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Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of Kaposi sarcoma (KS), the plasmablastic form of multicentric Castleman's disease, and primary effusion lymphoma. In sub-Saharan Africa, KS is the most common HIV-related malignancy and one of the most common childhood cancers. Immunosuppressed patients, including HIV-infected patients, are more prone to KSHV-associated disease. KSHV encodes a viral protein kinase (vPK) that is expressed from ORF36. KSHV vPK contributes to the optimal production of infectious viral progeny and upregulation of protein synthesis. To elucidate the interactions of vPK with cellular proteins in KSHV-infected cells, we used a bottom-up proteomics approach and identified host protein ubiquitin-specific peptidase 9X-linked (USP9X) as a potential interactor of vPK. Subsequently, we validated this interaction using a co-immunoprecipitation assay. We report that both the ubiquitin-like and the catalytic domains of USP9X are important for association with vPK. To uncover the biological relevance of the USP9X/vPK interaction, we investigated whether the knockdown of USP9X would modulate viral reactivation. Our data suggest that depletion of USP9X inhibits both viral reactivation and the production of infectious virions. Understanding how USP9X influences the reactivation of KSHV will provide insights into how cellular deubiquitinases regulate viral kinase activity and how viruses co-opt cellular deubiquitinases to propagate infection. Hence, characterizing the roles of USP9X and vPK during KSHV infection constitutes a first step toward identifying a potentially critical interaction that could be targeted by future therapeutics. IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of Kaposi sarcoma (KS), the plasmablastic form of multicentric Castleman's disease, and primary effusion lymphoma. In sub-Saharan Africa, KS is the most common HIV-related malignancy. KSHV encodes a viral protein kinase (vPK) that aids viral replication. To elucidate the interactions of vPK with cellular proteins in KSHV-infected cells, we used an affinity purification approach and identified host protein ubiquitin-specific peptidase 9X-linked (USP9X) as a potential interactor of vPK. Depletion of USP9X inhibits both viral reactivation and the production of infectious virions. Overall, our data suggest a proviral role for USP9X.
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Herpesvirus Humano 8 , Sarcoma de Kaposi , Ubiquitina Tiolesterasa , Niño , Humanos , Enzimas Desubicuitinizantes , Herpesvirus Humano 8/fisiología , Infecciones por VIH/complicaciones , Linfoma de Efusión Primaria , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Sarcoma de Kaposi/metabolismo , Sarcoma de Kaposi/patología , Sarcoma de Kaposi/virología , Ubiquitina Tiolesterasa/genética , Proteínas Virales/genéticaRESUMEN
The anaphase-promoting complex/cyclosome (APC/C) is an E3 ubiquitin ligase and critical regulator of cell cycle progression. Despite its vital role, it has remained challenging to globally map APC/C substrates. By combining orthogonal features of known substrates, we predicted APC/C substrates in silico. This analysis identified many known substrates and suggested numerous candidates. Unexpectedly, chromatin regulatory proteins are enriched among putative substrates, and we show experimentally that several chromatin proteins bind APC/C, oscillate during the cell cycle, and are degraded following APC/C activation, consistent with being direct APC/C substrates. Additional analysis revealed detailed mechanisms of ubiquitylation for UHRF1, a key chromatin regulator involved in histone ubiquitylation and DNA methylation maintenance. Disrupting UHRF1 degradation at mitotic exit accelerates G1-phase cell cycle progression and perturbs global DNA methylation patterning in the genome. We conclude that APC/C coordinates crosstalk between cell cycle and chromatin regulatory proteins. This has potential consequences in normal cell physiology, where the chromatin environment changes depending on proliferative state, as well as in disease.
