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The pogo transposable element derived zinc finger protein, POGZ, is notably associated with neurodevelopmental disorders through its role in gene transcription. Many proteins involved in neurological development are often dysregulated in cancer, suggesting a potential role for POGZ in tumor biology. Here, we provided experimental evidence that POGZ influences the growth and metastatic spread of triple negative breast cancers (TNBC). In well-characterized models of TNBC, POGZ exerted a dual role, both as a tumor promoter and metastasis suppressor. Mechanistically, loss of POGZ potentiated TGFß pathway activation to exert cytostatic effects while simultaneously increasing the mesenchymal and migratory properties of breast tumors. Whereas POGZ levels are elevated in human breast cancers, the most aggressive forms of TNBC tumors, including those with increased mesenchymal and metastatic properties, exhibit dampened POGZ levels, and low POGZ expression was associated with inferior clinical outcomes in these tumor types. Taken together, these data suggest that POGZ is a critical suppressor of the early stages of the metastatic cascade.
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DNA double-strand breaks (DSBs) elicit an elaborate response to signal damage and trigger repair via two major pathways: nonhomologous end-joining (NHEJ), which functions throughout the interphase, and homologous recombination (HR), restricted to S/G2 phases. The DNA damage response relies, on post-translational modifications of nuclear factors to coordinate the mending of breaks. Ubiquitylation of histones and chromatin-associated factors regulates DSB repair and numerous E3 ubiquitin ligases are involved in this process. Despite significant progress, our understanding of ubiquitin-mediated DNA damage response regulation remains incomplete. Here, we have performed a localization screen to identify RING/U-box E3 ligases involved in genome maintenance. Our approach uncovered 7 novel E3 ligases that are recruited to microirradiation stripes, suggesting potential roles in DNA damage signaling and repair. Among these factors, the DELTEX family E3 ligase DTX2 is rapidly mobilized to lesions in a poly ADP-ribosylation-dependent manner. DTX2 is recruited and retained at DSBs via its WWE and DELTEX conserved C-terminal domains. In cells, both domains are required for optimal binding to mono and poly ADP-ribosylated proteins with WWEs playing a prominent role in this process. Supporting its involvement in DSB repair, DTX2 depletion decreases HR efficiency and moderately enhances NHEJ. Furthermore, DTX2 depletion impeded BRCA1 foci formation and increased 53BP1 accumulation at DSBs, suggesting a fine-tuning role for this E3 ligase in repair pathway choice. Finally, DTX2 depletion sensitized cancer cells to X-rays and PARP inhibition and these susceptibilities could be rescued by DTX2 reexpression. Altogether, our work identifies DTX2 as a novel ADP-ribosylation-dependent regulator of HR-mediated DSB repair.
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The ubiquitin proteasome system performs the covalent attachment of lysine 48-linked polyubiquitin chains to substrate proteins, thereby targeting them for degradation, while deubiquitylating enzymes (DUBs) reverse this process. This posttranslational modification regulates key features both of innate and adaptative immunity, including antigen presentation, protein homeostasis and signal transduction. Here we show that loss of one of the most highly expressed DUBs, Otub1, results in changes in murine splenic B cell subsets, leading to a significant increase in marginal zone and transitional B cells and a concomitant decrease in follicular B cells. We demonstrate that Otub1 interacts with the γ-subunit of the heterotrimeric G protein, Gng2, and modulates its ubiquitylation status, thereby controlling Gng2 stability. Proximal mapping of Gng2 revealed an enrichment in partners associated with chemokine signaling, actin cytoskeleton and cell migration. In line with these findings, we show that Otub1-deficient B cells exhibit greater Ca2+ mobilization, F-actin polymerization and chemotactic responsiveness to Cxcl12, Cxcl13 and S1P in vitro, which manifests in vivo as altered localization of B cells within the spleen. Together, our data establishes Otub1 as a novel regulator of G-protein coupled receptor signaling in B cells, regulating their differentiation and positioning in the spleen.
