RESUMEN
Isoforms of microtubule-associated protein 2 (MAP2) differ from their homolog Tau in the sequence and interactions of the N-terminal region. Binding of the N-terminal region of MAP2c (N-MAP2c) to the dimerization/docking domains of the regulatory subunit RIIα of cAMP-dependent protein kinase (RIIDD2) and to the Src-homology domain 2 (SH2) of growth factor receptor-bound protein 2 (Grb2) have been described long time ago. However, the structural features of the complexes remained unknown due to the disordered nature of MAP2. Here, we provide structural description of the complexes. We have solved solution structure of N-MAP2c in complex with RIIDD2, confirming formation of an amphiphilic α-helix of MAP2c upon binding, defining orientation of the α-helix in the complex and showing that its binding register differs from previous predictions. Using chemical shift mapping, we characterized the binding interface of SH2-Grb2 and rat MAP2c phosphorylated by the tyrosine kinase Fyn in their complex and proposed a model explaining differences between SH2-Grb2 complexes with rat MAP2c and phosphopeptides with a Grb2-specific sequence. The results provide the structural basis of a potential role of MAP2 in regulating cAMP-dependent phosphorylation cascade via interactions with RIIDD2 and Ras signaling pathway via interactions with SH2-Grb2.
Asunto(s)
Proteína Adaptadora GRB2 , Proteínas Asociadas a Microtúbulos , Unión Proteica , Proteína Adaptadora GRB2/metabolismo , Proteína Adaptadora GRB2/química , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/química , Proteínas Asociadas a Microtúbulos/genética , Humanos , Transducción de Señal , Animales , Dominios Homologos src , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas c-fyn/química , Proteínas Proto-Oncogénicas c-fyn/genética , Dominios ProteicosRESUMEN
Microglia-mediated neuroinflammation is closely associated with many neurodegenerative diseases. Psoralen has potential for the treatment of many diseases, however, the anti-neuroinflammatory and neuroprotective effects of psoralen have been unclear. This study investigated the anti-neuroinflammatory and neuroprotective effects of psoralen and its regulation of microglial M1/M2 polarization. The LPS-induced mice model was used to test anti-neuroinflammatory effects, regulatory effects on microglia polarization, and neuroprotective effects of psoralen in vivo. The LPS-induced BV2 model was used to test the anti-neuroinflammatory effects and the regulatory effects and mechanisms on microglial M1/M2 polarization of psoralen in vitro. PC12 cell model induced by conditioned medium of BV2 cells was used to validate the protective effects of psoralen against neuroinflammation-induced neuronal damage. These results showed that psoralen inhibited the expression of iNOS, CD86, and TNF-α, and increased the expression of Arg-1, CD206, and IL-10. These results indicated that psoralen inhibited the M1 microglial phenotype and promoted the M2 microglial phenotype. Further studies showed that psoralen inhibited the phosphorylation of Fyn and PKCδ, thereby inhibiting activation of the MAPKs and NF-κB pathways and suppressing the expression of pro-inflammatory cytokines in microglia. Furthermore, psoralen reduced oxidative stress, neuronal damage, and apoptosis via inhibition of neuroinflammation. For the first time, this study showed that psoralen protected neurons and alleviated neuroinflammation by regulating microglial M1/M2 polarization, which may be mediated by inhibition of the Fyn-PKCδ pathway. Thus, psoralen may be a potential agent in the treatment of neuroinflammation-related diseases.
Asunto(s)
Ficusina , Lipopolisacáridos , Microglía , Enfermedades Neuroinflamatorias , Neuronas , Fármacos Neuroprotectores , Proteína Quinasa C-delta , Proteínas Proto-Oncogénicas c-fyn , Transducción de Señal , Animales , Microglía/efectos de los fármacos , Microglía/metabolismo , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteína Quinasa C-delta/metabolismo , Ratones , Ficusina/farmacología , Ficusina/uso terapéutico , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Células PC12 , Enfermedades Neuroinflamatorias/tratamiento farmacológico , Enfermedades Neuroinflamatorias/inmunología , Ratas , Transducción de Señal/efectos de los fármacos , Neuronas/efectos de los fármacos , Neuronas/patología , Masculino , Ratones Endogámicos C57BL , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Citocinas/metabolismoRESUMEN
Glioma, particularly glioblastomas (GBM), is incurable brain tumor. The most targeted receptor tyrosine kinase (RTKs) drugs did not bring benefit to GBM patients. The mechanism of glioma growth continues to be explored to find more effective treatment. Here, we reported that Ser/Thr protein kinase YANK2 (yet another kinase 2) is upregulated in glioma tissues and promotes the growth and proliferation of glioma in vitro and in vivo. Further, we confirmed that oncogene Fyn directly activated YANK2 through phosphorylation its Y110, and Fyn-mediated YANK2 phosphorylation at Y110 site promotes glioma growth by increasing its stability. Finally, YANK2 was proved to be a novel upstream kinase of p70S6K and promotes glioma growth by directly phosphorylating p70S6K at T389. Taken together, we found a new mTOR-independent p70S6K activation pathway, Fyn-YANK2-p70S6K, which promotes glioma growth, and YANK2 is a potential oncogene and serves as a novel therapeutic target for glioma.
