RESUMEN
Various lines of evidence suggest a mechanistic role for altered cAMP-CREB (cAMP response element - binding protein) signaling in depressive and affective disorders. However, the establishment and validation of human inter-individual differences in this and other major signaling pathways has proven difficult. Here, we describe a novel lentiviral methodology to investigate signaling variation over long periods of time directly in human primary fibroblasts. On a cellular level, this method showed surprisingly large inter-individual differences in three major signaling pathways in human subjects that nevertheless correlated with cellular measures of genome-wide transcription and drug toxicity. We next validated this method by establishing a likely role for cAMP-mediated signaling in a human neuroendocrine response to light - the light-dependent suppression of the circadian hormone melatonin - that shows wide inter-individual differences of unknown origin in vivo. Finally, we show an overall greater magnitude of cellular CREB signaling in individuals with bipolar disorder, suggesting a possible role for this signaling pathway in susceptibility to mental disease. Overall, our results suggest that genetic differences in major signaling pathways can be reliably detected with sensitive viral-based reporter profiling, and that these differences can be conserved across tissues and be predictive of physiology and disease susceptibility.
Asunto(s)
Trastorno Bipolar/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Luz , Melatonina/metabolismo , Adulto , Células Cultivadas , Estudios de Cohortes , Femenino , Fibroblastos/metabolismo , Vectores Genéticos , Humanos , Lentivirus/genética , Masculino , Persona de Mediana Edad , Estimulación Luminosa , Transducción de Señal , Población Blanca , Adulto JovenRESUMEN
The SS18-SSX fusion drives oncogenic transformation in synovial sarcoma by bridging SS18, a member of the mSWI/SNF (BAF) complex, to Polycomb repressive complex 1 (PRC1) target genes. Here we show that the ability of SS18-SSX to occupy H2AK119ub1-rich regions is an intrinsic property of its SSX C terminus, which can be exploited by fusion to transcriptional regulators beyond SS18. Accordingly, SS18-SSX recruitment occurs in a manner that is independent of the core components and catalytic activity of BAF. Alternative SSX fusions are also recruited to H2AK119ub1-rich chromatin and reproduce the expression signatures of SS18-SSX by engaging with transcriptional activators. Variant Polycomb repressive complex 1.1 (PRC1.1) acts as the main depositor of H2AK119ub1 and is therefore required for SS18-SSX occupancy. Importantly, the SSX C terminus not only depends on H2AK119ub1 for localization, but also further increases it by promoting PRC1.1 complex stability. Consequently, high H2AK119ub1 levels are a feature of murine and human synovial sarcomas. These results uncover a critical role for SSX-C in mediating gene deregulation in synovial sarcoma by providing specificity to chromatin and further enabling oncofusion binding by enhancing PRC1.1 stability and H2AK119ub1 deposition.
Asunto(s)
Sarcoma Sinovial , Humanos , Animales , Ratones , Sarcoma Sinovial/genética , Sarcoma Sinovial/metabolismo , Complejo Represivo Polycomb 1/genética , Activación Transcripcional , Núcleo Celular/metabolismo , Cromatina/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Proteínas de Ciclo Celular/metabolismoRESUMEN
Meiotic recombination rates vary across the genome, often involving localized crossover hotspots and coldspots. Studying the molecular basis and mechanisms underlying this variation has been challenging due to the high cost and effort required to construct individualized genome-wide maps of recombination crossovers. Here we introduce a new method, called ReMIX, to detect crossovers from gamete DNA of a single individual using Illumina sequencing of 10X Genomics linked-read libraries. ReMIX reconstructs haplotypes and identifies the valuable rare molecules spanning crossover breakpoints, allowing quantification of the genomic location and intensity of meiotic recombination. Using a single mouse and stickleback fish, we demonstrate how ReMIX faithfully recovers recombination hotspots and landscapes that have previously been built using hundreds of offspring. ReMIX provides a high-resolution, high-throughput, and low-cost approach to quantify recombination variation across the genome, providing an exciting opportunity to study recombination among multiple individuals in diverse organisms.
Asunto(s)
Mapeo Cromosómico/métodos , Biología Computacional/métodos , Genoma , Recombinación Homóloga/genética , Recombinación Genética , Algoritmos , Animales , Puntos de Rotura del Cromosoma , Intercambio Genético , Bases de Datos Genéticas , Evolución Molecular , Ligamiento Genético/genética , Genómica , Haplotipos , Secuenciación de Nucleótidos de Alto Rendimiento , Masculino , Meiosis , Ratones , Ratones Endogámicos C57BL , Espermatozoides , BazoRESUMEN
The importance of natural gene expression variation for human behavior is undisputed, but its impact on circadian physiology remains mostly unexplored. Using umbilical cord fibroblasts, we have determined by genome-wide association how common genetic variation impacts upon cellular circadian function. Gene set enrichment points to differences in protein catabolism as one major source of clock variation in humans. The two most significant alleles regulated expression of COPS7B, a subunit of the COP9 signalosome. We further show that the signalosome complex is imported into the nucleus in timed fashion to stabilize the essential circadian protein BMAL1, a novel mechanism to oppose its proteasome-mediated degradation. Thus, circadian clock properties depend in part upon a genetically-encoded competition between stabilizing and destabilizing forces, and genetic alterations in these mechanisms provide one explanation for human chronotype.
Asunto(s)
Variación Biológica Poblacional , Ritmo Circadiano , Regulación de la Expresión Génica , Variación Genética , Factores de Transcripción ARNTL/metabolismo , Complejo del Señalosoma COP9/metabolismo , Estudio de Asociación del Genoma Completo , Humanos , Estabilidad Proteica , Proteínas/metabolismoRESUMEN
Circadian clocks are present in most cells and are essential for maintenance of daily rhythms in physiology, mood, and cognition. Thus, not only neurons of the central circadian pacemaker but also many other peripheral tissues possess the same functional and self-sustained circadian clocks. Surprisingly, however, their properties vary widely within the human population. In recent years, this clock variance has been studied extensively both in health and in disease using robust lentivirus-based reporter technologies to probe circadian function in human peripheral cells as proxies for those in neurologically and physiologically relevant but inaccessible tissues. The same procedures can be used to investigate other conserved signal transduction cascades affecting multiple aspects of human physiology, behavior, and disease. Accessing gene expression variation within human populations via these powerful in vitro cell-based technologies could provide important insights into basic phenotypic diversity or to better interpret patterns of gene expression variation in disease.
Asunto(s)
Relojes Circadianos/fisiología , Animales , Perfilación de la Expresión Génica , Humanos , Ratones , Transducción de SeñalRESUMEN
The NONO protein has been characterized as an important transcriptional regulator in diverse cellular contexts. Here we show that loss of NONO function is a likely cause of human intellectual disability and that NONO-deficient mice have cognitive and affective deficits. Correspondingly, we find specific defects at inhibitory synapses, where NONO regulates synaptic transcription and gephyrin scaffold structure. Our data identify NONO as a possible neurodevelopmental disease gene and highlight the key role of the DBHS protein family in functional organization of GABAergic synapses.