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Ciclosoma-Complejo Promotor de la Anafase/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Cromatina/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ciclosoma-Complejo Promotor de la Anafase/fisiología , Proteínas Potenciadoras de Unión a CCAAT/genética , Proteínas Potenciadoras de Unión a CCAAT/fisiología , Ciclo Celular/fisiología , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Cromatina/genética , Simulación por Computador , Células HEK293 , Células HeLa , Humanos , Procesamiento Proteico-Postraduccional , Factores de Transcripción/metabolismo , Ubiquitina-Proteína Ligasas/fisiología , UbiquitinaciónRESUMEN
Nuclear factor erythroid 2-related factor 2 (NFE2L2, also known as NRF2) is a transcription factor and master regulator of cellular antioxidant response. Aberrantly high NRF2-dependent transcription is recurrent in human cancer, but conversely NRF2 activity diminishes with age and in neurodegenerative and metabolic disorders. Although NRF2-activating drugs are clinically beneficial, NRF2 inhibitors do not yet exist. Here, we describe use of a gain-of-function genetic screen of the kinome to identify new druggable regulators of NRF2 signaling. We found that the under-studied protein kinase brain-specific kinase 2 (BRSK2) and the related BRSK1 kinases suppress NRF2-dependent transcription and NRF2 protein levels in an activity-dependent manner. Integrated phosphoproteomics and RNAseq studies revealed that BRSK2 drives 5'-AMP-activated protein kinase α2 (AMPK) signaling and suppresses the mTOR pathway. As a result, BRSK2 kinase activation suppresses ribosome-RNA complexes, global protein synthesis and NRF2 protein levels. Collectively, our data illuminate the BRSK2 and BRSK1 kinases, in part by functionally connecting them to NRF2 signaling and mTOR. This signaling axis might prove useful for therapeutically targeting NRF2 in human disease.This article has an associated First Person interview with the first author of the paper.
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Factor 2 Relacionado con NF-E2 , Receptor EphA5 , Proteínas Quinasas Activadas por AMP/metabolismo , Mutación con Ganancia de Función , Humanos , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal/genéticaRESUMEN
BACKGROUND: Human papillomavirus-positive (HPV+) squamous cell carcinoma of the oropharynx (OPSCC) is the most prevalent HPV-associated malignancy in the United States. Favorable treatment outcomes have led to increased interest in treatment de-escalation to reduce treatment morbidity as well as the development of prognostic markers to identify appropriately low-risk patients. Intratumoral genomic heterogeneity and copy number alteration burden have been demonstrated to be predictive of poor outcomes in many other cancers; therefore, we sought to determine whether intratumor heterogeneity and genomic instability are associated with poor outcomes in HPV+ OPSCC. METHODS: Tumor heterogeneity estimates were made based on targeted exome sequencing of 45 patients with HPV+ OPSCC tumors. Analysis of an additional cohort of HPV+ OPSCC tumors lacking matched normal sequencing allowed copy number analysis of 99 patient tumors. RESULTS: High intratumorally genomic heterogeneity and high numbers of copy number alterations were strongly associated with worse recurrence-free survival. Tumors with higher heterogeneity and frequent copy number alterations were associated with loss of distal 11q, which encodes key genes related to double-strand break repair, including ATM and MRE11A. CONCLUSIONS: Both intratumor genomic heterogeneity and high-burden copy number alterations are strongly associated with poor recurrence-free survival in patients with HPV+ OPSCC. The drivers of genomic instability and heterogeneity in these tumors remains to be elucidated. However, 11q loss and defective DNA double-strand break repair have been associated with genomic instability in other solid tumors. Copy number alteration burden and intratumoral heterogeneity represent promising avenues for risk stratification of patients with HPV+OPSCC.