Asunto(s)
Proliferación Celular , Glioma , Proteínas Proto-Oncogénicas c-fyn , Proteínas Quinasas S6 Ribosómicas 70-kDa , Transducción de Señal , Serina-Treonina Quinasas TOR , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Proteínas Quinasas S6 Ribosómicas 70-kDa/genética , Humanos , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas c-fyn/genética , Serina-Treonina Quinasas TOR/metabolismo , Glioma/metabolismo , Glioma/patología , Glioma/genética , Animales , Línea Celular Tumoral , Fosforilación , Carcinogénesis/genética , Carcinogénesis/metabolismo , Ratones , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/genética , Ratones Desnudos , Regulación Neoplásica de la Expresión GénicaRESUMEN
OBJECTIVES: This study aimed to explore the expression, clinical significance, and functional mechanism of FYN in lower-grade gliomas (LGG). METHODS: The mRNA and protein expression of FYN in LGG tissues were detected using databases including OncoLnc, GEPIA, and Human protein atlas (HPA). The UCSC Xena browser, TIMER, STRING and Metascape databases were used to investigate Kaplan-Meier survival curves, correlations between FYN expression and various types of immune cell infiltration, protein interaction network and possible functional mechanism. RESULTS: FYN expression in LGG, IDH mutation or 1p19q co-deletion subgroup was significantly higher than in corresponding control groups (p < 0.05). Patients with higher FYN expression had longer overall survival (p < 0.05). Male or no 1p19q co-deletion groups with higher FYN expression also had longer overall survival (p < 0.05). FYN expression had close correlation with infiltrating levels of cell purity, CD4+T cells, macrophages, and CD8+T cells (p < 0.05). Protein interaction network result showed correlation among FYN, SH2D1A, LCK, CAV1, SRC, CBL and PTK2. Functional enrichment analysis revealed that FYN and its related genes mainly participated in bacterial invasion of epithelial cells and natural killer cell mediated cytotoxicity. Peptidyl-tyrosine phosphorylation, negative regulation of anoikis, immune effector process, transmembrane receptor protein tyrosine kinase signaling pathway, epidermal growth factor receptor signaling pathway, and negative regulation of protein modification process may be the critical biological process. CONCLUSIONS: FYN is up-expressed in LGG and related to its good prognosis. It participated in tumor pathophysiological processes and may be a therapeutic target for LGG.
Asunto(s)
Biomarcadores de Tumor , Neoplasias Encefálicas , Glioma , Proteínas Proto-Oncogénicas c-fyn , Humanos , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas c-fyn/genética , Glioma/metabolismo , Glioma/genética , Pronóstico , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genética , Masculino , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/genética , Femenino , Mapas de Interacción de ProteínasRESUMEN
Nuclear receptors (NRs) are important transcriptional factors that mediate autophagy, preventing podocyte injury and the progression of diabetic kidney disease (DKD). However, the role of nuclear receptor coactivators that are powerful enhancers for the transcriptional activity of NRs in DKD remains unclear. In this study, a significant decrease in Nuclear Receptor Coactivator 3 (NCOA3) is observed in injured podocytes caused by high glucose treatment. Additionally, NCOA3 overexpression counteracts podocyte damage by improving autophagy. Further, Src family member, Fyn is identified to be the target of NCOA3 that mediates the podocyte autophagy process. Mechanistically, NCOA3 regulates the transcription of Fyn in a nuclear receptor, PPAR-γ dependent way. Podocyte-specific NCOA3 knockout aggravates albuminuria, glomerular sclerosis, podocyte injury, and autophagy in DKD mice. However, the Fyn inhibitor, AZD0530, rescues podocyte injury of NCOA3 knockout DKD mice. Renal NCOA3 overexpression with lentivirus can ameliorate podocyte damage and improve podocyte autophagy in DKD mice. Taken together, the findings highlight a novel target, NCOA3, that protects podocytes from high glucose injury by maintaining autophagy.