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Quimiotaxis de Leucocito , Enzimas Desubicuitinizantes , Bazo , Ubiquitina , Animales , Ratones , Enzimas Desubicuitinizantes/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Transducción de Señal , Bazo/metabolismo , Ubiquitina/metabolismo , Ubiquitinación , Cisteína Endopeptidasas/metabolismo , Proteínas de Unión al GTP/metabolismo , Linfocitos B/metabolismo , Quimiotaxis de Leucocito/genéticaRESUMEN
Multiple myeloma (MM) is a hematological malignancy that emerges from antibody-producing plasma B cells. Proteasome inhibitors, including the US Food and Drug Administration-approved bortezomib (BTZ) and carfilzomib (CFZ), are frequently used for the treatment of patients with MM. Nevertheless, a significant proportion of patients with MM are refractory or develop resistance to this class of inhibitors, which represents a significant challenge in the clinic. Thus, identifying factors that determine the potency of proteasome inhibitors in MM is of paramount importance to bolster their efficacy in the clinic. Using genome-wide CRISPR-based screening, we identified a subunit of the mitochondrial pyruvate carrier (MPC) complex, MPC1, as a common modulator of BTZ response in 2 distinct human MM cell lines in vitro. We noticed that CRISPR-mediated deletion or pharmacological inhibition of the MPC complex enhanced BTZ/CFZ-induced MM cell death with minimal impact on cell cycle progression. In fact, targeting the MPC complex compromised the bioenergetic capacity of MM cells, which is accompanied by reduced proteasomal activity, thereby exacerbating BTZ-induced cytotoxicity in vitro. Importantly, we observed that the RNA expression levels of several regulators of pyruvate metabolism were altered in advanced stages of MM for which they correlated with poor patient prognosis. Collectively, this study highlights the importance of the MPC complex for the survival of MM cells and their responses to proteasome inhibitors. These findings establish mitochondrial pyruvate metabolism as a potential target for the treatment of MM and an unappreciated strategy to increase the efficacy of proteasome inhibitors in the clinic.
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Antineoplásicos , Mieloma Múltiple , Estados Unidos , Humanos , Inhibidores de Proteasoma/farmacología , Inhibidores de Proteasoma/uso terapéutico , Mieloma Múltiple/tratamiento farmacológico , Mieloma Múltiple/patología , Antineoplásicos/uso terapéutico , Transportadores de Ácidos Monocarboxílicos/uso terapéutico , Bortezomib/farmacología , Bortezomib/uso terapéutico , Piruvatos/uso terapéuticoRESUMEN
BACKGROUND: Imaging tests are one of the most frequently used diagnostic modalities in healthcare, but the benefits of their direct impacts on clinical decision-making have been countered by concerns that they can be overused. Assessing the relative value of imaging tests has largely focused on measures of test accuracy, which overlooks more comprehensive benefits and risks of imaging tests, particularly their impact on patient-centred outcomes (PCOs). We present the findings of the Patient Reported Outcomes of Diagnostics (PROD) research study in response to a methodological gap in the area of diagnostic test comparative effectiveness research. METHODS: Over a 3-year period, the PROD Study engaged with multiple stakeholders to identify existing conceptual models related to PCOs for imaging testing, conducted primary research and evidence synthesis, and developed consensus recommendations to describe and categorise PCOs related to imaging testing. RESULTS: The PROD framework categorises PCOs from imaging studies within four main domains: information or knowledge yielded, physical impact, emotional outcomes and test burden. PCOs interact with each other and influence effects across domains, and can be modified by factors related to the patient, clinical situation, healthcare team and the testing environment. CONCLUSIONS: Using PCOs to inform healthcare decision-making will require ways of collating and presenting information on PCOs in ways that can inform patient-provider decision-making, and developing methods to determine the relative importance of outcomes (including test accuracy) to one another.