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Alphapapillomavirus , Carcinoma de Células Escamosas , Neoplasias Orofaríngeas , Infecciones por Papillomavirus , Carcinoma de Células Escamosas/patología , Variaciones en el Número de Copia de ADN , Genómica , Humanos , Neoplasias Orofaríngeas/patología , Papillomaviridae/genética , Infecciones por Papillomavirus/complicaciones , Infecciones por Papillomavirus/genética , PronósticoRESUMEN
Leukocyte transendothelial migration is one of the most important step in launching an inflammatory immune response and chronic inflammation can lead to devastating diseases. Leukocyte migration inhibitors are considered as promising and potentially effective therapeutic agents to treat inflammatory and auto-immune disorders. In this study, based on previous trioxotetrahydropyrimidin based integrin inhibitors that suboptimally blocked leukocyte adhesion, twelve molecules with a modified scaffold were designed, synthesized, and tested in vitro for their capacity to block the transendothelial migration of immune cells. One of the molecules, namely, methyl 4-((2-(tert-butyl)-6-((2,4,6-trioxotetrahydropyrimidin-5(2H)-ylidene) methyl) phenoxy) methyl) benzoate, (compound 12), completely blocked leukocyte transendothelial migration, without any toxic effects on immune or endothelial cells (IC50â¯=â¯2.4⯵M). In vivo, compound 12 exhibited significant therapeutic effects in inflammatory bowel disease (IBD)/Crohn's disease, multiple sclerosis, fatty liver disease, and rheumatoid arthritis models. A detailed acute and chronic toxicity profile of the lead compound in vivo did not reveal any toxic effects. Such a type of molecule might therefore provide a unique starting point for designing a novel class of leukocyte transmigration blocking agents with broad therapeutic applications in inflammatory and auto-immune pathologies.
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Linfocitos B/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Monocitos/efectos de los fármacos , Pirimidinas/síntesis química , Linfocitos T/efectos de los fármacos , Migración Transcelular de la Célula/efectos de los fármacos , Migración Transendotelial y Transepitelial/efectos de los fármacos , Linfocitos B/inmunología , Adhesión Celular/efectos de los fármacos , Moléculas de Adhesión Celular/inmunología , Células Endoteliales de la Vena Umbilical Humana/inmunología , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Inflamación , Estructura Molecular , Monocitos/inmunología , Mucoproteínas/inmunología , Pirimidinas/química , Pirimidinas/farmacología , Linfocitos T/inmunología , Molécula 1 de Adhesión Celular Vascular/inmunologíaRESUMEN
BACKGROUND: The study objective was to obtain consensus on physical therapy (PT) in the rehabilitation of critical illness survivors after hospital discharge. Research questions were: what are PT goals, what are recommended measurement tools, and what constitutes an optimal PT intervention for survivors of critical illness? METHODS: A Delphi consensus study was conducted. Panelists were included based on relevant fields of expertise, years of clinical experience, and publication record. A literature review determined five themes, forming the basis for Delphi round one, which was aimed at generating ideas. Statements were drafted and ranked on a 5-point Likert scale in two additional rounds with the objective to reach consensus. Results were expressed as median and semi-interquartile range, with the consensus threshold set at ≤0.5. RESULTS: Ten internationally established researchers and clinicians participated in this Delphi panel, with a response rate of 80 %, 100 %, and 100 % across three rounds. Consensus was reached on 88.5 % of the statements, resulting in a framework for PT after hospital discharge. Essential handover information should include information on 15 parameters. A core set of outcomes should test exercise capacity, skeletal muscle strength, function in activities of daily living, mobility, quality of life, and pain. PT interventions should include functional exercises, circuit and endurance training, strengthening exercises for limb and respiratory muscles, education on recovery, and a nutritional component. Screening tools to identify impairments in other health domains and referral to specialists are proposed. CONCLUSIONS: A consensus-based framework for optimal PT after hospital discharge is proposed. Future research should focus on feasibility testing of this framework, developing risk stratification tools and validating core outcome measures for ICU survivors.
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Consenso , Enfermedad Crítica/rehabilitación , Modalidades de Fisioterapia/normas , Rehabilitación/métodos , Actividades Cotidianas , Técnica Delphi , Humanos , Alta del Paciente/tendencias , Rehabilitación/normas , SobrevivientesRESUMEN
The theory of heterodyne/stroboscopic detection of nuclear resonance scattering is developed, starting from the total scattering matrix as a product of the matrix of the reference sample and the sample under study. This general approach holds for all dynamical scattering channels. In the forward channel, which has been discussed in detail in the literature, the electronic scattering manifests itself only in an energy-independent diminution of the scattered intensity. In all other channels, complex resonance line shapes of the heterodyne/stroboscopic spectra are encountered, as a result of the interference of electronic and nuclear scattering. The grazing-incidence case will be evaluated and described in detail. Experimental data of classical X-ray reflectivity and their stroboscopically detected resonant counterpart spectra on the [(nat)Fe/(57)Fe]10 isotope periodic multilayer and antiferromagnetic [(57)Fe/Cr]20 superlattice are fitted simultaneously.
