RESUMEN
BACKGROUND/AIMS: Deubiquitinating enzymes (DUBs) are crucially involved in controlling signal transductions, and reverse ubiquitination by removing the ubiquitin from protein substrates. The Hippo signaling has an important role in tissue growth, cell proliferation, differentiation, and apoptosis. Since disruption of the Hippo signaling is associated with a number of diseases, it is imperative to investigate the molecular mechanism of the Hippo signaling. METHODS: DUB screening was performed using the kidney of the mouse unilateral ureteric obstruction (UUO) model to identify the cellular mechanism of the DUB-regulated Hippo signaling. In addition, kidney cells were used to confirm cell proliferation and protein levels in the Hippo signaling pathway. Densitometric analysis was conducted to compare the expression level of proteins using Image J. RESULTS: We found that YOD1, also known as OTU1, is downregulated in the mouse UUO model. We also demonstrated that YOD1 binds to and deubiquitinates neural precursor cell expressed developmentally down-regulated protein 4 (NEDD4). Furthermore, we observed that YOD1 suppresses NEDD4-induced cell proliferation. CONCLUSION: YOD1 regulates the Hippo signaling pathway through NEDD4, and the K63-linked polyubiquitin chain of NEDD4 plays an important role. Also, our results indicate that YOD1 plays an important role in kidney diseases.
Asunto(s)
Ubiquitina-Proteína Ligasas Nedd4/metabolismo , Transducción de Señal , Tioléster Hidrolasas/metabolismo , Animales , Línea Celular , Proliferación Celular , Modelos Animales de Enfermedad , Vía de Señalización Hippo , Humanos , Ratones , Mutagénesis , Ubiquitina-Proteína Ligasas Nedd4/química , Ubiquitina-Proteína Ligasas Nedd4/genética , Unión Proteica , Proteínas Serina-Treonina Quinasas/metabolismo , Insuficiencia Renal Crónica/metabolismo , Insuficiencia Renal Crónica/patología , Tioléster Hidrolasas/química , Ubiquitina/genética , Ubiquitina/metabolismo , UbiquitinaciónRESUMEN
Premature ovarian failure (POF) is defined as loss of ovarian function in women less than 40 years of age. The causes of POF are diverse and include environmental factors. Di-2-ethylhexyl phthalate (DEHP) is one factor that may cause POF. The ubiquitin-proteasome system maintains intracellular balance by promoting or inhibiting protein degradation. To investigate the differential expressions of deubiquitinating enzyme (DUB) genes in patients with POF, we developed two in vitro POF models by treating A2780 or OVCAR5 with DEHP. Using these models, a multiplex RT-PCR system for DUB genes was applied to identify biomarkers by comparing expression patterns and DUB mRNA levels; multiplex RT-PCR results were validated by qRT-PCR and Western blotting analyses. Observed differential expression levels of several DUB genes including USP12, COPS5, ATXN3L, USP49, and USP34 in A2780 and OVCAR5 cells at the mRNA and protein levels suggest that they should be investigated as potential biomarkers of POF.
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Enzimas Desubicuitinizantes/genética , Dietilhexil Ftalato/farmacología , Regulación Neoplásica de la Expresión Génica , Neoplasias Ováricas/tratamiento farmacológico , Ovario/efectos de los fármacos , Insuficiencia Ovárica Primaria/tratamiento farmacológico , Adulto , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Neoplasias Ováricas/genética , Insuficiencia Ovárica Primaria/genética , Complejo de la Endopetidasa Proteasomal/metabolismo , ARN Mensajero/metabolismoRESUMEN
Colored radiative cooling (CRC) offers an attractive alternative for surface and space cooling, while preserving the aesthetics of an object. However, there has been no study on the CRC using phosphors in regard to vivid coloration, sophisticated performance investigation, retention of properties, functionality, and structural flexibility all at once. Thus, to manage the entire solar spectrum, a colored cooling structure comprising a near-infrared (NIR)-reflective bottom layer and a top colored layer with a phosphor-embedded polymer matrix is proposed. The structure is paintable, vividly colored, hydrophobic, and ultraviolet (UV) and water resistant. In the daytime outdoor measurement, the structure with red, orange, and yellow colors exhibited lower temperature than a control group using commercial white paint by 4.7 °C, 7.2 °C, and 7.4 °C, respectively. After precise theoretical and experimental time-tracing temperature validation, the CRC performance enhancement from NIR reflection and photoluminescence effects was thoroughly analyzed, and a temperature reduction of up to 16.1 °C was achieved for the orange-colored structure. Furthermore, experiments of hydrophobicity infusion and exposure to UV and deionized water verified the durability of the colored cooling structure. In addition, flexible-film-type colored cooling structures were demonstrated using different bottom reflective layers, such as a silver thin film and porous aluminum oxide particle-embedded poly(vinylidene fluoride-co-hexafluoropropylene), suggesting the potential applicability of these colored cooling structures for vivid-colored, functional, and durable CRC.