Asunto(s)
Autofagia , Nefropatías Diabéticas , Ratones Noqueados , Coactivador 3 de Receptor Nuclear , Podocitos , Animales , Masculino , Ratones , Autofagia/genética , Nefropatías Diabéticas/metabolismo , Nefropatías Diabéticas/genética , Nefropatías Diabéticas/patología , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Coactivador 3 de Receptor Nuclear/metabolismo , Coactivador 3 de Receptor Nuclear/genética , Podocitos/metabolismo , Podocitos/patología , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas c-fyn/genética , HumanosRESUMEN
OBJECTIVE: Hypoxic-ischemic brain injury in infants often leads to hemiplegic motor dysfunction. The mechanism of their motor dysfunction has been attributed to deficiencies of the transcription factor sex-determining region (SRY) box 2 (Sox2) or the non-receptor-type tyrosine kinase Fyn (involved in neuronal signal transduction), which causes a defect in myelin formation. Constraint-induced movement therapy (CIMT) following cerebral hypoxia-ischemia may stimulate myelin growth by regulating Sox2/Fyn, Ras homolog protein family A (RhoA), and rho-associated kinase 2 (ROCK2) expression levels. This study investigated how Sox2/Fyn regulates myelin remodeling following CIMT to improve motor function in rats with hemiplegic cerebral palsy (HCP). METHODS: To investigate the mechanism of Sox2 involvement in myelin growth and neural function in rats with HCP, Lentivirus (Lenti)-Sox2 adeno-associated virus and negative control-Lenti-Sox2 (LS) adeno-associated virus were injected into the lateral ventricle. The rats were divided into a control group and an HCP group with different interventions (CIMT, LS, or negative control-LS [NS] treatment), yielding the HCP, HCP plus CIMT (HCP + CIMT), HCP + LS, HCP + LS + CIMT, HCP + NS, and HCP + NS + CIMT groups. Front-limb suspension and RotaRod tests, Golgi-Cox staining, transmission electron microscopy, immunofluorescence staining, western blotting, and quantitative polymerase chain reaction experiments were used to analyze the motor function, dendrite/axon area, myelin ultrastructure, and levels of expression of oligodendrocytes and Sox2/Fyn/RhoA/ROCK2 in the motor cortex. RESULTS: The rats in the HCP + LS + CIMT group had better values for motor function, dendrite/axon area, myelin ultrastructure, oligodendrocytes, and Sox2/Fyn/RhoA/ROCK2 expression in the motor cortex than rats in the HCP and HCP + NS groups. The improvement of motor function and myelin remodeling, the expression of oligodendrocytes, and the expression of Sox2/Fyn/RhoA/ROCK2 in the HCP + LS group were similar to those in the HCP + CIMT group. CONCLUSION: CIMT might overcome RhoA/ROCK2 signaling by upregulating the transcription of Sox2 to Fyn in the brain to induce the maturation and differentiation of oligodendrocytes, thereby promoting myelin remodeling and improving motor function in rats with HCP. IMPACT: The pathway mediated by Sox2/Fyn could be a promising therapeutic target for HCP.
Asunto(s)
Parálisis Cerebral , Vaina de Mielina , Proteínas Proto-Oncogénicas c-fyn , Factores de Transcripción SOXB1 , Animales , Ratas , Vaina de Mielina/metabolismo , Factores de Transcripción SOXB1/metabolismo , Parálisis Cerebral/fisiopatología , Parálisis Cerebral/rehabilitación , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Hemiplejía/fisiopatología , Hemiplejía/rehabilitación , Masculino , Transducción de Señal/fisiología , Quinasas Asociadas a rho/metabolismo , Ratas Sprague-Dawley , Proteína de Unión al GTP rhoA/metabolismo , Modelos Animales de Enfermedad , Proteínas de Unión al GTP rhoRESUMEN
Fyn, Blk, and Lyn are part of a group of proteins called Src family kinases. They are crucial in controlling cell communication and their response to the growth, changes, and immune system. Blocking these proteins with inhibitors can be a way to treat diseases where these proteins are too active. The primary mode of action of these inhibitors is to inhibit the phosphorylation of Fyn, Blk, and Lyn receptors, which in turn affects how signals pass within the cells. This review shows the structural and functional aspects of Fyn, Blk, and Lyn kinases, highlighting the significance of their dysregulation in diseases such as cancer and autoimmune disorders. The discussion encompasses the design strategies, SAR analysis, and chemical characteristics of effective inhibitors, shedding light on their specificity and potency. Furthermore, it explores the progress of clinical trials of these inhibitors, emphasizing their potential therapeutic applications.