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UNLABELLED: Kaposi's sarcoma-associated herpesvirus (KSHV, also called human herpesvirus 8) is linked to the development of Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD). KSHV expresses several proteins that modulate host cell signaling pathways. One of these proteins is viral interleukin-6 (vIL-6), which is a homolog of human IL-6 (hIL-6). vIL-6 is able to prevent apoptosis and promote proinflammatory signaling, angiogenesis, and cell proliferation. Although it can be secreted, vIL-6 is mainly an intracellular protein that is retained in the endoplasmic reticulum (ER). We performed affinity purification and mass spectrometry to identify novel vIL-6 binding partners and found that a cellular ER chaperone, hypoxia-upregulated protein 1 (HYOU1), interacts with vIL-6. Immunohistochemical staining reveals that both PEL and KS tumor tissues express significant amounts of HYOU1. We also show that HYOU1 increases endogenous vIL-6 protein levels and that HYOU1 facilitates vIL-6-induced JAK/STAT signaling, migration, and survival in endothelial cells. Furthermore, our data suggest that HYOU1 also modulates vIL-6's ability to induce CCL2, a chemokine involved in cell migration. Finally, we investigated the impact of HYOU1 on cellular hIL-6 signaling. Collectively, our data indicate that HYOU1 is important for vIL-6 function and may play a role in the pathogenesis of KSHV-associated cancers. IMPORTANCE: KSHV vIL-6 is detectable in all KSHV-associated malignancies and promotes tumorigenesis and inflammation. We identified a cellular protein, called hypoxia-upregulated protein 1 (HYOU1), that interacts with KSHV vIL-6 and is present in KSHV-infected tumors. Our data suggest that HYOU1 facilitates the vIL-6-induced signaling, migration, and survival of endothelial cells.
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Proteínas HSP70 de Choque Térmico/metabolismo , Herpesvirus Humano 8/metabolismo , Interleucina-6/metabolismo , Proteínas Virales/metabolismo , Línea Celular , Quimiocina CCL2/metabolismo , Células HEK293 , Infecciones por Herpesviridae/metabolismo , Humanos , Quinasas Janus/metabolismo , Factores de Transcripción STAT/metabolismo , Sarcoma de Kaposi/metabolismo , Sarcoma de Kaposi/virología , Transducción de Señal/fisiología , Regulación hacia Arriba/fisiologíaRESUMEN
The last two decades have seen important advances in our knowledge of maize domestication, thanks in part to the contributions of genetic data. Genetic studies have provided firm evidence that maize was domesticated from Balsas teosinte (Zea mays subspecies parviglumis), a wild relative that is endemic to the mid- to lowland regions of southwestern Mexico. An interesting paradox remains, however: Maize cultivars that are most closely related to Balsas teosinte are found mainly in the Mexican highlands where subspecies parviglumis does not grow. Genetic data thus point to primary diffusion of domesticated maize from the highlands rather than from the region of initial domestication. Recent archeological evidence for early lowland cultivation has been consistent with the genetics of domestication, leaving the issue of the ancestral position of highland maize unresolved. We used a new SNP dataset scored in a large number of accessions of both teosinte and maize to take a second look at the geography of the earliest cultivated maize. We found that gene flow between maize and its wild relatives meaningfully impacts our inference of geographic origins. By analyzing differentiation from inferred ancestral gene frequencies, we obtained results that are fully consistent with current ecological, archeological, and genetic data concerning the geography of early maize cultivation.
