RESUMEN
Glioblastoma multiforme (GBM) is a malignant tumour with a poor prognosis. Therefore, potential treatment strategies and novel therapeutic targets have gained increased attention. Our data showed that the ethanol extract of Vanilla planifolia stem (VAS) significantly decreased the viability and the colony formation of GBM cells. Moreover, VAS induced the cleavage of MAP1LC3, a marker of autophagy. Further RNA-seq and bioinformatic analysis revealed 4248 differentially expressed genes (DEGs) between VAS-treated GBM cells and the control cells. Protein-protein interactions between DEGs with fold changes less than -3 and more than 5 were further analysed, and we found that 16 and 9 hub DEGs, respectively, were correlated with other DEGs. Further qPCR experiments confirmed that 14 hub DEGs was significantly downregulated and 9 hub DEGs was significantly upregulated. In addition, another significantly downregulated DEG, p21-activated kinase 6 (PAK6), was correlated with the overall survival of GBM patients. Further validation experiments confirmed that VAS significantly reduced the mRNA and protein expression of PAK6, which led to the abolition of cell viability and colony formation. These findings demonstrated that VAS reduced cell viability, suppressed colony formation and induced autophagy and revealed PAK6 and other DEGs as potential therapeutic targets for GBM treatment.
Asunto(s)
Autofagia , Regulación Neoplásica de la Expresión Génica , Glioblastoma , Extractos Vegetales , Quinasas p21 Activadas , Glioblastoma/metabolismo , Glioblastoma/patología , Glioblastoma/tratamiento farmacológico , Glioblastoma/genética , Humanos , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Extractos Vegetales/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Línea Celular Tumoral , Autofagia/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Tallos de la Planta/química , Etanol , Proliferación Celular/efectos de los fármacos , Mapas de Interacción de Proteínas/efectos de los fármacos , Muerte Celular/efectos de los fármacosRESUMEN
PAK4 and PD-L1 have been suggested as novel therapeutic targets in human cancers. Moreover, PAK4 has been suggested to be a molecule closely related to the immune evasion of cancers. Therefore, this study evaluated the roles of PAK4 and PD-L1 in the progression of osteosarcomas in 32 osteosarcomas and osteosarcoma cells. In human osteosarcomas, immunohistochemical positivity for the expression of PAK4 (overall survival, p = 0.028) and PD-L1 (relapse-free survival, p = 0.002) were independent indicators for the survival of patients in a multivariate analysis. In osteosarcoma cells, the overexpression of PAK4 increased proliferation and invasiveness, while the knockdown of PAK4 suppressed proliferation and invasiveness. The expression of PAK4 was associated with the expression of the molecules related to cell cycle regulation, invasion, and apoptosis. PAK4 was involved in resistance to apoptosis under a treatment regime with doxorubicin for osteosarcoma. In U2OS cells, PAK4 was involved in the stabilization of PD-L1 from ubiquitin-mediated proteasomal degradation and the in vivo infiltration of immune cells such as regulatory T cells and PD1-, CD4-, and CD8-positive cells in mice tumors. In conclusion, this study suggests that PAK4 is involved in the progression of osteosarcoma by promoting proliferation, invasion, and resistance to doxorubicin and stabilized PD-L1 from proteasomal degradation.
Asunto(s)
Antígeno B7-H1 , Proliferación Celular , Doxorrubicina , Resistencia a Antineoplásicos , Osteosarcoma , Quinasas p21 Activadas , Osteosarcoma/patología , Osteosarcoma/tratamiento farmacológico , Osteosarcoma/metabolismo , Osteosarcoma/genética , Humanos , Antígeno B7-H1/metabolismo , Femenino , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Doxorrubicina/farmacología , Doxorrubicina/uso terapéutico , Animales , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Masculino , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Ratones , Apoptosis/efectos de los fármacos , Neoplasias Óseas/metabolismo , Neoplasias Óseas/patología , Neoplasias Óseas/tratamiento farmacológico , Neoplasias Óseas/genética , Adulto , Adolescente , Estabilidad Proteica/efectos de los fármacos , Ratones Desnudos , Adulto Joven , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Invasividad NeoplásicaRESUMEN
Our recent studies have identified p21-activated kinase 4 (PAK4) as a key regulator of lipid catabolism in the liver and adipose tissue, but its role in glucose homeostasis in skeletal muscle remains to be explored. In this study, we find that PAK4 levels are highly upregulated in the skeletal muscles of diabetic humans and mice. Skeletal muscle-specific Pak4 ablation or administering the PAK4 inhibitor in diet-induced obese mice retains insulin sensitivity, accompanied by AMPK activation and GLUT4 upregulation. We demonstrate that PAK4 promotes insulin resistance by phosphorylating AMPKα2 at Ser491, thereby inhibiting AMPK activity. We additionally show that skeletal muscle-specific expression of a phospho-mimetic mutant AMPKα2S491D impairs glucose tolerance, while the phospho-inactive mutant AMPKα2S491A improves it. In summary, our findings suggest that targeting skeletal muscle PAK4 may offer a therapeutic avenue for type 2 diabetes.
