing spondylitis, reactive arthritis, psoriatic arthritis, arthritis associated with inflammatory bowel disease, and undifferentiated spondyloarthritis. The exact etiology and pathogenesis of spondyloarthritis are still unknown, but five hypotheses explaining the pathogenesis exist. These hypotheses suggest that spondyloarthritis is caused by arthritogenic peptides, an unfolded protein response, HLA-B*27 homodimer formation, malfunctioning endoplasmic reticulum aminopeptidases, and, last but not least, gut inflammation and dysbiosis. Here we discuss the five hypotheses and the evidence supporting each. In all of these hypotheses, HLA-B*27 plays a central role. It is likely that a combination of these hypotheses, with HLA-B*27 taking center stage, will eventually explain the development of spondyloarthritis in predisposed individuals.
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Antígeno HLA-B27/inmunología , Inflamación/inmunología , Espondiloartritis/inmunología , Espondilitis Anquilosante/inmunología , Artritis Psoriásica/genética , Artritis Psoriásica/inmunología , Artritis Psoriásica/metabolismo , Artritis Psoriásica/patología , Artritis Reactiva/genética , Artritis Reactiva/inmunología , Artritis Reactiva/metabolismo , Artritis Reactiva/patología , Antígeno HLA-B27/genética , Humanos , Inflamación/genética , Inflamación/metabolismo , Inflamación/patología , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/patología , Articulaciones/inmunología , Articulaciones/patología , Columna Vertebral/inmunología , Columna Vertebral/patología , Espondiloartritis/genética , Espondiloartritis/metabolismo , Espondiloartritis/patología , Espondilitis Anquilosante/genética , Espondilitis Anquilosante/metabolismo , Espondilitis Anquilosante/patología , Respuesta de Proteína Desplegada/genética , Respuesta de Proteína Desplegada/inmunologíaRESUMEN
Background: The purpose of this study was to determine whether plasma levels of the collagen triple helix repeat containing 1 (CTHRC1) protein can serve as a blood-based biomarker for improved diagnosis of rheumatoid arthritis (RA) patients and monitoring of RA disease activity. Methods: We measured levels of CTHRC1 in the plasma of patients diagnosed with RA, osteoarthritis (OA), reactive arthritis (ReA), as well as in healthy individuals. We then assessed the correlation between CTHRC1 protein and a range of indices including the 28-joint disease activity score (DAS28), rheumatoid factor (RF), C-reactive protein (CRP), anti-citrullinated protein antibodies (ACPA), erythrocyte sedimentation rate (ESR), as well as a panel of cytokines, including interleukin 1 beta (IL-1ß), interleukin 6 (IL-6), interleukin 8 (IL-8), and interferon gamma (IFNγ). Receiver operating characteristic (ROC) analysis was further performed to assess the diagnostic value of CTHRC1. Results: CTHRC1 plasma levels were significantly elevated in RA patients compared to healthy individuals, OA and ReA patients. ROC curve and risk score analysis suggested that plasma CTHRC1 can accurately discriminate patients with RA from healthy controls and may have practical value for RA diagnosis. CTHRC1 levels were positively associated with RF, ACPA, CRP, and disease activity based on the combined index of DAS28 with CRP (DAS28-CRP), and also strongly correlated with IL-1ß, IL-6, IL-8, and IFNγ. Conclusion: Our studies show that CTHRC1 is a sensitive and easy-to-measure plasma marker that differentiates between RA and healthy status and also distinguishes between RA and other forms of arthritis, such as OA and ReA. At the current level of understanding, plasma CTHRC1 levels may improve the diagnosis of RA and these findings warrant confirmation in a larger, more comprehensive patient population.