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Demografía , Variación Genética , Genética de Población , Polimorfismo de Nucleótido Simple/genética , Zea mays/genética , Bases de Datos Genéticas , Frecuencia de los Genes , Flujo Genético , Genotipo , Geografía , México , Análisis de Componente Principal , Especificidad de la EspecieRESUMEN
PURPOSE: Multiple studies have explored the needs and experiences of patients, family members, and healthcare professionals regarding hospital-to-home transitions. Our study aimed to identify, critically appraise, and summarize these studies in a qualitative meta-synthesis. MATERIALS AND METHODS: Medline, CINAHL and Embase were systematically searched to identify eligible articles from inception to June 2024. Qualitative studies were included and critically appraised using the Critical Appraisal Skills Program. Insufficient-quality papers were excluded. We performed a meta-synthesis following (1) open coding by two independent researchers and (2) discussing codes during reflexivity meetings. RESULTS: Ninety-eight studies were appraised, of which 53 were included. We reached thematic saturation, four themes were constructed: (1) care coordination and continuity, (2) communication, (3) patient and family involvement, and (4) individualized support and information exchange. For patients and families, tailored information and support are prerequisites for a seamless transition and an optimal recovery trajectory after hospital discharge. It is imperative that healthcare professionals communicate effectively within and across care settings to ensure multidisciplinary collaboration and care continuity. CONCLUSIONS: This study identifies essential elements of optimal transitional care. These findings could be supportive to researchers and healthcare professionals when (re)designing transitional care interventions to ensure care continuity after hospital discharge.
Patients and their families need to receive tailored information and support, which are prerequisites for a seamless transition from hospital to homeProfessionals must communicate effectively within and across hospital and primary care settingsProfessional roles should be clarified to ensure effective collaboration and continued high-quality care after hospital discharge.Integrated allied health pathways addressing coordination and communication are needed to ensure seamless transitions.
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The nuclear factor erythroid 2-related factor 2 (NRF2) transcription factor activates cytoprotective and metabolic gene expression in response to various electrophilic stressors. Constitutive NRF2 activity promotes cancer progression, whereas decreased NRF2 function contributes to neurodegenerative diseases. We used proximity proteomic analysis to define protein networks for NRF2 and its family members NRF1, NRF3, and the NRF2 heterodimer MAFG. A functional screen of co-complexed proteins revealed previously uncharacterized regulators of NRF2 transcriptional activity. We found that ZNF746 (also known as PARIS), a zinc finger transcription factor implicated in Parkinson's disease, physically associated with NRF2 and MAFG, resulting in suppression of NRF2-driven transcription. ZNF746 overexpression increased oxidative stress and apoptosis in a neuronal cell model of Parkinson's disease, phenotypes that were reversed by chemical and genetic hyperactivation of NRF2. This study presents a functionally annotated proximity network for NRF2 and suggests a link between ZNF746 overexpression in Parkinson's disease and inhibition of NRF2-driven neuroprotection.
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Enfermedad de Parkinson , Humanos , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/metabolismo , Proteínas Represoras/metabolismo , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Proteínas Co-Represoras , ProteómicaRESUMEN
Acyl-protein thioesterases 1 and 2 (APT1 and APT2) reverse S-acylation, a potential regulator of systemic glucose metabolism in mammals. Palmitoylation proteomics in liver-specific knockout mice shows that APT1 predominates over APT2, primarily depalmitoylating mitochondrial proteins, including proteins linked to glutamine metabolism. miniTurbo-facilitated determination of the protein-protein proximity network of APT1 and APT2 in HepG2 cells reveals APT proximity networks encompassing mitochondrial proteins including the major translocases Tomm20 and Timm44. APT1 also interacts with Slc1a5 (ASCT2), the only glutamine transporter known to localize to mitochondria. High-fat-diet-fed male mice with dual (but not single) hepatic deletion of APT1 and APT2 have insulin resistance, fasting hyperglycemia, increased glutamine-driven gluconeogenesis, and decreased liver mass. These data suggest that APT1 and APT2 regulation of hepatic glucose metabolism and insulin signaling is functionally redundant. Identification of substrates and protein-protein proximity networks for APT1 and APT2 establishes a framework for defining mechanisms underlying metabolic disease.