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Citocromo P-450 CYP2B1 , Evaluación de Resultado en la Atención de Salud , Humanos , Medición de Resultados Informados por el PacienteRESUMEN
DNA repair by homologous recombination (HR) is critical for the maintenance of genome stability. Germline and somatic mutations in HR genes have been associated with an increased risk of developing breast (BC) and ovarian cancers (OvC). However, the extent of factors and pathways that are functionally linked to HR with clinical relevance for BC and OvC remains unclear. To gain a broader understanding of this pathway, we used multi-omics datasets coupled with machine learning to identify genes that are associated with HR and to predict their sub-function. Specifically, we integrated our phylogenetic-based co-evolution approach (CladePP) with 23 distinct genetic and proteomic screens that monitored, directly or indirectly, DNA repair by HR. This omics data integration analysis yielded a new database (HRbase) that contains a list of 464 predictions, including 76 gold standard HR genes. Interestingly, the spliceosome machinery emerged as one major pathway with significant cross-platform interactions with the HR pathway. We functionally validated 6 spliceosome factors, including the RNA helicase SNRNP200 and its co-factor SNW1. Importantly, their RNA expression correlated with BC/OvC patient outcome. Altogether, we identified novel clinically relevant DNA repair factors and delineated their specific sub-function by machine learning. Our results, supported by evolutionary and multi-omics analyses, suggest that the spliceosome machinery plays an important role during the repair of DNA double-strand breaks (DSBs).
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The heterochromatin protein HP1 plays a central role in the maintenance of genome stability but little is known about how HP1 is controlled. Here, we show that the zinc finger protein POGZ promotes the presence of HP1 at DNA double-strand breaks (DSBs) in human cells. POGZ depletion delays the resolution of DSBs and sensitizes cells to different DNA-damaging agents, including cisplatin and talazoparib. Mechanistically, POGZ promotes homology-directed DNA repair by retaining the BRCA1/BARD1 complex at DSBs in an HP1-dependent manner. In vivo CRISPR inactivation of Pogz is embryonically lethal. Pogz haploinsufficiency (Pogz+ /delta) results in developmental delay, impaired intellectual abilities, hyperactive behaviour and a compromised humoral immune response in mice, recapitulating the main clinical features of the White Sutton syndrome (WHSUS). Pogz+ /delta mice are further radiosensitive and accumulate DSBs in diverse tissues, including the spleen and brain. Altogether, our findings identify POGZ as an important player in homology-directed DNA repair both in vitro and in vivo.
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Homólogo de la Proteína Chromobox 5 , Reparación del ADN , Discapacidad Intelectual , Reparación del ADN por Recombinación , Transposasas , Animales , Homólogo de la Proteína Chromobox 5/genética , Homólogo de la Proteína Chromobox 5/metabolismo , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , ADN , Roturas del ADN de Doble Cadena , Humanos , Discapacidad Intelectual/genética , Ratones , Transposasas/genética , Transposasas/metabolismoRESUMEN
BACKGROUND: Imaging tests are a widely used tool in primary care with many known benefits. Without an understanding of which outcomes matter the most to patients, clinicians are challenged to balance the benefits and harms of imaging tests. This study aimed to explore the perceived impacts imaging tests have on patients from the perspective of the primary care providers (PCPs) and determine PCPs' understanding of patient-centered outcomes (PCOs) from imaging tests. METHODS: Recruitment of PCPs occurred at 4 family medicine clinics in Washington and Idaho. Primary care physicians, nurse practitioners, or physician assistants who order imaging tests were eligible to participate. Semistructured interviews explored providers' perceptions of patient experiences during the process of ordering, performing and following up on imaging tests. Classic content analysis generated themes and subthemes. RESULTS: Sixteen PCPs, including 11 physicians, 3 physician assistants, and 2 nurse practitioners, completed interviews. Two themes were identified: 1) perceived PCOs, and 2) factors influencing the incorporation of PCOs into clinical management. Perceived outcomes included emotions related to the answer a test provides and costs to the patient such as monetary, physical, and added risk. Patient expectations, provider-patient communication, and inadequate knowledge all contributed as barriers to incorporating PCOs into clinical management. DISCUSSION: PCPs recognize different outcomes of imaging tests that they consider important for patients. While providers are perceptive to patient outcomes there remains a challenge to how patient outcomes are used to improve care. Communication with patients and improving provider knowledge are needed to incorporate identified PCOs.