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Artritis Reumatoide/diagnóstico , Biomarcadores/sangre , Proteínas de la Matriz Extracelular/sangre , Osteoartritis/diagnóstico , Adulto , Anciano , Anticuerpos Antiproteína Citrulinada/metabolismo , Citocinas/metabolismo , Progresión de la Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Prohibitinas , Índice de Severidad de la Enfermedad , Regulación hacia ArribaRESUMEN
Liver fibrosis is the result of a deregulated wound healing process characterized by the excessive deposition of extracellular matrix. Hepatic stellate cells (HSCs), which are activated in response to liver injury, are the major source of extracellular matrix and drive the wound healing process. However, chronic liver damage leads to perpetual HSC activation, progressive formation of pathological scar tissue and ultimately, cirrhosis and organ failure. HSC activation is triggered largely in response to mechanosignaling from the microenvironment, which induces a profibrotic nuclear transcription program that promotes HSC proliferation and extracellular matrix secretion thereby setting up a positive feedback loop leading to matrix stiffening and self-sustained, pathological, HSC activation. Despite the significant progress in our understanding of liver fibrosis, the molecular mechanisms through which the extracellular matrix promotes HSC activation are not well understood and no effective therapies have been approved to date that can target this early, reversible, stage in liver fibrosis. Several new lines of investigation now provide important insight into this area of study and identify two nuclear targets whose inhibition has the potential of reversing liver fibrosis by interfering with HSC activation: Yes-associated protein (YAP), a transcriptional co-activator and effector of the mechanosensitive Hippo pathway, and bromodomain-containing protein 4 (BRD4), an epigenetic regulator of gene expression. YAP and BRD4 activity is induced in response to mechanical stimulation of HSCs and each protein independently controls waves of early gene expression necessary for HSC activation. Significantly, inhibition of either protein can revert the chronic activation of HSCs and impede pathological progression of liver fibrosis in clinically relevant model systems. In this review we will discuss the roles of these nuclear co-activators in HSC activation, their mechanism of action in the fibrotic process in the liver and other organs, and the potential of targeting their activity with small molecule drugs for fibrosis reversal.
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Phosphatidylinositol-4,5-bisphosphate (PI4,5P(2)) is a critical regulator of cell migration, but the roles of the type I phosphatidylinositol-4-phosphate 5-kinases (PIPKIs), which synthesize PI4,5P(2), have yet to be fully defined in this process. In this study, we report that one kinase, PIPKI-alpha, is a novel upstream regulator of Rac1 that links activated integrins to the regulation of cell migration. We show that PIPKI-alpha controls integrin-induced translocation of Rac1 to the plasma membrane and thereby regulates Rac1 activation. Strikingly, this function is not shared with other PIPKI isoforms, is independent of catalytic activity, and requires physical interaction of PIPKI-alpha with the Rac1 polybasic domain. Consistent with its role in Rac1 activation, depletion of PIPKI-alpha causes pronounced defects in membrane ruffling, actin organization, and focal adhesion formation, and ultimately affects the directional persistence of migration. Thus, our study defines the role of PIPKI-alpha in cell migration and describes a new mechanism for the spatial regulation of Rac1 activity that is critical for cell migration.
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Membrana Celular/metabolismo , Movimiento Celular/fisiología , Isoenzimas/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Actinas/metabolismo , Animales , Adhesión Celular/fisiología , Membrana Celular/ultraestructura , Activación Enzimática , Células HeLa , Humanos , Integrinas/metabolismo , Isoenzimas/genética , Fosfatidilinositol 4,5-Difosfato/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Estructura Terciaria de Proteína , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Proteína de Unión al GTP rac1/genéticaRESUMEN
Cell migration requires the regulated disassembly of focal adhesions, but the underlying mechanisms remain poorly defined. We have previously shown that focal adhesion disassembly requires the dynamin 2- and clathrin-dependent endocytosis of ligand-activated beta1 integrins. Here, we identify type I phosphatidylinositol phosphate kinase beta (PIPKIbeta), an enzyme that generates phosphatidylinositol-4,5-bisphosphate (PI4,5P(2)), as a key regulator of this process. We found that knockdown of PIPKIbeta by RNA interference blocks the internalization of active beta1 integrins and impairs focal adhesion turnover and cell migration. These defects are caused by the failure to target the endocytic machinery, including clathrin adaptors and dynamin 2, to focal adhesion sites. As a consequence, depletion of PIPKIbeta blocks clathrin assembly at adhesion plaques and prevents complex formation between dynamin 2 and focal adhesion kinase (FAK), a critical step in focal adhesion turnover. Together, our findings identify PIPKIbeta as a novel regulator of focal adhesion disassembly and suggest that PIPKIbeta spatially regulates integrin endocytosis at adhesion sites to control cell migration.