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Proteoma , Tioléster Hidrolasas , Masculino , Ratones , Animales , Proteoma/metabolismo , Tioléster Hidrolasas/genética , Tioléster Hidrolasas/metabolismo , Glutamina/metabolismo , Mitocondrias/metabolismo , Hígado/metabolismo , Proteínas Mitocondriales/metabolismo , Glucosa/metabolismo , Lípidos , Mamíferos/metabolismoRESUMEN
Histone methyltransferases play essential roles in the organization and function of chromatin. They are also frequently mutated in human diseases including cancer1. One such often mutated methyltransferase, SETD2, associates co-transcriptionally with RNA polymerase II and catalyzes histone H3 lysine 36 trimethylation (H3K36me3) - a modification that contributes to gene transcription, splicing, and DNA repair2. While studies on SETD2 have largely focused on the consequences of its catalytic activity, the non-catalytic functions of SETD2 are largely unknown. Here we report a catalysis-independent function of SETD2 in maintaining nuclear lamina stability and genome integrity. We found that SETD2, via its intrinsically disordered N-terminus, associates with nuclear lamina proteins including lamin A/C, lamin B1, and emerin. Depletion of SETD2, or deletion of its N-terminus, resulted in widespread nuclear morphology abnormalities and genome stability defects that were reminiscent of a defective nuclear lamina. Mechanistically, the N-terminus of SETD2 facilitates the association of the mitotic kinase CDK1 with lamins, thereby promoting lamin phosphorylation and depolymerization required for nuclear envelope disassembly during mitosis. Taken together, our findings reveal an unanticipated link between the N-terminus of SETD2 and nuclear lamina organization that may underlie how SETD2 acts as a tumor suppressor.
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OBJECTIVE: NRF2 is a master transcription factor that regulates the stress response. NRF2 is frequently mutated and activated in human esophageal squamous cell carcinoma (ESCC), which drives resistance to chemotherapy and radiation therapy. Therefore, a great need exists for NRF2 inhibitors for targeted therapy of NRF2high ESCC. DESIGN: We performed high-throughput screening of two compound libraries from which hit compounds were further validated in human ESCC cells and a genetically modified mouse model. The mechanism of action of one compound was explored by biochemical assays. RESULTS: Using high-throughput screening of two small molecule compound libraries, we identified 11 hit compounds as potential NRF2 inhibitors with minimal cytotoxicity at specified concentrations. We then validated two of these compounds, pyrimethamine and mitoxantrone, by demonstrating their dose- and time-dependent inhibitory effects on the expression of NRF2 and its target genes in two NRF2Mut human ESCC cells (KYSE70 and KYSE180). RNAseq and qPCR confirmed the suppression of global NRF2 signaling by these two compounds. Mechanistically, pyrimethamine reduced NRF2 half-life by promoting NRF2 ubiquitination and degradation in KYSE70 and KYSE180 cells. Expression of an Nrf2E79Q allele in mouse esophageal epithelium (Sox2CreER;LSL-Nrf2E79Q/+) resulted in an NRF2high phenotype, which included squamous hyperplasia, hyperkeratinization, and hyperactive glycolysis. Treatment with pyrimethamine (30 mg/kg/day, p.o.) suppressed the NRF2high esophageal phenotype with no observed toxicity. CONCLUSION: We have identified and validated pyrimethamine as an NRF2 inhibitor that may be rapidly tested in the clinic for NRF2high ESCC.
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Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Humanos , Animales , Ratones , Carcinoma de Células Escamosas de Esófago/genética , Carcinoma de Células Escamosas de Esófago/patología , Carcinoma de Células Escamosas de Esófago/terapia , Neoplasias Esofágicas/genética , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Pirimetamina/farmacología , Pirimetamina/uso terapéutico , Hiperplasia , Línea Celular Tumoral , Proliferación CelularRESUMEN
This study aimed at understanding animal research technicians (ART) work activity to identify difficulties encountered by workers and their determinants which may increase musculoskeletal disorders (MSD) risks. The methods for the work activity analysis combined interviews, observations, events and operations chronicles as well as inclinometry. From the work activity analysis of the three main tasks (changing mouse cages, preparation of water bottles and unloading dirty material), difficulties such as awkward postures, heavy load handling, repetitiveness, high workload, supplementary tasks, interruptions and difficult social interactions emerged. The work activity analysis further allowed the identification of determinants of these difficulties. Some are related to the physical, organizational or social work environment, and others to the interdependence between these determinants. Such an improved understanding of ART work activity will lead to solutions best suited for MSDs prevention in this understudied setting.