Asunto(s)
Proteínas Tirosina Quinasas , Proteínas Proto-Oncogénicas , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Familia-src Quinasas , FosforilaciónRESUMEN
Glutamatergic synapses encode information from extracellular inputs using dynamic protein interaction networks (PINs) that undergo widespread reorganization following synaptic activity, allowing cells to distinguish between signaling inputs and generate coordinated cellular responses. Here, we investigate how Fragile X Messenger Ribonucleoprotein (FMRP) deficiency disrupts signal transduction through a glutamatergic synapse PIN downstream of NMDA receptor or metabotropic glutamate receptor (mGluR) stimulation. In cultured cortical neurons or acute cortical slices from P7, P17 and P60 FMR1-/y mice, the unstimulated protein interaction network state resembled that of wildtype littermates stimulated with mGluR agonists, demonstrating resting state pre-activation of mGluR signaling networks. In contrast, interactions downstream of NMDAR stimulation were similar to WT. We identified the Src family kinase (SFK) Fyn as a network hub, because many interactions involving Fyn were pre-activated in FMR1-/y animals. We tested whether targeting SFKs in FMR1-/y mice could modify disease phenotypes, and found that Saracatinib (SCB), an SFK inhibitor, normalized elevated basal protein synthesis, novel object recognition memory and social behavior in FMR1-/y mice. However, SCB treatment did not normalize the PIN to a wild-type-like state in vitro or in vivo, but rather induced extensive changes to protein complexes containing Shank3, NMDARs and Fyn. We conclude that targeting abnormal nodes of a PIN can identify potential disease-modifying drugs, but behavioral rescue does not correlate with PIN normalization.
Asunto(s)
Benzodioxoles , Modelos Animales de Enfermedad , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Síndrome del Cromosoma X Frágil , Neuronas , Proteínas Proto-Oncogénicas c-fyn , Familia-src Quinasas , Animales , Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil/tratamiento farmacológico , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Ratones , Familia-src Quinasas/metabolismo , Benzodioxoles/farmacología , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Neuronas/metabolismo , Neuronas/efectos de los fármacos , Masculino , Receptores de Glutamato Metabotrópico/metabolismo , Transducción de Señal/efectos de los fármacos , Fenotipo , Sinapsis/metabolismo , Sinapsis/efectos de los fármacos , Mapas de Interacción de Proteínas/efectos de los fármacos , Receptores de N-Metil-D-Aspartato/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Corteza Cerebral/metabolismo , Corteza Cerebral/efectos de los fármacos , QuinazolinasRESUMEN
The tyrosine kinase Fyn is a member of the SRC family of kinases, and its sustained activation is closely linked to tumor cell migration, proliferation, and cell metabolism. Recently, Fyn has been found to be expressed in various tumor tissues, and the expression and function of Fyn vary between tumors, with Fyn acting as an oncogene to promote proliferation and metastasis in some tumors. This article summarizes the recent studies on the role of Fyn in different human tumors, focusing on the role of Fyn in melanoma, breast cancer, glioma, lung cancer, and peripheral T-cell lymphoma in order to provide a basis for future research and targeted therapy in different human tumors.(AU)
Asunto(s)
Humanos , Proteínas Proto-Oncogénicas c-fyn/genética , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Neoplasias/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Regulación Neoplásica de la Expresión Génica , FosforilaciónRESUMEN
The tyrosine kinase Fyn is a member of the SRC family of kinases, and its sustained activation is closely linked to tumor cell migration, proliferation, and cell metabolism. Recently, Fyn has been found to be expressed in various tumor tissues, and the expression and function of Fyn vary between tumors, with Fyn acting as an oncogene to promote proliferation and metastasis in some tumors. This article summarizes the recent studies on the role of Fyn in different human tumors, focusing on the role of Fyn in melanoma, breast cancer, glioma, lung cancer, and peripheral T-cell lymphoma in order to provide a basis for future research and targeted therapy in different human tumors.
Asunto(s)
Melanoma , Proteínas Tirosina Quinasas , Humanos , Fosforilación , Proteínas Proto-Oncogénicas c-fyn/genética , Proteínas Proto-Oncogénicas c-fyn/metabolismoRESUMEN
OBJECTIVES: Oxidative stress-mediated colistin's nephrotoxicity is associated with the diminished activity of nuclear factor erythroid 2-related factor 2 (Nrf2) that is primarily correlated with cellular PH domain and leucine-rich repeat protein phosphatase (PHLPP2) levels. This study investigated the possible modulation of PHLPP2/protein kinase B (Akt) trajectory as a critical regulator of Nrf2 stability by rosuvastatin (RST) to guard against colistin-induced oxidative renal damage in rats. METHODS: Colistin (300,000 IU/kg/day; i.p.) was injected for 6 consecutive days, and rats were treated simultaneously with RST orally at 10 or 20 mg/kg. KEY FINDINGS: RST enhanced renal nuclear Nrf2 translocation as revealed by immunohistochemical staining to boost the renal antioxidants, superoxide dismutase (SOD) and reduced glutathione (GSH) along with a marked reduction in caspase-3. Accordingly, rats treated with RST showed significant restoration of normal renal function and histological features. On the molecular level, RST effectively decreased the mRNA expression of PHLPP2 to promote Akt phosphorylation. Consequently, it deactivated GSK-3ß and reduced the gene expression of Fyn kinase in renal tissues. CONCLUSIONS: RST could attenuate colistin-induced oxidative acute kidney injury via its suppressive effect on PHLPP2 to endorse Nrf2 activity through modulating Akt/GSK3 ß/Fyn kinase trajectory.