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Endocitosis/fisiología , Adhesiones Focales/metabolismo , Integrina beta1/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proteínas Reguladoras de la Apoptosis , Adhesión Celular/fisiología , Línea Celular , Movimiento Celular/fisiología , Clatrina/metabolismo , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , Dinamina II/genética , Dinamina II/metabolismo , Proteína-Tirosina Quinasas de Adhesión Focal/genética , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Glicoproteínas/genética , Glicoproteínas/metabolismo , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Interferencia de ARN , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Supresoras de Tumor , ZixinaRESUMEN
Cell migration requires the controlled disassembly of focal adhesions, but the underlying mechanisms remain poorly understood. Here, we show that adhesion turnover is mediated through dynamin- and clathrin-dependent endocytosis of activated beta1 integrins. Consistent with this, clathrin and the clathrin adaptors AP-2 and disabled-2 (DAB2) distribute along with dynamin 2 to adhesion sites prior to adhesion disassembly. Moreover, knockdown of either dynamin 2 or both clathrin adaptors blocks beta1 integrin internalization, leading to impaired focal adhesion disassembly and cell migration. Together, these results provide important insight into the mechanisms underlying adhesion disassembly and identify novel components of the disassembly pathway.
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Clatrina/fisiología , Endocitosis/fisiología , Integrinas/fisiología , Línea Celular Tumoral , Técnica del Anticuerpo Fluorescente , Humanos , ARN Interferente PequeñoRESUMEN
Angiogenesis plays a key role in promoting tumorigenesis and metastasis. The 16-kDa fragment of prolactin (16k PRL) is an NH(2)-terminal natural breakdown fragment of the intact 23-kDa prolactin and has been shown to have potent antiangiogenic and antitumor activities. The mechanism(s) involved in the action of 16k PRL in endothelial cells remains unclear. In this study, we showed that 16k PRL reduced rat aortic endothelial cell (RAEC) migration in a wound-healing assay and in a Matrigel tube formation assay, suggesting that 16k PRL inhibits endothelial cell migration, an important activity involved in angiogenesis and tumorigenesis. We further investigated how 16k PRL attenuates endothelial cell migration. We first showed that RAEC migration is mediated through the Rho GTPase Rac1, as Rac1 inhibition by the Rac1-specific inhibitor NSC27366 or Rac1 knockdown by small interfering RNA both blocked RAEC migration. We next showed that 16k PRL reduced the activation of Rac1 in a concentration-dependent manner. Furthermore, 16k PRL inhibition of Rac1 is mediated through the suppression of T lymphoma invasion and metastasis 1 (Tiam1) and its upstream activator Ras in a phosphoinositide-3-kinase-independent manner. 16k PRL also down-regulated the phosphorylation of the downstream effector of Rac1, p21-activating kinase 1 (Pak1), and inhibited its translocation to the leading edge of migrating cells. Thus, 16k PRL inhibits cell migration by blocking the Ras-Tiam1-Rac1-Pak1 signaling pathway in endothelial cells.
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Regulación hacia Abajo , Células Endoteliales/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Proteínas de Neoplasias/metabolismo , Prolactina/fisiología , Quinasas p21 Activadas/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Proteínas ras/metabolismo , Inhibidores de la Angiogénesis/uso terapéutico , Animales , Movimiento Celular , Humanos , Prolactina/metabolismo , Ratas , Proteína 1 de Invasión e Inducción de Metástasis del Linfoma-TRESUMEN
The PH domain-containing proteins Slm1 and Slm2 were previously identified as effectors of the phosphatidylinositol-4,5-bisphosphate (PI4,5P(2)) and TORC2 signaling pathways. Here, we demonstrate that Slm1 and Slm2 are also targets of sphingolipid signaling during the heat shock response. We show that upon depletion of cellular sphingolipid levels, Slm1 function becomes essential for survival under heat stress. We further demonstrate that Slm proteins are regulated by a phosphorylation/dephosphorylation cycle involving the sphingolipid-activated protein kinases Pkh1 and Pkh2 and the calcium/calmodulin-dependent protein phosphatase calcineurin. By using a combination of mass spectrometry and mutational analysis, we identified serine residue 659 in Slm1 as a site of phosphorylation. Characterization of Slm1 mutants that mimic dephosphorylated and phosphorylated states demonstrated that phosphorylation at serine 659 is vital for survival under heat stress and promotes the proper polarization of the actin cytoskeleton. Finally, we present evidence that Slm proteins are also required for the trafficking of the raft-associated arginine permease Can1 to the plasma membrane, a process that requires sphingolipid synthesis and actin polymerization. Together with previous work, our findings suggest that Slm proteins are subject to regulation by multiple signals, including PI4,5P(2), TORC2, and sphingolipids, and may thus integrate inputs from different signaling pathways to temporally and spatially control actin polarization.