Asunto(s)
Proteínas Quinasas Activadas por AMP , Diabetes Mellitus Tipo 2 , Glucosa , Resistencia a la Insulina , Músculo Esquelético , Quinasas p21 Activadas , Animales , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Glucosa/metabolismo , Fosforilación , Músculo Esquelético/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Humanos , Ratones , Diabetes Mellitus Tipo 2/metabolismo , Masculino , Transportador de Glucosa de Tipo 4/metabolismo , Transportador de Glucosa de Tipo 4/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/metabolismoRESUMEN
A growing number of patients presenting severe combined immunodeficiencies attributed to monoallelic RAC2 variants have been identified. The expression of the RHO GTPase RAC2 is restricted to the hematopoietic lineage. RAC2 variants have been described to cause immunodeficiencies associated with high frequency of infection, leukopenia, and autoinflammatory features. Here, we show that specific RAC2 activating mutations induce the NLRP3 inflammasome activation leading to the secretion of IL-1ß and IL-18 from macrophages. This activation depends on the activation state of the RAC2 variant and is mediated by the downstream kinase PAK1. Inhibiting the RAC2-PAK1-NLRP3 inflammasome pathway might be considered as a potential treatment for these patients.
Asunto(s)
Mutación con Ganancia de Función , Inflamasomas , Interleucina-1beta , Macrófagos , Proteína con Dominio Pirina 3 de la Familia NLR , Proteína RCA2 de Unión a GTP , Quinasas p21 Activadas , Proteínas de Unión al GTP rac , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Inflamasomas/metabolismo , Inflamasomas/inmunología , Humanos , Macrófagos/inmunología , Macrófagos/metabolismo , Proteínas de Unión al GTP rac/genética , Proteínas de Unión al GTP rac/metabolismo , Interleucina-1beta/metabolismo , Interleucina-1beta/genética , Animales , Quinasas p21 Activadas/genética , Quinasas p21 Activadas/metabolismo , Ratones , Interleucina-18/genética , Interleucina-18/metabolismo , Transducción de SeñalRESUMEN
The GTPase Cdc42 regulates polarized growth in most eukaryotes. In the bipolar yeast Schizosaccharomyces pombe, Cdc42 activation cycles periodically at sites of polarized growth. These periodic cycles are caused by alternating positive feedback and time-delayed negative feedback loops. At each polarized end, negative feedback is established when active Cdc42 recruits the Pak1 kinase to prevent further Cdc42 activation. It is unclear how Cdc42 activation returns to each end after Pak1-dependent negative feedback. We find that disrupting branched actin-mediated endocytosis disables Cdc42 reactivation at the cell ends. Using experimental and mathematical approaches, we show that endocytosis-dependent Pak1 removal from the cell ends allows the Cdc42 activator Scd1 to return to that end to enable reactivation of Cdc42. Moreover, we show that Pak1 elicits its own removal via activation of endocytosis. These findings provide a deeper insight into the self-organization of Cdc42 regulation and reveal previously unknown feedback with endocytosis in the establishment of cell polarity.
Asunto(s)
Complejo 2-3 Proteico Relacionado con la Actina , Polaridad Celular , Endocitosis , Retroalimentación Fisiológica , Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Proteína de Unión al GTP cdc42 , Quinasas p21 Activadas , Schizosaccharomyces/metabolismo , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Complejo 2-3 Proteico Relacionado con la Actina/metabolismo , Complejo 2-3 Proteico Relacionado con la Actina/genética , Proteína de Unión al GTP cdc42/metabolismo , Proteína de Unión al GTP cdc42/genética , Actinas/metabolismoRESUMEN
Psychiatric disorders are highly inheritable, and most psychiatric disorders exhibit genetic overlap. Recent studies associated the 3q29 recurrent deletion with schizophrenia (SCZ) and autism spectrum disorder (ASD). In this study, we investigated the association of genes in the 3q29 region with SCZ and ASD. TM4SF19 and PAK2 were chosen as candidate genes for this study based on evidence from previous research. We sequenced TM4SF19 and PAK2 in 437 SCZ cases, 187 ASD cases and 524 controls in the Japanese population. Through targeted sequencing, we identified 6 missense variants among the cases (ASD & SCZ), 3 missense variants among controls, and 1 variant common to both cases and controls; however, no loss-of-function variants were identified. Fisher's exact test showed a significant association of variants in TM4SF19 among cases (p=0.0160). These results suggest TM4SF19 variants affect the etiology of SCZ and ASD in the Japanese population. Further research examining 3q29 region genes and their association with SCZ and ASD is thus needed.