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Técnicos de Animales , Enfermedades Musculoesqueléticas , Enfermedades Profesionales , Humanos , Enfermedades Musculoesqueléticas/etiología , Enfermedades Musculoesqueléticas/prevención & control , Enfermedades Profesionales/etiología , Enfermedades Profesionales/prevención & control , Postura , Factores de Riesgo , Carga de TrabajoRESUMEN
A new high radial resolution 2D multichannel Charge eXchange Imaging (CXI) diagnostic is under development for deployment at DIII-D. The diagnostic system will measure low-to-intermediate radial wavenumber carbon density fluctuations by observing the n = 8 - 7 (λ = 529.06 nm) C-VI emission line, resulting from charge exchange collisions between heating neutral beam atoms and the intrinsic carbon ion density. The new CXI diagnostic will provide measurements with ΔR â¼ 0.4 cm to access higher kr instabilities (kr < 8 cm-1) predicted to arise in the steep-gradient region of the H-mode pedestal. The CXI system will feature 60 fiber bundles in a 12 × 5 arrangement, with each bundle consisting of four 1 mm fibers. A custom optical system has been designed to filter and image incoming signals onto an 8 × 8 avalanche photodiode array. Additionally, a novel electronics suite has been designed and commissioned to amplify and digitize the relatively low-intensity carbon signal at a 2 MHz bandwidth. Forward modeling results of the active C-VI emission suggest sufficient signal to noise ratios to resolve turbulent fluctuations. Prototype measurements demonstrate the ability to perform high frequency pedestal measurements.
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Studies have shown that Nrf2E79Q/+ is one of the most common mutations found in human tumors. To elucidate how this genetic change contributes to lung cancer, we compared lung tumor development in a genetically-engineered mouse model (GEMM) with dual Trp53/p16 loss, the most common mutations found in human lung tumors, in the presence or absence of Nrf2E79Q/+. Trp53/p16-deficient mice developed combined-small cell lung cancer (C-SCLC), a mixture of pure-SCLC (P-SCLC) and large cell neuroendocrine carcinoma. Mice possessing the LSL-Nrf2E79Q mutation showed no difference in the incidence or latency of C-SCLC compared with Nrf2+/+ mice. However, these tumors did not express NRF2 despite Cre-induced recombination of the LSL-Nrf2E79Q allele. Trp53/p16-deficient mice also developed P-SCLC, where activation of the NRF2E79Q mutation associated with a higher incidence of this tumor type. All C-SCLCs and P-SCLCs were positive for NE-markers, NKX1-2 (a lung cancer marker) and negative for P63 (a squamous cell marker), while only P-SCLC expressed NRF2 by immunohistochemistry. Analysis of a consensus NRF2 pathway signature in human NE+-lung tumors showed variable activation of NRF2 signaling. Our study characterizes the first GEMM that develops C-SCLC, a poorly-studied human cancer and implicates a role for NRF2 activation in SCLC development.
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Carcinoma Neuroendocrino , Neoplasias Pulmonares , Carcinoma Pulmonar de Células Pequeñas , Animales , Carcinoma Neuroendocrino/patología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Proteínas de Homeodominio/metabolismo , Humanos , Incidencia , Neoplasias Pulmonares/patología , Ratones , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Proteínas Nucleares/metabolismo , Carcinoma Pulmonar de Células Pequeñas/patología , Factores de Transcripción/metabolismo , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Neuronal morphogenesis involves dramatic plasma membrane expansion, fueled by soluble N-ethylmaleimide-sensitive factor attachment protein eceptors (SNARE)-mediated exocytosis. Distinct fusion modes described at synapses include full-vesicle fusion (FVF) and kiss-and-run fusion (KNR). During FVF, lumenal cargo is secreted and vesicle membrane incorporates into the plasma membrane. During KNR, a transient fusion pore secretes cargo but closes without membrane addition. In contrast, fusion modes are not described in developing neurons. Here, we resolve individual exocytic events in developing murine cortical neurons and use classification tools to identify four distinguishable fusion modes: two FVF-like modes that insert membrane material and two KNR-like modes that do not. Discrete fluorescence profiles suggest distinct behavior of the fusion pore. Simulations and experiments agree that FVF-like exocytosis provides sufficient membrane material for morphogenesis. We find the E3 ubiquitin ligase TRIM67 promotes FVF-like exocytosis in part by limiting incorporation of the Qb/Qc SNARE SNAP47 into SNARE complexes and, thus, SNAP47 involvement in exocytosis.