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Proteínas Portadoras/metabolismo , Membrana Celular/fisiología , Proteínas de Unión al ARN/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Esfingolípidos/fisiología , Proteínas Quinasas Dependientes de 3-Fosfoinosítido , Sistemas de Transporte de Aminoácidos Básicos/metabolismo , Calcineurina/metabolismo , Proteínas Portadoras/genética , Proteínas del Citoesqueleto , Ácidos Grasos Monoinsaturados/farmacología , Respuesta al Choque Térmico , Microdominios de Membrana/metabolismo , Fosforilación , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas , Estructura Terciaria de Proteína , Transporte de Proteínas , Proteínas de Unión al ARN/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Transducción de SeñalRESUMEN
Dendritic arborization and spine formation are critical for the functioning of neurons. Although many proteins have been identified recently as regulators of dendritic morphogenesis, the intracellular signaling pathways that control these processes are not well understood. Here we report that the Ras-phosphatidylinositol 3-kinase (PI3K)-Akt-mammalian target of rapamycin (mTOR) signaling pathway plays pivotal roles in the regulation of many aspects of dendrite formation. Whereas the PI3K-Akt-mTOR pathway alone controlled soma and dendrite size, a coordinated activation together with the Ras-mitogen-activated protein kinase signaling pathway was required for increasing dendritic complexity. Chronic inhibition of PI3K or mTOR reduced soma and dendrite size and dendritic complexity, as well as density of dendritic filopodia and spines, whereas a short-term inhibition promoted the formation of mushroom-shaped spines on cells expressing constitutively active mutants of Ras, PI3K, or Akt, or treated with the upstream activator BDNF. Together, our data underscore the central role of a spatiotemporally regulated key cell survival and growth pathway on trophic regulation of the coordinated development of dendrite size and shape.
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Dendritas/fisiología , Quinasas de Proteína Quinasa Activadas por Mitógenos/fisiología , Fosfatidilinositol 3-Quinasas/fisiología , Proteínas Quinasas/fisiología , Proteínas Proto-Oncogénicas c-akt/fisiología , Proteínas ras/fisiología , Animales , Células Cultivadas , Dendritas/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Activación Enzimática/fisiología , Inhibidores Enzimáticos/farmacología , Técnicas In Vitro , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Morfogénesis/efectos de los fármacos , Morfogénesis/fisiología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Ratas , Transducción de Señal/fisiología , Serina-Treonina Quinasas TORRESUMEN
The V(0) complex forms the proteolipid pore of an ATPase that acidifies vesicles. In addition, an independent function in membrane fusion has been proposed largely based on yeast vacuolar fusion experiments. We have isolated mutations in the largest V(0) component vha100-1 in flies in an unbiased genetic screen for synaptic malfunction. The protein is only required in neurons, colocalizes with markers for synaptic vesicles as well as active zones, and interacts with t-SNAREs. Loss of vha100-1 leads to vesicle accumulation in synaptic terminals, suggesting a deficit in release. The amplitude of spontaneous release events and release with hypertonic stimulation indicate normal levels of neurotransmitter loading, yet mutant embryos display severe defects in evoked synaptic transmission and FM1-43 uptake. Our data suggest that Vha100-1 functions downstream of SNAREs in synaptic vesicle fusion.