Asunto(s)
Pueblo Asiatico , Trastorno del Espectro Autista , Predisposición Genética a la Enfermedad , Esquizofrenia , Humanos , Trastorno del Espectro Autista/genética , Esquizofrenia/genética , Femenino , Masculino , Japón , Pueblo Asiatico/genética , Predisposición Genética a la Enfermedad/genética , Quinasas p21 Activadas/genética , Cromosomas Humanos Par 3/genética , Adulto , Mutación Missense/genética , Estudios de Casos y Controles , Estudios de Asociación Genética , Pueblos del Este de AsiaRESUMEN
Background: Lenvatinib is the most common multitarget receptor tyrosine kinase inhibitor for the treatment of advanced hepatocellular carcinoma (HCC). Acquired resistance to lenvatinib is one of the major factors leading to the failure of HCC treatment, but the underlying mechanism has not been fully characterized. Methods: We established lenvatinib-resistant cell lines, cell-derived xenografts (CDXs) and patient-derived xenografts (PDXs) and obtained lenvatinib-resistant HCC tumor tissues for further study. Results: We found that ubiquitin-specific protease 14 (USP14) was significantly increased in lenvatinib-resistant HCC cells and tumors. Silencing USP14 significantly attenuated lenvatinib resistance in vitro and in vivo. Mechanistically, USP14 directly interacts with and stabilizes calcium- and integrin-binding protein 1 (CIB1) by reversing K48-linked proteolytic ubiquitination at K24, thus facilitating the P21-activated kinase 1 (PAK1)-ERK1/2 signaling axis. Moreover, in vivo adeno-associated virus 9 mediated transduction of CIB1 promoted lenvatinib resistance in PDXs, whereas CIB1 knockdown resensitized the response of PDXs to lenvatinib. Conclusions: These findings provide new insights into the role of CIB1/PAK1-ERK1/2 signaling in lenvatinib resistance in HCC. Targeting CIB1 and its pathways may be a novel pharmaceutical intervention for the treatment of lenvatinib-resistant HCC.
Asunto(s)
Carcinoma Hepatocelular , Resistencia a Antineoplásicos , Neoplasias Hepáticas , Compuestos de Fenilurea , Quinolinas , Ubiquitina Tiolesterasa , Quinasas p21 Activadas , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/tratamiento farmacológico , Humanos , Quinolinas/farmacología , Quinolinas/uso terapéutico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/tratamiento farmacológico , Ubiquitina Tiolesterasa/metabolismo , Ubiquitina Tiolesterasa/genética , Compuestos de Fenilurea/farmacología , Compuestos de Fenilurea/uso terapéutico , Animales , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Ratones , Línea Celular Tumoral , Sistema de Señalización de MAP Quinasas , Ratones Desnudos , UbiquitinaciónRESUMEN
BACKGROUND: An increasing number of studies have demonstrated the association of circular RNAs (circRNAs) with the pathological processes of various diseases and their involvement in the onset and progression of multiple cancers. Nevertheless, the functional roles and underlying mechanisms of circRNAs in the autophagy regulation of gastric cancer (GC) have not been fully elucidated. METHODS: We used transmission electron microscopy and the mRFP-GFP-LC3 dual fluorescent autophagy indicator to investigate autophagy regulation. The cell counting kit-8 assay, colony formation assay, 5-ethynyl-2'-deoxyuridine incorporation assay, Transwell assay, and Western blot assay were conducted to confirm circPTPN22's influence on GC progression. Dual luciferase reporter assays validated the binding between circPTPN22 and miR-6788-5p, as well as miR-6788-5p and p21-activated kinase-1 (PAK1). Functional rescue experiments assessed whether circPTPN22 modulates PAK1 expression by competitively binding miR-6788-5p, affecting autophagy and other biological processes in GC cells. We investigated the impact of circPTPN22 on in vivo GC tumors using a nude mouse xenograft model. Bioinformatics tools predicted upstream regulatory transcription factors and binding proteins of circPTPN22, while chromatin immunoprecipitation and ribonucleoprotein immunoprecipitation assays confirmed the binding status. RESULTS: Upregulation of circPTPN22 in GC has been shown to inhibit autophagy and promote cell proliferation, migration, and invasion. Mechanistically, circPTPN22 directly binds to miR-6788-5p, subsequently regulating the expression of PAK1, which activates protein kinase B (Akt) and extracellular signal-regulated kinase (Erk) phosphorylation. This modulation ultimately affects autophagy levels in GC cells. Additionally, runt-related transcription factor 1 (RUNX1) negatively regulates circPTPN22 expression, while RNA-binding proteins such as FUS (fused in sarcoma) and ELAVL1 (recombinant ELAV-like protein 1) positively regulate its expression. Inhibition of the autophagy pathway can increase FUS expression, further upregulating circPTPN22 in GC cells, thereby exacerbating the progression of GC. CONCLUSION: Under the regulation of the transcription factor RUNX1 and RNA-binding proteins FUS and ELAVL1, circPTPN22 activates the phosphorylation of Akt and Erk through the miR-6788-5p/PAK1 axis, thereby modulating autophagy in GC cells. Inhibition of autophagy increases FUS, which in turn upregulates circPTPN22, forming a positive feedback loop that ultimately accelerates the progression of GC.