Asunto(s)
Lesión Renal Aguda , Proteínas Proto-Oncogénicas c-akt , Ratas , Animales , Proteínas Proto-Oncogénicas c-akt/metabolismo , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Rosuvastatina Cálcica/farmacología , Colistina/metabolismo , Colistina/farmacología , Transducción de Señal , Glucógeno Sintasa Quinasa 3/metabolismo , Glucógeno Sintasa Quinasa 3/farmacología , Estrés Oxidativo , Antioxidantes/farmacología , Antioxidantes/metabolismo , Riñón , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/tratamiento farmacológico , Lesión Renal Aguda/prevención & control , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas c-fyn/farmacologíaRESUMEN
Tubulointerstitial fibrosis is a common pathological change in end-stage renal disease. However, limited treatment methods are developed, and unexplained potential mechanisms of renal diseases are urgent problems to be solved. In the present research, we first elucidated the role of podocarpusflavone (POD), a biflavone compound, in unilateral ureteral obstruction (UUO) in rodent model which is characterized by inflammation and fibrosis. The changes in histology and immunohistochemistry were observed that POD exerted renoprotective effects by retarding the infiltration of macrophage and aberrant deposition of É-SMA, Col1a1, and fibronectin. Consistent with in vivo assay, POD treatment also ameliorated the process of fibrosis in TGF-ß1-stimulated renal tubular epithelial cells and inflammation in LPS-induced RAW264.7 cells in vitro. In terms of mechanism, our results showed that treatment with POD inhibited the aggravated activation of Fyn in the UUO group, and weakened the level of phosphorylation of Stat3 which indicated that POD may alleviate the process of fibrosis by the Fyn/Stat3 signaling pathway. Furthermore, the gain of function assay by lentivirus-mediated exogenous forced expression of Fyn abrogated the therapeutic effect of the POD on renal fibrosis and inflammation. Collectively, it can be concluded that POD exerted a protective effect on renal fibrosis by mediating Fyn/Stat3 signaling pathway.
Asunto(s)
Enfermedades Renales , Obstrucción Ureteral , Ratones , Fibrosis , Inflamación/metabolismo , Riñón/metabolismo , Riñón/patología , Enfermedades Renales/tratamiento farmacológico , Enfermedades Renales/patología , Proteínas Proto-Oncogénicas c-fyn/efectos de los fármacos , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Células RAW 264.7/efectos de los fármacos , Células RAW 264.7/metabolismo , Transducción de Señal , Factor de Transcripción STAT3/metabolismo , Obstrucción Ureteral/complicaciones , Obstrucción Ureteral/tratamiento farmacológico , Obstrucción Ureteral/metabolismo , AnimalesRESUMEN
Src family protein kinases (SFKs) play a key role in cell adhesion, invasion, proliferation, survival, apoptosis, and angiogenesis during tumor development. In humans, SFKs consists of eight family members with similar structure and function. There is a high level of overexpression or hyperactivity of SFKs in tumor, and they play an important role in multiple signaling pathways involved in tumorigenesis. FYN is a member of the SFKs that regulate normal cellular processes. Additionally, FYN is highly expressed in many cancers and promotes cancer growth and metastasis through diverse biological functions such as cell growth, apoptosis, and motility migration, as well as the development of drug resistance in many tumors. Moreover, FYN is involved in the regulation of multiple cancer-related signaling pathways, including interactions with ERK, COX-2, STAT5, MET and AKT. FYN is therefore an attractive therapeutic target for various tumor types, and suppressing FYN can improve the prognosis and prolong the life of patients. The purpose of this review is to provide an overview of FYN's structure, expression, upstream regulators, downstream substrate molecules, and biological functions in tumors.
