RESUMEN
Priority effects, where arrival order and initial relative abundance modulate local species interactions, can exert taxonomic, functional, and evolutionary influences on ecological communities by driving them to alternative states. It remains unclear if these wide-ranging consequences of priority effects can be explained systematically by a common underlying factor. Here, we identify such a factor in an empirical system. In a series of field and laboratory studies, we focus on how pH affects nectar-colonizing microbes and their interactions with plants and pollinators. In a field survey, we found that nectar microbial communities in a hummingbird-pollinated shrub, Diplacus (formerly Mimulus) aurantiacus, exhibited abundance patterns indicative of alternative stable states that emerge through domination by either bacteria or yeasts within individual flowers. In addition, nectar pH varied among D. aurantiacus flowers in a manner that is consistent with the existence of these alternative stable states. In laboratory experiments, Acinetobacter nectaris, the bacterium most commonly found in D. aurantiacus nectar, exerted a strongly negative priority effect against Metschnikowia reukaufii, the most common nectar-specialist yeast, by reducing nectar pH. This priority effect likely explains the mutually exclusive pattern of dominance found in the field survey. Furthermore, experimental evolution simulating hummingbird-assisted dispersal between flowers revealed that M. reukaufii could evolve rapidly to improve resistance against the priority effect if constantly exposed to A. nectaris-induced pH reduction. Finally, in a field experiment, we found that low nectar pH could reduce nectar consumption by hummingbirds, suggesting functional consequences of the pH-driven priority effect for plant reproduction. Taken together, these results show that it is possible to identify an overarching factor that governs the eco-evolutionary dynamics of priority effects across multiple levels of biological organization.
Asunto(s)
Néctar de las Plantas , Polinización , Animales , Flores , Aves , Plantas , Levaduras , BacteriasRESUMEN
During a critical period in the developing mammalian brain, there is a major switch in the nature of GABAergic transmission from depolarizing and excitatory, the pattern of the neonatal brain, to hyperpolarizing and inhibitory, the pattern of the mature brain. This switch is believed to play a major role in determining neuronal connectivity via activity-dependent mechanisms. The GABAergic developmental switch may also be particularly vulnerable to dysfunction leading to seizure disorders. The developmental GABA switch is mediated primarily by KCC2, a neuronal K+/Cl- cotransporter that determines the intracellular concentration of Cl- and, hence, the reversal potential for GABA. Here, we report that kazachoc (kcc) mutations that reduce the level of the sole K+/Cl- cotransporter in the fruitfly Drosophila melanogaster render flies susceptible to epileptic-like seizures. Drosophila kcc protein is widely expressed in brain neuropil, and its level rises with developmental age. Young kcc mutant flies with low kcc levels display behavioral seizures and demonstrate a reduced threshold for seizures induced by electroconvulsive shock. The kcc mutation enhances a series of other Drosophila epilepsy mutations indicating functional interactions leading to seizure disorder. Both genetic and pharmacological experiments suggest that the increased seizure susceptibility of kcc flies occurs via excitatory GABAergic signaling. The kcc mutants provide an excellent model system in which to investigate how modulation of GABAergic signaling influences neuronal excitability and epileptogenesis.