Asunto(s)
Autofagia , Movimiento Celular , Proliferación Celular , Subunidad alfa 2 del Factor de Unión al Sitio Principal , Proteína 1 Similar a ELAV , MicroARNs , ARN Circular , Proteína FUS de Unión a ARN , Neoplasias Gástricas , Quinasas p21 Activadas , Humanos , Neoplasias Gástricas/genética , Neoplasias Gástricas/patología , Neoplasias Gástricas/metabolismo , ARN Circular/genética , ARN Circular/metabolismo , Autofagia/genética , MicroARNs/genética , MicroARNs/metabolismo , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Proliferación Celular/genética , Proteína FUS de Unión a ARN/metabolismo , Proteína FUS de Unión a ARN/genética , Movimiento Celular/genética , Línea Celular Tumoral , Animales , Proteína 1 Similar a ELAV/metabolismo , Proteína 1 Similar a ELAV/genética , Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Regulación Neoplásica de la Expresión Génica , Ratones Desnudos , Ratones , Invasividad Neoplásica , Ratones Endogámicos BALB CRESUMEN
Neurofibromatosis type 2 is a genetic disorder that results in the formation and progressive growth of schwannomas, ependymomas, and/or meningiomas. The NF2 gene encodes the Merlin protein, which links cell cortical elements to the actin cytoskeleton and regulates a number of key enzymes including Group I p21-activated kinases (PAKs), the Hippo-pathway kinase LATS, and mTORC. While PAK1 and PAK2 directly bind Merlin and transmit proliferation and survival signals when Merlin is mutated or absent, inhibition of Group 1 PAKs alone has not proven sufficient to completely stop the growth of NF2-deficient meningiomas or schwannomas in vivo, suggesting the need for a second pathway inhibitor. As the Hippo pathway is also activated in NF2-deficient cells, several inhibitors of the Hippo pathway have recently been developed in the form of YAP-TEAD binding inhibitors. These inhibitors prevent activation of pro-proliferation and anti-apoptotic Hippo pathway effectors. In this study, we show that PAK inhibition slows cell proliferation while TEAD inhibition promotes apoptotic cell death. Finally, we demonstrate the efficacy of PAK and TEAD inhibitor combinations in several NF2-deficient Schwannoma cell lines.
Asunto(s)
Proliferación Celular , Vía de Señalización Hippo , Neurilemoma , Neurofibromina 2 , Proteínas Serina-Treonina Quinasas , Transducción de Señal , Factores de Transcripción , Quinasas p21 Activadas , Humanos , Neurilemoma/metabolismo , Neurilemoma/genética , Neurilemoma/patología , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/antagonistas & inhibidores , Quinasas p21 Activadas/genética , Proliferación Celular/efectos de los fármacos , Neurofibromina 2/genética , Neurofibromina 2/deficiencia , Neurofibromina 2/metabolismo , Transducción de Señal/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Línea Celular Tumoral , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Apoptosis/efectos de los fármacos , Sinergismo Farmacológico , Neurofibromatosis 2/metabolismo , Neurofibromatosis 2/genética , Proteínas Señalizadoras YAP/metabolismo , Proteínas Señalizadoras YAP/genética , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ADN/genéticaRESUMEN
Acral lentiginous melanoma (ALM) is the most common subtype of acral melanoma. Even though recent genetic studies are reported in acral melanomas, the genetic differences between in-situ and invasive ALM remain unclear. We aimed to analyze specific genetic changes in ALM and compare genetic differences between in-situ and invasive lesions to identify genetic changes associated with the pathogenesis and progression of ALM. We performed whole genome sequencing of 71 tissue samples from 29 patients with ALM. Comparative analyses were performed, pairing in-situ ALMs with normal tissues and, furthermore, invasive ALMs with normal and in-situ tissues. Among 21 patients with in-situ ALMs, 3 patients (14.3%) had SMIM14 , SLC9B1 , FRG1 , FAM205A , ESRRA , and ESPN mutations, and copy number (CN) gains were identified in only 2 patients (9.5%). Comparing 13 invasive ALMs with in-situ tissues, CN gains were identified in GAB2 in 8 patients (61.5%), PAK1 in 6 patients (46.2%), and UCP2 and CCND1 in 5 patients (38.5%). Structural variants were frequent in in-situ and invasive ALM lesions. Both in-situ and invasive ALMs had very low frequencies of common driver mutations. Structural variants were common in both in-situ and invasive ALMs. Invasive ALMs had markedly increased CN gains, such as GAB2 , PAK1 , UCP2 , and CCND1 , compared with in-situ lesions. These results suggest that they are associated with melanoma invasion.