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Proteínas del Citoesqueleto/metabolismo , Exocitosis , Neurogénesis , Proteínas Qb-SNARE/metabolismo , Proteínas Qc-SNARE/metabolismo , Sinapsis/metabolismo , Proteínas de Motivos Tripartitos/metabolismo , Animales , Proteínas del Citoesqueleto/genética , Femenino , Ratones , Ratones Noqueados , Proteínas Qb-SNARE/genética , Proteínas Qc-SNARE/genética , Proteínas SNARE/genética , Proteínas SNARE/metabolismo , Sinapsis/genética , Proteínas de Motivos Tripartitos/genéticaRESUMEN
OBJECTIVES: Patients with laryngeal squamous cell carcinoma (LSCC) often fail radiation therapy (RT), when received as monotherapy or in combination with other treatment modalities. Mechanisms for RT failure are poorly understood. We hypothesized that tumors failing RT would have increased rates of somatic mutations in genes associated with radiation resistance, particularly in genes associated with the NFE2L2 oxidative stress pathway. Using targeted exome sequencing on pretreated LSCC tumors, we retrospectively compared somatic mutation profile with clinical data and response to treatment. METHODS: Tumors were classified as either radiation-resistant (RR) or radiation-sensitive (RS). RR was defined as persistent or recurrent disease within 2 years of receiving full-dose RT. Early stage (ES) LSCC was defined as Stage I or II tumors without lymph node involvement. Eight genes associated with radiation resistance were prioritized for analysis. RT-qPCR was performed on five NFE2L2 pathway genes. RESULTS: Twenty LSCC tumors were included and classified as either RR (n = 8) or RS (n = 12). No differences in individual rates of somatic mutations by genes associated with radiation resistance were identified. Higher rates of total mutational burden (TMB) and increased alterations associated with the NFE2L2 pathway was observed in RR vs RS tumors (P < .05). In an analysis of only ES-LSCC patients (RR, n = 3 and RS, n = 3), RR tumors had increased NFE2L2 somatic pathway mutations (P = .014) and increased NQO1 mRNA expression (P = .05). CONCLUSION: Increased TMB and NFE2L2 pathway alterations were associated with radiation resistance in LSCC. NQO1 mRNA expression may serve as a biomarker for RT response in ES-LSCC.Level of Evidence: II1.
RESUMEN
TRIM9 and TRIM67 are neuronally enriched E3 ubiquitin ligases essential for appropriate morphogenesis of cortical and hippocampal neurons and fidelitous responses to the axon guidance cue netrin-1. Deletion of murine Trim9 or Trim67 results in neuroanatomical defects and striking behavioral deficits, particularly in spatial learning and memory. TRIM9 and TRIM67 interact with cytoskeletal and exocytic proteins, but the full interactome is not known. Here we performed the unbiased proximity-dependent biotin identification (BioID) approach to define TRIM9 and TRIM67 protein-protein proximity network in developing cortical neurons and identified putative neuronal TRIM interaction partners. Candidates included cytoskeletal regulators, cytosolic protein transporters, exocytosis and endocytosis regulators, and proteins necessary for synaptic regulation. A subset of high-priority candidates was validated, including Myo16, Coro1A, MAP1B, ExoC1, GRIP1, PRG-1, and KIF1A. For a subset of validated candidates, we utilized total internal reflection fluorescence microscopy to demonstrate dynamic colocalization with TRIM proteins at the axonal periphery, including at the tips of filopodia. Further analysis demonstrated that the RNA interference-based knockdown of the unconventional myosin Myo16 in cortical neurons altered growth cone filopodia density and axonal branching patterns in a TRIM9- and netrin-1-dependent manner. Future analysis of other validated candidates will likely identify novel proteins and mechanisms by which TRIM9 and TRIM67 regulate neuronal form and function. [Media: see text].