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Drosophila melanogaster/metabolismo , Exocitosis/fisiología , Fusión de Membrana/fisiología , Membranas Sinápticas/metabolismo , Vesículas Sinápticas/metabolismo , ATPasas de Translocación de Protón Vacuolares/metabolismo , Animales , Drosophila melanogaster/ultraestructura , Embrión no Mamífero/metabolismo , Embrión no Mamífero/ultraestructura , Anomalías del Ojo/genética , Soluciones Hipertónicas/farmacología , Microscopía Electrónica , Mutación/genética , Células Fotorreceptoras de Invertebrados/anomalías , Células Fotorreceptoras de Invertebrados/ultraestructura , Subunidades de Proteína/genética , Subunidades de Proteína/metabolismo , Compuestos de Piridinio/farmacocinética , Compuestos de Amonio Cuaternario/farmacocinética , Proteínas SNARE , Membranas Sinápticas/ultraestructura , Transmisión Sináptica/genética , Vesículas Sinápticas/ultraestructura , ATPasas de Translocación de Protón Vacuolares/genética , Proteínas de Transporte Vesicular/metabolismoRESUMEN
Phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P(2)] is a key second messenger that regulates actin and membrane dynamics, as well as other cellular processes. Many of the effects of PtdIns(4,5)P(2) are mediated by binding to effector proteins that contain a pleckstrin homology (PH) domain. Here, we identify two novel effectors of PtdIns(4,5)P(2) in the budding yeast Saccharomyces cerevisiae: the PH domain containing protein Slm1 and its homolog Slm2. Slm1 and Slm2 serve redundant roles essential for cell growth and actin cytoskeleton polarization. Slm1 and Slm2 bind PtdIns(4,5)P(2) through their PH domains. In addition, Slm1 and Slm2 physically interact with Avo2 and Bit61, two components of the TORC2 signaling complex, which mediates Tor2 signaling to the actin cytoskeleton. Together, these interactions coordinately regulate Slm1 targeting to the plasma membrane. Our results thus identify two novel effectors of PtdIns(4,5)P(2) regulating cell growth and actin organization and suggest that Slm1 and Slm2 integrate inputs from the PtdIns(4,5)P(2) and TORC2 to modulate polarized actin assembly and growth.
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Actinas/metabolismo , Proteínas Portadoras/metabolismo , Citoesqueleto/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Proteínas de Unión al ARN/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , 1-Fosfatidilinositol 4-Quinasa/genética , 1-Fosfatidilinositol 4-Quinasa/metabolismo , Secuencia de Aminoácidos , Proteínas Sanguíneas/química , Proteínas Portadoras/química , Proteínas Portadoras/genética , Membrana Celular/metabolismo , Proteínas del Citoesqueleto , Citoesqueleto/química , Datos de Secuencia Molecular , Mutación/genética , Fosfatidilinositol 4,5-Difosfato/biosíntesis , Fosfoproteínas/química , Fosfotransferasas/genética , Fosfotransferasas/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol) , Unión Proteica , Proteína Quinasa C/genética , Proteína Quinasa C/metabolismo , Estructura Terciaria de Proteína , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/genética , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Alineación de Secuencia , Proteína de Unión al GTP cdc42 de Saccharomyces cerevisiae/genética , Proteína de Unión al GTP cdc42 de Saccharomyces cerevisiae/metabolismo , Proteínas de Unión al GTP rho/genética , Proteínas de Unión al GTP rho/metabolismoRESUMEN
Type I and type II phosphatidylinositol phosphate (PIP) kinases generate the lipid second messenger phosphatidylinositol (PtdIns) 4,5-bisphosphate and thus play fundamental roles in the regulation of many cellular processes. Although the two kinase families are highly homologous, they phosphorylate distinct substrates and are functionally non-redundant. Type I PIP kinases phosphorylate PtdIns 4-phosphate at the D-5 hydroxyl group and are consequently PtdIns 4-phosphate 5-kinases. By contrast, type II PIP kinases are PtdIns 5-phosphate 4-kinases that phosphorylate PtdIns 5-phosphate at the D-4 position. Type I PIP kinases, in addition, also phosphorylate other phosphoinositides in vitro and in vivo and thus have the potential to generate multiple lipid second messengers. To understand how these enzymes differentiate between stereoisomeric substrates, we used a site-directed mutagenesis approach. We show that a single amino acid substitution in the activation loop, A381E in IIbeta and the corresponding mutation E362A in Ibeta, is sufficient to swap substrate specificity between these PIP kinases. In addition to its role in substrate specificity, the type I activation loop is also key in subcellular targeting. The Ibeta(E362A) mutant and other mutants with reduced PtdIns 4-phosphate binding affinity were largely cytosolic when expressed in mammalian cells in contrast to wild-type Ibeta which targets to the plasma membrane. These results clearly establish the role of the activation loop in determining both signaling specificity and plasma membrane targeting of type I PIP kinases.