Asunto(s)
Biomarcadores de Tumor , Ciclina D1 , Variaciones en el Número de Copia de ADN , Melanoma , Invasividad Neoplásica , Neoplasias Cutáneas , Secuenciación Completa del Genoma , Quinasas p21 Activadas , Humanos , Melanoma/genética , Melanoma/patología , Masculino , Femenino , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología , Quinasas p21 Activadas/genética , Persona de Mediana Edad , Anciano , Ciclina D1/genética , Biomarcadores de Tumor/genética , Adulto , Proteínas Adaptadoras Transductoras de Señales/genética , Anciano de 80 o más Años , Predisposición Genética a la Enfermedad , Mutación , Fenotipo , Proteína Desacopladora 2RESUMEN
This study aimed to explore the expression, function, and mechanisms of TBC1D10B in colon cancer, as well as its potential applications in the diagnosis and treatment of the disease.The expression levels of TBC1D10B in colon cancer were assessed by analyzing the TCGA and CCLE databases. Immunohistochemistry analysis was conducted using tumor and adjacent non-tumor tissues from 68 colon cancer patients. Lentiviral infection techniques were employed to silence and overexpress TBC1D10B in colon cancer cells. The effects on cell proliferation, migration, and invasion were evaluated using CCK-8, EDU, wound healing, and Transwell invasion assays. Additionally, GSEA enrichment analysis was used to explore the association of TBC1D10B with biological pathways related to colon cancer. TBC1D10B was significantly upregulated in colon cancer and closely associated with patient prognosis. Silencing of TBC1D10B notably inhibited proliferation, migration, and invasion of colon cancer cells and promoted apoptosis. Conversely, overexpression of TBC1D10B enhanced these cellular functions. GSEA analysis revealed that TBC1D10B is enriched in the AKT/PI3K/mTOR signaling pathway and highly correlated with PAK4. The high expression of TBC1D10B in colon cancer is associated with poor prognosis. It influences cancer progression by regulating the proliferation, migration, and invasion capabilities of colon cancer cells, potentially acting through the AKT/PI3K/mTOR signaling pathway. These findings provide new targets and therapeutic strategies for the treatment of colon cancer.
Asunto(s)
Apoptosis , Movimiento Celular , Proliferación Celular , Neoplasias del Colon , Proteínas Activadoras de GTPasa , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt , Transducción de Señal , Serina-Treonina Quinasas TOR , Quinasas p21 Activadas , Femenino , Humanos , Masculino , Persona de Mediana Edad , Apoptosis/genética , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Neoplasias del Colon/genética , Neoplasias del Colon/patología , Neoplasias del Colon/metabolismo , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica , Proteínas Activadoras de GTPasa/metabolismo , Proteínas Activadoras de GTPasa/genética , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Pronóstico , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Serina-Treonina Quinasas TOR/metabolismo , Serina-Treonina Quinasas TOR/genéticaRESUMEN
By taking advantage of forward genetic analysis in mice, we have demonstrated that Pak1 plays a crucial role during DMBA/TPA skin carcinogenesis. Although Pak1 has been considered to promote cancer development, its overall function remains poorly understood. To clarify the functional significance of Pak1 in detail, we sought to evaluate the possible effect of an allosteric inhibitor against PAK1 (NVS-PAK1-1) on a syngeneic mouse model. To this end, we established two cell lines, 9AS1 and 19AS1, derived from DMBA/TPA-induced squamous cell carcinoma (SCC) that engrafted in FVB mice. Based on our present results, NVS-PAK1-1 treatment significantly inhibited the growth of tumors derived from 9AS1 and 19AS1 cells in vitro and in vivo. RNA-sequencing analysis on the engrafted tumors indicates that NVS-PAK1-1 markedly potentiates the epidermal cell differentiation and enhances the immune response in the engrafted tumors. Consistent with these observations, we found an expansion of Pan-keratin-positive regions and potentially elevated infiltration of CD8-positive immune cells in NVS-PAK1-1-treated tumors as examined by immunohistochemical analyses. Together, our present findings strongly suggest that PAK1 is tightly linked to the development of SCC, and that its inhibition is a promising therapeutic strategy against SCC.
Asunto(s)
Carcinoma de Células Escamosas , Modelos Animales de Enfermedad , Neoplasias Cutáneas , Quinasas p21 Activadas , Animales , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Quinasas p21 Activadas/antagonistas & inhibidores , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/patología , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/genética , Ratones , Línea Celular Tumoral , 9,10-Dimetil-1,2-benzantraceno/toxicidad , Femenino , Diferenciación Celular/efectos de los fármacos , Acetato de Tetradecanoilforbol , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/metabolismo , Proliferación Celular/efectos de los fármacosRESUMEN
The p21-activated kinase (PAK) family of proteins regulates various processes requiring dynamic cytoskeleton organization such as cell adhesion, migration, proliferation, and apoptosis. Among the six members of the protein family, PAK2 is specifically involved in apoptosis, angiogenesis, or the development of endothelial cells. We report a novel de novo heterozygous missense PAK2 variant, p.(Thr406Met), found in a newborn with clinical manifestations of Knobloch syndrome. In vitro experiments indicated that this and another reported variant, p.(Asp425Asn), result in substantially impaired protein kinase activity. Similar findings were described previously for the PAK2 p.(Glu435Lys) variant found in two siblings with proposed Knobloch syndrome type 2 (KNO2). These new variants support the association of PAK2 kinase deficiency with a second, autosomal dominant form of Knobloch syndrome: KNO2.