Asunto(s)
Neoplasias , Proteínas Proto-Oncogénicas c-fyn , Transducción de Señal , Humanos , Movimiento Celular , Neoplasias/genética , Neoplasias/terapia , Proteínas Proto-Oncogénicas c-fyn/genética , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Familia-src Quinasas/metabolismoRESUMEN
Although Src is one of the oldest and most investigated oncoproteins, its function in tumor malignancy remains to be defined further. In this study, we demonstrated that the inhibition of Src activity by ponatinib effectively suppressed several malignant phenotypes of esophageal squamous cell carcinoma (ESCC) both in vitro and in vivo, whereas it did not produce growth-inhibitory effects on normal esophageal epithelial cells (NEECs). Importantly, we combined phosphoproteomics and several cellular and molecular biologic strategies to identify that Src interacted with the members of Src-family kinases (SFKs), such as Fyn or Lyn, to form heterodimers. Src interactions with Fyn and Lyn phosphorylated the tyrosine sites in SH2 (Fyn Tyr185 or Lyn Tyr183) and kinase domains (Fyn Tyr420 or Lyn Tyr397), which critically contributed to ESCC development. By contrast, Src could not form heterodimers with Fyn or Lyn in NEECs. We used RNA sequencing to comprehensively demonstrate that the inhibition of Src activity effectively blocked several critical tumor-promoting pathways, such as JAK/STAT, mTOR, stemness-related, and metabolism-related pathways. Results of the real-time polymerase chain reaction (RT-PCR) assay confirmed that Lyn and Fyn were critical effectors for the Src-mediated expression of tumor growth or metastasis-related molecules. Furthermore, results of the clinical ESCC samples showed that the hyperactivation of pSrc Tyr419, Fyn Tyr185 or Tyr420, and Lyn Tyr183 or Tyr397 could be biomarkers of ESCC prognosis. This study illustrates that Src/Fyn and Src/Lyn heterodimers serve as targets for the treatment of ESCC.
Asunto(s)
Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Humanos , Proteínas Proto-Oncogénicas c-fyn/genética , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas , Carcinoma de Células Escamosas de Esófago/diagnóstico , Neoplasias Esofágicas/diagnóstico , Neoplasias Esofágicas/tratamiento farmacológico , Neoplasias Esofágicas/genética , Familia-src Quinasas/genética , Familia-src Quinasas/metabolismo , Tirosina/metabolismo , FosforilaciónRESUMEN
Fyn is a Src kinase that controls critical signalling cascades and has been implicated in learning and memory. Postsynaptic enrichment of Fyn underpins synaptotoxicity in dementias such as Alzheimer's disease and frontotemporal lobar degeneration with Tau pathology (FTLD-Tau). The FLTD P301L mutant Tau is associated with a higher propensity to undergo liquid-liquid phase separation (LLPS) and form biomolecular condensates. Expression of P301L mutant Tau promotes aberrant trapping of Fyn in nanoclusters within hippocampal dendrites by an unknown mechanism. Here, we used single-particle tracking photoactivated localisation microscopy to demonstrate that the opening of Fyn into its primed conformation promotes its nanoclustering in dendrites leading to increased Fyn/ERK/S6 downstream signalling. Preventing the auto-inhibitory closed conformation of Fyn through phospho-inhibition or through perturbation of its SH3 domain increased Fyn's nanoscale trapping, whereas inhibition of the catalytic domain had no impact. By combining pharmacological and genetic approaches, we demonstrate that P301L Tau enhanced both Fyn nanoclustering and Fyn/ERK/S6 signalling via its ability to form biomolecular condensates. Together, our findings demonstrate that Fyn alternates between a closed and an open conformation, the latter being enzymatically active and clustered. Furthermore, pathogenic immobilisation of Fyn relies on the ability of P301L Tau to form biomolecular condensates, thus highlighting the critical importance of LLPS in controlling nanoclustering and downstream intracellular signalling events.
Asunto(s)
Enfermedad de Alzheimer , Demencia Frontotemporal , Degeneración Lobar Frontotemporal , Humanos , Proteínas tau/genética , Proteínas tau/metabolismo , Condensados Biomoleculares , Proteínas Proto-Oncogénicas c-fyn/genética , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Enfermedad de Alzheimer/genética , Degeneración Lobar Frontotemporal/metabolismoRESUMEN
N-methyl-D-aspartate receptors (NMDARs) are important postsynaptic receptors that contribute to normal synaptic function and cell survival; however, when overactivated, as in Huntington's disease (HD), NMDARs cause excitotoxicity. HD-affected striatal neurons show altered NMDAR currents and augmented ratio of surface to internal GluN2B-containing NMDARs, with augmented accumulation at extrasynaptic sites. Fyn protein is a member of the Src kinase family (SKF) with an important role in NMDARs phosphorylation and synaptic localization and function; recently, we demonstrated that Fyn is reduced in several HD models. Thus, in this study, we aimed to explore the impact of HD-mediated altered Fyn levels at post-synaptic density (PSD), and their role in distorted NMDARs function and localization, and intracellular neuroprotective pathways in YAC128 mouse primary striatal neurons. We show that reduced synaptic Fyn levels and activity in HD mouse striatal neurons is related to decreased phosphorylation of synaptic GluN2B-composed NMDARs; this occurs concomitantly with augmented extrasynaptic NMDARs activity and currents and reduced cAMP response element-binding protein (CREB) activation, along with induction of cell death pathways. Importantly, expression of a constitutive active form of SKF reestablishes NMDARs localization, phosphorylation, and function at PSD in YAC128 mouse neurons. Enhanced SKF levels and activity also promotes CREB activation and reduces caspase-3 activation in YAC128 mouse striatal neurons. This work supports, for the first time, a relevant role for Fyn protein in PSD modulation, controlling NMDARs synaptic function in HD, and favoring neuroprotective pathways and cell survival. In this respect, Fyn Tyr kinase constitutes an important potential HD therapeutic target directly acting at PSD.
