RESUMEN
BACKGROUND: Strains from a collection of Drosophila GFP protein trap lines express GFP in the normal tissues where the endogenous protein is present. This collection can be used to screen for proteins distributed in the nucleus in a non-uniform pattern. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed four lines that show peripheral or punctate nuclear staining. One of these lines affects an uncharacterized gene named CG11138. The CG11138 protein shows a punctate distribution in the nuclear periphery similar to that of Drosophila insulator proteins but does not co-localize with known insulators. Interestingly, mutations in Lamin proteins result in alterations in CG11138 localization, suggesting that this protein may be a novel component of the nuclear lamina. A second line affects the Decondensation factor 31 (Df31) gene, which encodes a protein with a unique nuclear distribution that appears to segment the nucleus into four different compartments. The X-chromosome of males is confined to one of these compartments. We also find that Drosophila Nucleoplasmin (dNlp) is present in regions of active transcription. Heat shock leads to loss of dNlp from previously transcribed regions of polytene chromosome without redistribution to the heat shock genes. Analysis of Stonewall (Stwl), a protein previously found to be necessary for the maintenance of germline stem cells, shows that Stwl is present in a punctate pattern in the nucleus that partially overlaps with that of known insulator proteins. Finally we show that Stwl, dNlp, and Df31 form part of a highly interactive network. The properties of other components of this network may help understand the role of these proteins in nuclear biology. CONCLUSIONS/SIGNIFICANCE: These results establish screening of GFP protein trap alleles as a strategy to identify factors with novel cellular functions. Information gained from the analysis of CG11138 Stwl, dNlp, and Df31 sets the stage for future studies of these proteins.
Asunto(s)
Núcleo Celular/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citología , Drosophila melanogaster/metabolismo , Proteínas Fluorescentes Verdes/metabolismo , Animales , Diferenciación Celular , Diploidia , Drosophila melanogaster/enzimología , Células Germinativas/citología , Células Germinativas/metabolismo , Respuesta al Choque Térmico , Lamina Tipo A/metabolismo , Masculino , Modelos Biológicos , Lámina Nuclear/enzimología , Nucleoplasminas , Cromosomas Politénicos/metabolismo , Mapas de Interacción de Proteínas , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Insulators are multiprotein-DNA complexes thought to affect gene expression by mediating inter- and intrachromosomal interactions. Drosophila insulators contain specific DNA-binding proteins plus common components, such as CP190, that facilitate these interactions. Here, we examine changes in the distribution of Drosophila insulator proteins during the heat-shock and ecdysone responses. We find that CP190 recruitment to insulator sites is the main regulatable step in controlling insulator function during heat shock. In contrast, both CP190 and DNA-binding protein recruitment are regulated during the ecdysone response. CP190 is necessary to stabilize specific chromatin loops and for proper activation of transcription of genes regulated by this hormone. These findings suggest that cells may regulate recruitment of insulator proteins to DNA to activate insulator activity at specific sites and create distinct patterns of nuclear organization that are necessary to achieve proper gene expression in response to different stimuli.
Asunto(s)
Cromatina/metabolismo , Regulación del Desarrollo de la Expresión Génica , Animales , Cromosomas/química , ADN/química , Proteínas de Unión al ADN/metabolismo , Drosophila , Proteínas de Drosophila/metabolismo , Ecdisona/farmacología , Perfilación de la Expresión Génica , Genoma , Proteínas de Choque Térmico/metabolismo , Microscopía Fluorescente/métodos , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Nucleares/metabolismo , Unión Proteica , Transcripción GenéticaRESUMEN
Increasing the college graduation rates of underrepresented minority students in science disciplines is essential to attain a diverse workforce for the 21st century. The Research Internship and Science Education (RISE) program attempts to motivate and prepare students from the Atlanta Public School system, where underrepresented minority (URM) students comprise a majority of the population, for biomedical science careers by offering the opportunity to participate in an original research project. Students work in a research laboratory from the summer of their sophomore year until graduation, mentored by undergraduate and graduate students and postdoctoral fellows (postdocs). In addition, they receive instruction in college-level biology, scholastic assessment test (SAT) preparation classes, and help with the college application process. During the last 4 yr, RISE students have succeeded in the identification and characterization of a series of proteins involved in the regulation of nuclear organization and transcription. All but 1 of 39 RISE students have continued on to 4-year college undergraduate studies and 61% of those students are currently enrolled in science-related majors. These results suggest that the use of research-based experiences at the high school level may contribute to the increased recruitment of underrepresented students into science-related careers.