Asunto(s)
Quinasas p21 Activadas , Humanos , Quinasas p21 Activadas/genética , Desprendimiento de Retina/genética , Desprendimiento de Retina/patología , Desprendimiento de Retina/congénito , Degeneración Retiniana/genética , Degeneración Retiniana/patología , Masculino , Recién Nacido , Femenino , Mutación Missense/genética , EncefaloceleAsunto(s)
Carcinoma de Células Renales , Neoplasias Renales , Terapia Molecular Dirigida , Carcinoma de Células Renales/tratamiento farmacológico , Carcinoma de Células Renales/metabolismo , Carcinoma de Células Renales/patología , Humanos , Neoplasias Renales/tratamiento farmacológico , Neoplasias Renales/metabolismo , Neoplasias Renales/patología , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Quinasas p21 Activadas/antagonistas & inhibidores , Inmunoterapia/métodos , Quimera Dirigida a la ProteólisisRESUMEN
BACKGROUND: Erectile dysfunction (ED) is a common male sexual dysfunction, with an increasing incidence, and the current treatment is often ineffective. METHODS: Vascular endothelial growth factor (VEGFA) was used to treat bone marrow-derived mesenchymal stem cells (BM-MSCs), and their cell migration rates were determined by Transwell assays. The expression of the von Willebrand Factor (vWF)VE-cadherin, and endothelial nitric oxide synthase(eNOS) endothelial markers was determined by qRTâPCR and Western blot analyses. The MALAT1-induced differentiation of BM-MCs to ECs via the CDC42/PAK1/paxillin pathway was explored by transfecting VEGFA-induced BM-MSC with si-MALAT1 and overexpressing CDC42 and PAK1. The binding capacity between CDC42, PAK1, and paxillin in VEGFA-treated and non-VEGFA-treated BM-MSCs was examined by protein immunoprecipitation. MiR-206 was overexpressed in VEGFA-induced BM-MSC, and the binding sites of MALAT1, miR-206, and CDC42 were identified using a luciferase assay. Sixty male SpragueâDawley rats were divided into six groups (n = 10/group). DMED modelling was demonstrated by APO experiments and was assessed by measuring blood glucose levels. Erectile function was assessed by measuring the intracavernosa pressure (ICP) and mean arterial pressure (MAP). Penile erectile tissue was analysed by qRTâPCR, Western blot analysis, and immunohistochemical staining. RESULTS: MALAT1 under VEGFA treatment conditions regulates the differentiation of BM-MSCs into ECs by modulating the CDC42/PAK1/paxillin axis. In vitro experiments demonstrated that interference with CDC42 and MALAT1 expression inhibited the differentiation of BM-MSCs to ECs. CDC42 binds to PAK1, and PAK1 binds to paxillin. In addition, CDC42 in the VEGFA group had a greater ability to bind to PAK1, whereas PAK1 in the VEGFA group had a greater ability to bind to paxillin. Overexpression of miR-206 in VEGFA-induced BM-MSCs demonstrated that MALAT1 competes with the CDC42 3'-UTR for binding to miR-206, which in turn is involved in the differentiation of BM-MSCs to ECs. Compared to the DMED model group, the ICP/MAP ratio was significantly greater in the three BM-MSCs treatment groups. CONCLUSIONS: MALAT1 facilitates BM-MSC differentiation into ECs by regulating the miR-206/CDC42/PAK1/paxillin axis to improve ED. The present findings revealed the vital role of MALAT1 in the repair of BM-MSCs for erectile function and provided new mechanistic insights into the BM-MSC-mediated repair of DMED.