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Actitud del Personal de Salud , Evaluación del Resultado de la Atención al Paciente , Atención Primaria de Salud/organización & administración , Adulto , Anciano , Competencia Clínica , Comunicación , Femenino , Humanos , Masculino , Persona de Mediana Edad , Enfermeras Practicantes/psicología , Enfermeras Practicantes/estadística & datos numéricos , Asistentes Médicos/psicología , Asistentes Médicos/estadística & datos numéricos , Médicos de Atención Primaria/psicología , Médicos de Atención Primaria/estadística & datos numéricos , Atención Primaria de Salud/economía , Investigación Cualitativa , Mejoramiento de la Calidad , Radiografía/psicología , Radiografía/estadística & datos numéricos , Ultrasonografía/economía , Ultrasonografía/psicología , Ultrasonografía/estadística & datos numéricosRESUMEN
The homologous recombination repair (HRR) pathway repairs DNA double-strand breaks in an error-free manner. Mutations in HRR genes can result in increased mutation rate and genomic rearrangements, and are associated with numerous genetic disorders and cancer. Despite intensive research, the HRR pathway is not yet fully mapped. Phylogenetic profiling analysis, which detects functional linkage between genes using coevolution, is a powerful approach to identify factors in many pathways. Nevertheless, phylogenetic profiling has limited predictive power when analyzing pathways with complex evolutionary dynamics such as the HRR. To map novel HRR genes systematically, we developed clade phylogenetic profiling (CladePP). CladePP detects local coevolution across hundreds of genomes and points to the evolutionary scale (e.g., mammals, vertebrates, animals, plants) at which coevolution occurred. We found that multiscale coevolution analysis is significantly more biologically relevant and sensitive to detect gene function. By using CladePP, we identified dozens of unrecognized genes that coevolved with the HRR pathway, either globally across all eukaryotes or locally in different clades. We validated eight genes in functional biological assays to have a role in DNA repair at both the cellular and organismal levels. These genes are expected to play a role in the HRR pathway and might lead to a better understanding of missing heredity in HRR-associated cancers (e.g., heredity breast and ovarian cancer). Our platform presents an innovative approach to predict gene function, identify novel factors related to different diseases and pathways, and characterize gene evolution.
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Evolución Molecular , Reparación del ADN por Recombinación , Programas Informáticos , Animales , Enzimas Reparadoras del ADN/genética , Sitios Genéticos , Filogenia , Plantas/genéticaRESUMEN
DNA double-strand breaks (DSBs) can be repaired by two major pathways: non-homologous end-joining (NHEJ) and homologous recombination (HR). DNA repair pathway choice is governed by the opposing activities of 53BP1, in complex with its effectors RIF1 and REV7, and BRCA1. However, it remains unknown how the 53BP1/RIF1/REV7 complex stimulates NHEJ and restricts HR to the S/G2 phases of the cell cycle. Using a mass spectrometry (MS)-based approach, we identify 11 high-confidence REV7 interactors and elucidate the role of SHLD2 (previously annotated as FAM35A and RINN2) as an effector of REV7 in the NHEJ pathway. FAM35A depletion impairs NHEJ-mediated DNA repair and compromises antibody diversification by class switch recombination (CSR) in B cells. FAM35A accumulates at DSBs in a 53BP1-, RIF1-, and REV7-dependent manner and antagonizes HR by limiting DNA end resection. In fact, FAM35A is part of a larger complex composed of REV7 and SHLD1 (previously annotated as C20orf196 and RINN3), which promotes NHEJ and limits HR Together, these results establish SHLD2 as a novel effector of REV7 in controlling the decision-making process during DSB repair.