RESUMEN
Polymerase chain reaction (PCR)-based diagnostic kits for point-of-care (POC) testing are highly desirable to prevent the spread of infectious diseases. Here, we demonstrate a rapid PCR testing kit that involves integrating a lateral flow paper strip with a nichrome-based thin film heater. The use of a paper membrane as a PCR-solution container results in fast thermocycling without a cooler because the membrane can contain the solution with a high specific surface area where Joule heating is applied. After PCR, amplified products are simultaneously detected at the lateral flow paper strip with the naked eye. Severe acute respiratory syndrome ß-coronavirus RNA can be detected within 30 min after PCR solution injection. This work reveals that the paper membrane can act as not only a capillary flow channel but also as a promising platform for fast PCR and detection.
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COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , Reacción en Cadena de la Polimerasa/métodos , Prueba de COVID-19 , Pruebas en el Punto de AtenciónRESUMEN
The expression of High-temperature requirement factor A4 (HtrA4) mRNA is significantly lower in the chorionic villi of patients with recurrent pregnancy loss (RPL) than in the control group. We conducted an investigation into the cellular functions of HtrA4 using the CRISPR/Cas9 system and shRNA-HtrA4 to create knockout BeWo cells and HtrA4 knockdown JEG3 cells. Our results indicated that the knockout BeWo cells exhibited reduced capacity for invasion and fusion, but increased levels of proliferation and migration, with a significantly shortened cell cycle compared to wild-type cells. Wild-type BeWo cells highly expressed cell invasion- and fusion-related factors, while knockout BeWo cells highly expressed migration-, proliferation-, and cell cycle-related factors. The shRNA-HtrA4 JEG3 cells showed a decreased capacity for invasion, but an increased capacity for migration, accompanied by a decrease in the expression of cell invasion-related factors and an increase in migration-related factors. Moreover, our ELISA results revealed that the serum HtrA4 level was lower in patients with RPL than in the controls. These findings suggest that HtrA4 depletion may be associated with placental dysfunction.
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Placenta , Preeclampsia , Embarazo , Humanos , Femenino , Placenta/metabolismo , Temperatura , Línea Celular Tumoral , Serina Proteasas/metabolismo , Preeclampsia/metabolismoRESUMEN
Radiation proctitis is the collateral damage that occurs to healthy cells during radiation treatment of pelvic malignancies. Conservative treatment of radiation proctitis can mitigate inflammatory symptoms, but, to date, no therapeutic options are available for direct recovery of the damaged colonic epithelium. The present study assessed the ability of colon organoid-based regeneration to treat radiation proctitis. Radiation proctitis was induced in mice by irradiating their recta, followed by enema-based transplantation of mouse colon organoids. The transplanted colon organoids were found to successfully engraft onto the damaged rectal mucosa of the irradiated mice, reconstituting epithelial structure and integrity. Lgr5+ stem cells were shown to be pivotal to colon organoid mediated regeneration. Endoscopic examination showed the efficacy of localized transplantation of colon organoids with fibrin glue to irradiated sites. These findings provide useful insights into the use of colon organoid-based regenerative therapy for the treatment of radiation proctitis.