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Diferenciación Celular , Disfunción Eréctil , Células Madre Mesenquimatosas , MicroARNs , Paxillin , ARN Largo no Codificante , Ratas Sprague-Dawley , Transducción de Señal , Proteína de Unión al GTP cdc42 , Quinasas p21 Activadas , Masculino , Animales , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Diferenciación Celular/genética , Proteína de Unión al GTP cdc42/metabolismo , Proteína de Unión al GTP cdc42/genética , Ratas , Quinasas p21 Activadas/genética , Quinasas p21 Activadas/metabolismo , Células Madre Mesenquimatosas/metabolismo , Disfunción Eréctil/terapia , Disfunción Eréctil/genética , Disfunción Eréctil/metabolismo , Paxillin/metabolismo , Paxillin/genética , Células Endoteliales/metabolismo , Células Cultivadas , Factor A de Crecimiento Endotelial Vascular/metabolismo , Factor A de Crecimiento Endotelial Vascular/genéticaRESUMEN
BACKGROUND: The intermediate filament protein vimentin is widely recognized as a molecular marker of epithelial-to-mesenchymal transition. Although vimentin expression is strongly associated with cancer metastatic potential, the exact role of vimentin in cancer metastasis and the underlying mechanism of its pro-metastatic functions remain unclear. RESULTS: This study revealed that vimentin can enhance integrin ß1 surface expression and induce integrin-dependent clustering of cells, shielding them against anoikis cell death. The increased integrin ß1 surface expression in suspended cells was caused by vimentin-mediated protection of the internal integrin ß1 pool against lysosomal degradation. Additionally, cell detachment was found to induce vimentin Ser38 phosphorylation, allowing the translocation of internal integrin ß1 to the plasma membrane. Furthermore, the use of an inhibitor of p21-activated kinase PAK1, one of the kinases responsible for vimentin Ser38 phosphorylation, significantly reduced cancer metastasis in animal models. CONCLUSIONS: These findings suggest that vimentin can act as an integrin buffer, storing internalized integrin ß1 and releasing it when needed. Overall, this study provides insights regarding the strong correlation between vimentin expression and cancer metastasis and a basis for blocking metastasis using this novel therapeutic mechanism.
Asunto(s)
Anoicis , Integrina beta1 , Vimentina , Vimentina/metabolismo , Vimentina/genética , Integrina beta1/metabolismo , Integrina beta1/genética , Humanos , Animales , Supervivencia Celular , Ratones , Línea Celular Tumoral , Fosforilación , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genéticaRESUMEN
The intricate orchestration of osteoporosis (OP) pathogenesis remains elusive. Mounting evidence suggests that angiogenesis-driven osteogenesis serves as a crucial foundation for maintaining bone homeostasis. This study aimed to explore the potential of the endothelial platelet-derived growth factor receptor-ß (PDGFR-ß) in mitigating bone loss through its facilitation of H-type vessel formation. Our findings demonstrate that the expression level of endothelial PDGFR-ß is reduced in samples obtained from individuals suffering from OP, as well as in ovariectomy mice. Depletion of PDGFR-ß in endothelial cells ameliorates angiogenesis-mediated bone formation in mice. The regulatory influence of endothelial PDGFR-ß on H-type vessels is mediated through the PDGFRß-P21-activated kinase 1-Notch1 intracellular domain signaling cascade. In particular, the endothelium-specific enhancement of PDGFR-ß facilitates H-type vessels and their associated bone formation in OP. Hence, the strategic targeting of endothelial PDGFR-ß emerges as a promising therapeutic approach for the management of OP in the near future.
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Neovascularización Fisiológica , Osteogénesis , Osteoporosis , Receptor Notch1 , Receptor beta de Factor de Crecimiento Derivado de Plaquetas , Transducción de Señal , Quinasas p21 Activadas , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/genética , Animales , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/genética , Humanos , Femenino , Ratones , Receptor Notch1/metabolismo , Receptor Notch1/genética , Osteoporosis/metabolismo , Osteoporosis/patología , Células Endoteliales/metabolismo , Ratones Endogámicos C57BL , Células Endoteliales de la Vena Umbilical Humana/metabolismo , AngiogénesisRESUMEN
AIMS: Porocarcinoma is a malignant sweat gland tumour differentiated toward the upper part of the sweat duct and may arise from the transformation of a preexisting benign poroma. In 2019, Sekine et al. demonstrated the presence of YAP1::MAML2 and YAP1::NUTM1 fusions in most poromas and porocarcinomas. Recently, our group identified PAK2-fusions in a subset of benign poromas. Herein we report a series of 12 porocarcinoma cases harbouring PAK1/2/3 fusions. METHODS AND RESULTS: Five patients were male and the median age was 79 years (ranges: 59-95). Tumours were located on the trunk (n = 7), on the thigh (n = 3), neck (n = 1), or groin area (n = 1). Four patients developed distant metastases. Microscopically, seven cases harboured a benign poroma component and a malignant invasive part. Ductal formations were observed in all, while infundibular/horn cysts and cells with vacuolated cytoplasm were detected in seven and six tumours, respectively. In three cases, the invasive component consisted of a proliferation of elongated cells, some of which formed pseudovascular spaces, whereas the others harboured a predominant solid or trabecular growth pattern. Immunohistochemical staining for CEA and EMA confirmed the presence of ducts. Focal androgen receptor expression was detected in three specimens. Whole RNA sequencing evidenced LAMTOR1::PAK1 (n = 2), ZDHHC5::PAK1 (n = 2), DLG1::PAK2, CTDSP1::PAK1, CTNND1::PAK1, SSR1::PAK3, CTNNA1::PAK2, RNF13::PAK2, ROBO1::PAK2, and CD47::PAK2. Activating mutation of HRAS (G13V, n = 3, G13R, n = 1, Q61L, n = 2) was present in six cases. CONCLUSION: Our study suggests that PAK1/2/3 fusions is the oncogenic driver of a subset of porocarcinomas lacking YAP1 rearrangement.