Asunto(s)
Drosophila melanogaster/genética , Predisposición Genética a la Enfermedad , Mutación , Convulsiones/genética , Simportadores/genética , Animales , Mapeo Cromosómico , Drosophila melanogaster/metabolismo , Fenotipo , Convulsiones/fisiopatología , Transducción de Señal , Simportadores/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Cotransportadores de K ClRESUMEN
The opportunity to engage in scientific research is an important, but often neglected, component of undergraduate training in biology. We describe the curriculum for an innovative, course-based undergraduate research experience (CURE) appropriate for a large, introductory cell and molecular biology laboratory class that leverages students' high level of interest in cancer. The course is highly collaborative and emphasizes the analysis and interpretation of original scientific data. During the course, students work in teams to characterize a collection of mutations in the human p53 tumor suppressor gene via expression and analysis in yeast. Initially, student pairs use both qualitative and quantitative assays to assess the ability of their p53 mutant to activate expression of reporter genes, and they localize their mutation within the p53 structure. Through facilitated discussion, students suggest possible molecular explanations for the transactivation defects displayed by their p53 mutants and propose experiments to test these hypotheses that they execute during the second part of the course. They use a western blot to determine whether mutant p53 levels are reduced, a DNA-binding assay to test whether recognition of any of three p53 target sequences is compromised, and fluorescence microscopy to assay nuclear localization. Students studying the same p53 mutant periodically convene to discuss and interpret their combined data. The course culminates in a poster session during which students present their findings to peers, instructors, and the greater biosciences community. Based on our experience, we provide recommendations for the development of similar large introductory lab courses. © 2016 by The International Union of Biochemistry and Molecular Biology, 45(2):161-178, 2017.
Asunto(s)
Bioensayo/métodos , Investigación Biomédica/educación , Biología Celular/educación , Biología Molecular/educación , Mutación/genética , Aprendizaje Basado en Problemas/métodos , Saccharomyces cerevisiae/genética , Proteína p53 Supresora de Tumor/genética , Curriculum , Educación de Pregrado en Medicina , Genes Supresores de Tumor , Humanos , Laboratorios/normas , EstudiantesRESUMEN
Suppressor mutations provide potentially powerful tools for examining mechanisms underlying neurological disorders and identifying novel targets for pharmacological intervention. Here we describe mutations that suppress seizures in a Drosophila model of human epilepsy. A screen utilizing the Drosophila easily shocked (eas) "epilepsy" mutant identified dominant suppressors of seizure sensitivity. Among several mutations identified, neuronal escargot (esg) reduced eas seizures almost 90%. The esg gene encodes a member of the snail family of transcription factors. Whereas esg is normally expressed in a limited number of neurons during a defined period of nervous system development, here normal esg was expressed in all neurons and throughout development. This greatly ameliorated both the electrophysiological and the behavioral epilepsy phenotypes of eas. Neuronal esg appears to act as a general seizure suppressor in the Drosophila epilepsy model as it reduces the susceptibility of several seizure-prone mutants. We observed that esg must be ectopically expressed during nervous system development to reduce seizure susceptibility in adults. Furthermore, induction of esg in a small subset of neurons (interneurons) will reduce seizure susceptibility. A combination of microarray and computational analyses revealed 100 genes that represent possible targets of neuronal esg. We anticipate that some of these genes may ultimately serve as targets for novel antiepileptic drugs.
Asunto(s)
Proteínas de Drosophila/genética , Drosophila/genética , Epilepsia/genética , Mutación , Convulsiones/genética , Animales , Modelos Animales de Enfermedad , GenotipoRESUMEN
We present an innovative course-based undergraduate research experience curriculum focused on the characterization of single point mutations in p53, a tumor suppressor gene that is mutated in more than 50% of human cancers. This course is required of all introductory biology students, so all biology majors engage in a research project as part of their training. Using a set of open-ended written prompts, we found that the course shifts student conceptions of what it means to think like a scientist from novice to more expert-like. Students at the end of the course identified experimental repetition, data analysis, and collaboration as important elements of thinking like a scientist. Course exams revealed that students showed gains in their ability to analyze and interpret data. These data indicate that this course-embedded research experience has a positive impact on the development of students' conceptions and practice of scientific thinking.