Asunto(s)
Enfermedad de Huntington , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Receptores de N-Metil-D-Aspartato , Animales , Caspasa 3/metabolismo , Cuerpo Estriado/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Enfermedad de Huntington/metabolismo , Ratones , Receptores de N-Metil-D-Aspartato/metabolismoRESUMEN
FYN is a non-receptor tyrosine kinase of the SRC family that facilitates virus entry across epithelial tight junctions. However, the role of FYN in mammalian testes in maintaining the blood-testis barrier (BTB) integrity and the adhesion of germ cells to Sertoli cells are not well defined. Here, we show that FYN is a component of the BTB and the apical ectoplasmic specialization (ES) at Sertoli-Sertoli and Sertoli-spermatid interfaces, respectively, and is expressed extensively in mouse testes during postnatal development. FYN was shown to be structurally linked to the actin and microtubule-based cytoskeletons. An in vivo model was used to explore the modulatory effect of FYN on BTB and apical ES dynamics within the testes when adult mice were treated intraperitoneally with CdCl2 (3 mg/kg body weight). The CdCl2-induced epithelial restructuring was associated with a transient increase in the interaction between FYN and the actin branching/nucleation protein Arp3, as well as an induction of Arp3 phosphorylation, which possibly lead to actin cytoskeleton remodeling, resulting in BTB damage and germ cell loss in the seminiferous epithelium. Based on the results, we propose a model in which FYN and Arp3 form a protein complex that is responsible for junction reorganization events at the apical ES and the BTB. It is also possible for viruses to break through the BTB and enter the immunoprivileged testicular microenvironment via this mechanism.
Asunto(s)
Barrera Hematotesticular , Testículo , Actinas/metabolismo , Animales , Barrera Hematotesticular/metabolismo , Adhesión Celular , Masculino , Mamíferos/metabolismo , Ratones , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Espermatogénesis , Testículo/metabolismoRESUMEN
Src family kinases (SFKs) have been implicated in the pathogenesis of kidney fibrosis. However, the specific mechanism by which SFKs contribute to the progression of diabetic kidney disease (DKD) remains unclear. Our preliminary transcriptome analysis suggested that SFK expression was increased in diabetic kidneys and that the expression of Fyn (a member of the SFKs), along with genes related to unfolded protein responses from the endoplasmic reticulum (ER) stress signaling pathway, was upregulated in the tubules of human diabetic kidneys. Thus, we examined whether SFK-induced ER stress is associated with DKD progression. Mouse proximal tubular (mProx24) cells were transfected with Fyn or Lyn siRNA and exposed to high glucose and palmitate (HG-Pal). Streptozotocin-induced diabetic rats were treated with KF-1607, a novel pan-Src kinase inhibitor (SKI) with low toxicity. The effect of KF-1607 was compared to that of losartan, a standard treatment for patients with DKD. Among the SFK family members, the Fyn and Lyn kinases were upregulated under diabetic stress. HG-Pal induced p70S6 kinase and JNK/CHOP signaling and promoted tubular injury. Fyn knockdown but not Lyn knockdown inhibited this detrimental signaling pathway. In addition, diabetic rats treated with KF-1607 showed improved kidney function and decreased ER stress, inflammation, and fibrosis compared with those treated with losartan. Collectively, these findings indicate that Fyn kinase is a specific member of the SFKs implicated in ER stress activation leading to proximal tubular injury in the diabetic milieu and that pan-SKI treatment attenuates kidney injury in diabetic rats. These data highlight Fyn kinase as a viable target for the development of therapeutic agents for DKD.