Asunto(s)
Porocarcinoma Ecrino , Neoplasias de las Glándulas Sudoríparas , Quinasas p21 Activadas , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Masculino , Persona de Mediana Edad , Porocarcinoma Ecrino/patología , Porocarcinoma Ecrino/genética , Proteínas de Fusión Oncogénica/genética , Quinasas p21 Activadas/genética , Quinasas p21 Activadas/metabolismo , Neoplasias de las Glándulas Sudoríparas/patología , Neoplasias de las Glándulas Sudoríparas/genéticaRESUMEN
BACKGROUND: Ovarian cancer is the most common cause of gynecological cancer death. Pak4 has been proved to be tumorigenic in many types of cancers, but its role in ovarian cancer is still not clarified. METHODS: In this study, we used immunohistochemistry to investigate into Pak4 expression in different histological types of ovarian cancer. TIMER, TISCH2, GEPIA, ualcan, KM plotter, GSCA and GeneMANIA were used to identify the prognostic roles and gene regulation networks of Pak4 in ovarian cancer. Immune infiltration levels were investigated using TIMER database. RESULTS: Pak4 was highly expressed in ovarian cancers, regardless of different FIGO stages and histological grades. Single cell sequencing database proved that Pak4 was highly expressed in malignant ovarian cancer cells. Pak4 level was significantly correlated with different histological types of ovarian cancer. Pak4 expression was negatively connected with OS and PFS of ovarian cancer patients. Functions of Pak4 and its interacted genes were mainly involved in protein serine/threonine kinase activity, regulation of actin filament-based process and regulation of cytoskeleton organization. Pak4 level was negatively correlated with immune biomarkers of B cell infiltration (p = 2.39e-05), CD8 + T cell infiltration (p = 1.51e-04), neutrophil (p = 1.74e-06) and dendritic cell (p = 4.41e-08). Close correlation was found between Pak4 expression and T cell exhaustion (p < 0.05). CONCLUSIONS: Our results demonstrated the expression level, gene interaction networks and immune infiltration levels of Pak4 in ovarian cancer. And the results revealed role of Pak4 in tumorigenesis and the possibility to be a potential immunotherapeutic target.
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Linfocitos Infiltrantes de Tumor , Neoplasias Ováricas , Quinasas p21 Activadas , Femenino , Humanos , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genética , Carcinogénesis/genética , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes , Linfocitos Infiltrantes de Tumor/inmunología , Neoplasias Ováricas/genética , Neoplasias Ováricas/inmunología , Neoplasias Ováricas/patología , Quinasas p21 Activadas/genética , Quinasas p21 Activadas/inmunología , PronósticoRESUMEN
Rabies virus (RABV) is highly lethal and triggers severe neurological symptoms. The neuropathogenic mechanism remains poorly understood. Ras-related C3 botulinum toxin substrate 1 (Rac1) is a Rho-GTPase that is involved in actin remodeling and has been reported to be closely associated with neuronal dysfunction. In this study, by means of a combination of pharmacological inhibitors, small interfering RNA, and specific dominant-negatives, we characterize the crucial roles of dynamic actin and the regulatory function of Rac1 in RABV infection, dominantly in the viral entry phase. The data show that the RABV phosphoprotein interacts with Rac1. RABV phosphoprotein suppress Rac1 activity and impedes downstream Pak1-Limk1-Cofilin1 signaling, leading to the disruption of F-actin-based structure formation. In early viral infection, the EGFR-Rac1-signaling pathway undergoes a biphasic change, which is first upregulated and subsequently downregulated, corresponding to the RABV entry-induced remodeling pattern of F-actin. Taken together, our findings demonstrate for the first time the role played by the Rac1 signaling pathway in RABV infection and may provide a clue for an explanation for the etiology of rabies neurological pathogenesis.IMPORTANCEThough neuronal dysfunction is predominant in fatal rabies, the detailed mechanism by which rabies virus (RABV) infection causes neurological symptoms remains in question. The actin cytoskeleton is involved in numerous viruses infection and plays a crucial role in maintaining neurological function. The cytoskeletal disruption is closely associated with abnormal nervous symptoms and induces neurogenic diseases. In this study, we show that RABV infection led to the rearrangement of the cytoskeleton as well as the biphasic kinetics of the Rac1 signal transduction. These results help elucidate the mechanism that causes the aberrant neuronal processes by RABV infection and may shed light on therapeutic development aimed at ameliorating neurological disorders.