Asunto(s)
Investigación Biomédica , Movilidad Laboral , Estudiantes , Escolaridad , Georgia , MentoresRESUMEN
Appropriate cell number and organ size in a multicellular organism are determined by coordinated cell growth, proliferation, and apoptosis. Disruption of these processes can cause cancer. Recent studies have identified the Large tumor suppressor (Lats)/Warts (Wts) protein kinase as a key component of a pathway that controls the coordination between cell proliferation and apoptosis. Here we describe growth inhibitory functions for a Mob superfamily protein, termed Mats (Mob as tumor suppressor), in Drosophila. Loss of Mats function results in increased cell proliferation, defective apoptosis, and induction of tissue overgrowth. We show that mats and wts function in a common pathway. Mats physically associates with Wts to stimulate the catalytic activity of the Wts kinase. A human Mats ortholog (Mats1) can rescue the lethality associated with loss of Mats function in Drosophila. As Mats1 is mutated in human tumors, Mats-mediated growth inhibition and tumor suppression is likely conserved in humans.
Asunto(s)
Apoptosis/fisiología , Proliferación Celular , Transformación Celular Neoplásica/metabolismo , Proteínas de Drosophila/metabolismo , Neoplasias/metabolismo , Proteínas Quinasas/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Animales , Línea Celular , Transformación Celular Neoplásica/genética , Secuencia Conservada , ADN Complementario/análisis , ADN Complementario/genética , Drosophila , Proteínas de Drosophila/genética , Proteínas de Drosophila/aislamiento & purificación , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica/genética , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Microscopía Electrónica de Rastreo , Datos de Secuencia Molecular , Mutación/genética , Neoplasias/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Transferasas (Grupos de Otros Fosfatos Sustitutos) , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/aislamiento & purificaciónRESUMEN
Yan is a nuclear DNA-binding protein that acts as a general inhibitor of cellular differentiation and proliferation in Drosophila melanogaster. The genetic and biochemical mechanisms required for regulating Yan protein function are well understood, however, the molecular mechanism of yan gene transcriptional regulation has not been fully elucidated. Here we show that the dynamic expression of the yan gene is specified by distinct spatial and temporal cis-acting regulatory elements in embryos and larval tissues. Each of these distinct elements is thus capable of replicating vital aspects of endogenous yan gene expression.
Asunto(s)
Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Proteínas del Ojo/genética , Regulación del Desarrollo de la Expresión Génica/genética , Secuencias Reguladoras de Ácidos Nucleicos/genética , Proteínas Represoras/genética , Animales , Sistema Nervioso Central/embriología , Sistema Nervioso Central/crecimiento & desarrollo , Sistema Nervioso Central/metabolismo , ADN , Proteínas de Drosophila/fisiología , Drosophila melanogaster/embriología , Drosophila melanogaster/crecimiento & desarrollo , Embrión no Mamífero/fisiología , Ojo/embriología , Ojo/crecimiento & desarrollo , Ojo/metabolismo , Proteínas del Ojo/fisiología , Femenino , Genes Reporteros , Operón Lac , Larva/fisiología , Ovario/embriología , Ovario/crecimiento & desarrollo , Ovario/metabolismo , Proteínas Represoras/fisiologíaRESUMEN
Receptor tyrosine kinase (RTK) signaling plays an instructive role in cell fate decisions, whereas Notch signaling is often involved in restricting cellular competence for differentiation. Genetic interactions between these two evolutionarily conserved pathways have been extensively documented. The underlying molecular mechanisms, however, are not well understood. Here, we show that Yan, an Ets transcriptional repressor that blocks cellular potential for specification and differentiation, is a target of Notch signaling during Drosophila eye development. The Suppressor of Hairless (Su[H]) protein of the Notch pathway is required for activating yan expression, and Su(H) binds directly to an eye-specific yan enhancer in vitro. In contrast, yan expression is repressed by Pointed (Pnt), which is a key component of the RTK pathway. Pnt binds specifically to the yan enhancer and competes with Su(H) for DNA binding. This competition illustrates a potential mechanism for RTK and Notch signals to oppose each other. Thus, yan serves as a common target of Notch/Su(H) and RTK/Pointed signaling pathways during cell fate specification.