Asunto(s)
Diabetes Mellitus Experimental , Nefropatías Diabéticas , Animales , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/patología , Nefropatías Diabéticas/tratamiento farmacológico , Nefropatías Diabéticas/patología , Estrés del Retículo Endoplásmico , Fibrosis , Humanos , Riñón/patología , Losartán , Ratones , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-fyn/genética , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Ratas , Familia-src Quinasas/metabolismoRESUMEN
Protein phosphatase 2A (PP2A) is a major phospho-Ser/Thr phosphatase and a key regulator of cellular signal transduction pathways. While PP2A dysfunction has been linked to human cancer and neurodegenerative disorders such as Alzheimer's disease (AD), PP2A regulation remains relatively poorly understood. It has been reported that the PP2A catalytic subunit (PP2Ac) is inactivated by a single phosphorylation at the Tyr307 residue by tyrosine kinases such as v-Src. However, multiple mass spectrometry studies have revealed the existence of other putative PP2Ac phosphorylation sites in response to activation of Src and Fyn, two major Src family kinases (SFKs). Here, using PP2Ac phosphomutants and novel phosphosite-specific PP2Ac antibodies, we show that cellular pools of PP2Ac are instead phosphorylated on both Tyr127 and Tyr284 upon Src activation, and on Tyr284 following Fyn activation. We found these phosphorylation events enhanced the interaction of PP2Ac with SFKs. In addition, we reveal SFK-mediated phosphorylation of PP2Ac at Y284 promotes dissociation of the regulatory Bα subunit, altering PP2A substrate specificity; the phosphodeficient Y127/284F and Y284F PP2Ac mutants prevented SFK-mediated phosphorylation of Tau at the CP13 (pSer202) epitope, a pathological hallmark of AD, and SFK-dependent activation of ERK, a major growth regulatory kinase upregulated in many cancers. Our findings demonstrate a novel PP2A regulatory mechanism that challenges the existing dogma on the inhibition of PP2A catalytic activity by Tyr307 phosphorylation. We propose dysregulation of SFK signaling in cancer and AD can lead to alterations in PP2A phosphorylation and subsequent deregulation of key PP2A substrates, including ERK and Tau.
Asunto(s)
Proteína Fosfatasa 2 , Proteínas Proto-Oncogénicas c-fyn , Familia-src Quinasas , Enfermedad de Alzheimer/metabolismo , Humanos , Fosfoproteínas Fosfatasas , Fosforilación , Proteína Fosfatasa 2/metabolismo , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Proteínas Proto-Oncogénicas pp60(c-src)/metabolismo , Tirosina/metabolismo , Familia-src Quinasas/genética , Familia-src Quinasas/metabolismo , Proteínas tau/metabolismoRESUMEN
Current pathophysiological findings indicate that damage to the alveolar epithelium plays a decisive role in the development of idiopathic pulmonary fibrosis (IPF). The available pharmacological interventions (i.e., oral pirfenidone and nintedanib) only slow down progression of the disease, but do not offer a cure. In order to develop new drug candidates, the pathophysiology of IPF needs to be better understood on a molecular level. It has previously been reported that a loss of caveolin-1 (Cav-1) contributes to profibrotic processes by causing reduced alveolar barrier function and fibrosis-like alterations of the lung-parenchyma. Conversely, overexpression of caveolin-1 appears to counteract the development of fibrosis by inhibiting the inflammasome NLRP3 and the associated expression of interleukin-1ß. In this study, the interaction between Fyn-kinase and caveolin-1 in the alveolar epithelium of various bleomycin (BLM)/TGF-ß damage models using precision-cut lung slices (PCLS), wildtype (WT) and caveolin-1 knockout (KO) mice as well as the human NCI-H441 cell line, were investigated. In WT mouse lung tissues, strong signals for Fyn-kinase were detected in alveolar epithelial type I cells, whereas in caveolin-1 KO animals, expression shifted to alveolar epithelial type II cells. Caveolin-1 and Fyn-kinase were found to be co-localized in isolated lipid rafts of NCI-H441 cell membrane fractions. These findings were corroborated by co-immunoprecipitation studies in which a co-localization of Cav-1 and Fyn-kinase was detected in the cell membrane of the alveolar epithelium. After TGF-ß and BLM-induced damage to the alveolar epithelium both in PCLS and cell culture experiments, a decrease in caveolin-1 and Fyn-kinase was found. Furthermore, TEER (transepithelial electrical resistance) measurements indicated that TGF-ß and BLM have a damaging effect on cell-cell contacts and thus impair the barrier function in NCI-H441 cell monolayers. This effect was attenuated after co-incubation with the Fyn-kinase inhibitor, PP-2. Our data suggest an involvement of Fyn-kinase and caveolin-1 in TGF-ß/bleomycin-induced impairment of alveolar barrier function and thus a possible role in the early stages of pulmonary fibrosis. Fyn-kinase and/or its complex with caveolin-1 might, therefore, be novel therapeutic targets in IPF.