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Proteínas de Ciclo Celular/metabolismo , Roturas del ADN de Doble Cadena , Reparación del ADN por Unión de Extremidades , Proteínas de Unión al ADN/metabolismo , Proteínas Mad2/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Unión al ADN/genética , Fase G2/genética , Células HEK293 , Humanos , Proteínas Mad2/genética , Fase S/genética , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo , Proteína 1 de Unión al Supresor Tumoral P53/genética , Proteína 1 de Unión al Supresor Tumoral P53/metabolismoRESUMEN
Encouraging patients and consumers to use data and other information in choosing health care providers is an important way to enhance patient engagement and improve the quality of care. The growing use of technology, including smart phones and near-ubiquitous Internet access, provides consumers with easy access to websites that collect and report assessments and ratings of providers, primarily physicians and hospitals. In addition to new technology, recent laws and changes in society and the delivery of care are laying the foundation for greater use by consumers of provider performance report cards. Such use could be accelerated if the shortcomings of current report card efforts were addressed. Recommendations include making online report cards easier to use and more understandable, engaging, substantive, and relevant to consumers' health and medical concerns and choices.
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Comportamiento del Consumidor/estadística & datos numéricos , Servicios de Información/estadística & datos numéricos , Internet/estadística & datos numéricos , Mejoramiento de la Calidad , Indicadores de Calidad de la Atención de Salud , Femenino , Encuestas de Atención de la Salud/métodos , Encuestas de Atención de la Salud/estadística & datos numéricos , Humanos , Difusión de la Información/métodos , Masculino , Prioridad del Paciente/estadística & datos numéricos , Pautas de la Práctica en Medicina/estadística & datos numéricos , Estados UnidosRESUMEN
Consumers Union, publisher of Consumer Reports magazine, has used the drug class reviews of the Drug Effectiveness Review Project (DERP) as one critical component of a free public information project on the comparative effectiveness, safety, and cost of prescription drugs. The project translates the DERP findings for consumers. Drawing on other sources and adding information on drug costs, the project chooses Best Buy drugs in each category it evaluates. This guidance can help consumers save up to thousands of dollars per year, and it has the potential to reduce overall drug spending.
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Revisión de la Utilización de Medicamentos/economía , Medicina Basada en la Evidencia , Organizaciones del Consumidor , Análisis Costo-Beneficio , Toma de Decisiones , Eficiencia Organizacional , Formularios Farmacéuticos como Asunto , Federación para Atención de Salud , Humanos , EdiciónRESUMEN
A key strategy for driving improvements in health care quality is providing comparative quality information to consumers. This strategy will not work, and could even be counterproductive, unless (1) consumers are convinced that quality problems are real and consequential and that quality can be improved; (2) purchasers and policymakers make sure that quality reporting is standardized and universal; (3) consumers are given quality information that is relevant and easy to understand and use; (4) the dissemination of quality information is improved; and (5) purchasers reward quality improvements and providers create the information and organizational infrastructure to achieve them.
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Benchmarking , Comportamiento del Consumidor , Revelación/normas , Difusión de la Información , Garantía de la Calidad de Atención de Salud/métodos , Indicadores de Calidad de la Atención de Salud , Defensa del Consumidor , Toma de Decisiones , Guías como Asunto , Humanos , Estados UnidosRESUMEN
The growth in mass media drug advertising has coincided with a rapid rise in spending on prescription drugs in the United States. Our research examines the potential link between increased ad spending and the rising use and sale of these drugs. We found strong circumstantial evidence that ad spending is one of many important elements driving the sharp rise in pharmaceutical sales to consumers.