Asunto(s)
Formación de Concepto , Curriculum , Investigación/educación , Ciencia/educación , Estudiantes , Pensamiento , Universidades , Femenino , Humanos , Laboratorios , Masculino , Evaluación de Programas y Proyectos de Salud , Estadística como AsuntoRESUMEN
The kcc(DHS1) allele of kazachoc (kcc) was identified as a seizure-enhancer mutation exacerbating the bang-sensitive (BS) paralytic behavioral phenotypes of several seizure-sensitive Drosophila mutants. On their own, young kcc(DHS1) flies also display seizure-like behavior and demonstrate a reduced threshold for seizures induced by electroconvulsive shock. The product of kcc shows substantial homology to KCC2, the mammalian neuronal K(+)-Cl(-) cotransporter. The kcc(DHS1) allele is a hypomorph, and its seizure-like phenotype reflects reduced expression of the kcc gene. We report here that kcc functions as a K(+)-Cl(-) cotransporter when expressed heterologously in Xenopus laevis oocytes: under hypotonic conditions that induce oocyte swelling, oocytes that express Drosophila kcc display robust ion transport activity observed as a Cl(-)-dependent uptake of the K(+) congener (86)Rb(+). Ectopic, spatially restricted expression of a UAS-kcc(+) transgene was used to determine where cotransporter function is required in order to rescue the kcc(DHS1) BS paralytic phenotype. Interestingly, phenotypic rescue is largely accounted for by targeted, circumscribed expression in the mushroom bodies (MBs) and the ellipsoid body (EB) of the central complex. Intriguingly, we observed that MB induction of kcc(+) functioned as a general seizure suppressor in Drosophila. Drosophila MBs have generated considerable interest especially for their role as the neural substrate for olfactory learning and memory; they have not been previously implicated in seizure susceptibility. We show that kcc(DHS1) seizure sensitivity in MB neurons acts via a weakening of chemical synaptic inhibition by GABAergic transmission and suggest that this is due to disruption of intracellular Cl(-) gradients in MB neurons.
Asunto(s)
Encéfalo/metabolismo , Drosophila melanogaster/metabolismo , Proteínas de Insectos/genética , Cuerpos Pedunculados/metabolismo , Convulsiones/genética , Simportadores/genética , Simportadores/metabolismo , Animales , Conducta Animal , Encéfalo/citología , Encéfalo/fisiología , Susceptibilidad a Enfermedades/metabolismo , Drosophila melanogaster/citología , Drosophila melanogaster/fisiología , Fenómenos Electrofisiológicos , Femenino , Expresión Génica , Humanos , Proteínas de Insectos/metabolismo , Masculino , Cuerpos Pedunculados/citología , Cuerpos Pedunculados/fisiología , Plasticidad Neuronal/genética , Fenotipo , Sinapsis/metabolismo , Transmisión Sináptica , Xenopus laevis/genética , Ácido gamma-Aminobutírico/metabolismo , Cotransportadores de K ClRESUMEN
Olfaction is of considerable importance to many insects in behaviors critical for survival and reproduction, including location of food sources, selection of mates, recognition of colony con-specifics, and determination of oviposition sites. An ubiquitous, but poorly understood, component of the insect's olfactory system is a group of odorant-binding proteins (OBPs) that are present at high concentrations in the aqueous lymph surrounding the dendrites of olfactory receptor neurons. OBPs are believed to shuttle odorants from the environment to the underlying odorant receptors, for which they could potentially serve as odorant presenters. Here we show that the Drosophila genome carries 51 potential OBP genes, a number comparable to that of its odorant-receptor genes. We find that the majority (73%) of these OBP-like genes occur in clusters of as many as nine genes, in contrast to what has been observed for the Drosophila odorant-receptor genes. Two of the presumptive OBP gene clusters each carries an odorant-receptor gene. We also report an intriguing subfamily of 12 putative OBPs that share a unique C-terminal structure with three conserved cysteines and a conserved proline. Members of this subfamily have not previously been described for any insect. We have performed phylogenetic analyses of the OBP-related proteins in Drosophila as well as other insects, and we discuss the duplication and divergence of the genes for this large family. [The sequence data from this study have been submitted to FlyBase. Annotations for these sequences are available as supplementary material at http://www.genome.org.]