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Proctitis , Traumatismos por Radiación , Animales , Colon , Mucosa Intestinal , Ratones , Organoides , Proctitis/terapia , Traumatismos por Radiación/terapiaRESUMEN
The ubiquitin-proteasome system (UPS) is responsible for proteasomal degradation, regulating the half-life of the protein. Deubiquitinating enzymes (DUBs) are components of the UPS and inhibit degradation by removing ubiquitins from protein substrates. Herpesvirus-associated ubiquitin-specific protease (HAUSP) is one such deubiquitinating enzyme and has been closely associated with tumor development. In a previous study, we isolated putative HAUSP binding substrates by two-dimensional electrophoresis (2-DE) and identified them by matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF/MS) analysis. The analysis showed that pyruvate kinase isoenzyme M2 (PKM2) was likely to be one of the substrates for HAUSP. Further study revealed that PKM2 binds to HAUSP, confirming the interaction between these proteins, and that PKM2 possesses the putative HAUSP binding motif, E or P/AXXS. Therefore, we generated mutant forms of PKM2 S57A, S97A, and S346A, and found that S57A had less binding affinity. In a previous study, we demonstrated that PKM2 is regulated by the UPS, and that HAUSP- as a DUB-acted on PKM2, thus siRNA for HAUSP increases PKM2 ubiquitination. Our present study newly highlights the direct interaction between HAUSP and PKM2.
RESUMEN
Deubiquitinating enzymes (DUBs) play an important role in the ubiquitin-proteasome system (UPS) by eliminating ubiquitins from substrates and inhibiting proteasomal degradation. Protein phosphatase methylesterase 1 (PME-1) inactivates protein phosphatase 2A (PP2A) and enhances the ERK and Akt signaling pathways, which increase cell proliferation and malignant cell transformation. In this study, we demonstrate that USP36 regulates PME-1 through its deubiquitinating enzyme activity. USP36 increases PME-1 stability, and depletion of USP36 decreases the PME-1 expression level. Furthermore, we demonstrate that USP36 promotes the ERK and Akt signaling pathways. In summary, it is suggested that USP36 regulates PME-1 as a DUB and participates in the ERK and Akt signaling pathways.
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Hidrolasas de Éster Carboxílico/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Ubiquitina Tiolesterasa/metabolismo , Progresión de la Enfermedad , Estabilidad de Enzimas , Células HEK293 , Células HeLa , Humanos , Complejo de la Endopetidasa Proteasomal/metabolismo , Dominios Proteicos , Ubiquitina/metabolismo , Ubiquitina Tiolesterasa/químicaRESUMEN
We developed three types of dithieno[3,2-b;2',3'-d]thiophene (DTT)-based organic sensitizers for high-performance thin photoactive TiO2 films and investigated the simple but powerful molecular engineering of different types of bonding between the triarylamine electron donor and the conjugated DTT π-bridge by the introduction of single, double, and triple bonds. As a result, with only 1.3 µm transparent and 2.5-µm TiO2 scattering layers, the triple-bond sensitizer (T-DAHTDTT) shows the highest power conversion efficiency (η = 8.4%; VOC = 0.73 V, JSC = 15.4 mA·cm-2, and FF = 0.75) in an iodine electrolyte system under one solar illumination (AM 1.5, 1000 W·m-2), followed by the single-bond sensitizer (S-DAHTDTT) (η = 7.6%) and the double-bond sensitizer (D-DAHTDTT) (η = 6.4%). We suggest that the superior performance of T-DAHTDTT comes from enhanced intramolecular charge transfer (ICT) induced by the triple bond. Consequently, T-DAHTDTT exhibits the most active photoelectron injection and charge transport on a TiO2 film during operation, which leads to the highest photocurrent density among the systems studied. We analyzed these correlations mainly in terms of charge injection efficiency, level of photocharge storage, and charge-transport kinetics. This study suggests that the molecular engineering of a triple bond between the electron donor and the π-bridge of a sensitizer increases the performance of dye-sensitized solar cell (DSC) with a thin photoactive film by enhancing not only JSC through improved ICT but also VOC through the evenly distributed sensitizer surface coverage.