RESUMEN
Carbon dioxide (CO2) elicits different olfactory behaviors across species. In Drosophila, neurons that detect CO2 are located in the antenna, form connections in a ventral glomerulus in the antennal lobe, and mediate avoidance. By contrast, in the mosquito these neurons are in the maxillary palps (MPs), connect to medial sites, and promote attraction. We found in Drosophila that loss of a microRNA, miR-279, leads to formation of CO2 neurons in the MPs. miR-279 acts through down-regulation of the transcription factor Nerfin-1. The ectopic neurons are hybrid cells. They express CO2 receptors and form connections characteristic of CO2 neurons, while exhibiting wiring and receptor characteristics of MP olfactory receptor neurons (ORNs). We propose that this hybrid ORN reveals a cellular intermediate in the evolution of species-specific behaviors elicited by CO2.
Asunto(s)
Dióxido de Carbono/análisis , Proteínas de Drosophila/genética , Drosophila/fisiología , MicroARNs/metabolismo , Neuronas Receptoras Olfatorias/fisiología , Receptores de Superficie Celular/metabolismo , Factores de Transcripción/genética , Regiones no Traducidas 3' , Animales , Animales Modificados Genéticamente , Dióxido de Carbono/metabolismo , Drosophila/genética , Proteínas de Drosophila/metabolismo , Regulación de la Expresión Génica , Células Híbridas/fisiología , MicroARNs/genética , Mutación , Neuronas Receptoras Olfatorias/citología , Órganos de los Sentidos/fisiología , Especificidad de la Especie , Factores de Transcripción/metabolismoRESUMEN
Blood-feeding insects, including the malaria mosquito Anopheles gambiae, use highly specialized and sensitive olfactory systems to locate their hosts. This is accomplished by detecting and following plumes of volatile host emissions, which include carbon dioxide (CO2). CO2 is sensed by a population of olfactory sensory neurons in the maxillary palps of mosquitoes and in the antennae of the more genetically tractable fruitfly, Drosophila melanogaster. The molecular identity of the chemosensory CO2 receptor, however, remains unknown. Here we report that CO2-responsive neurons in Drosophila co-express a pair of chemosensory receptors, Gr21a and Gr63a, at both larval and adult life stages. We identify mosquito homologues of Gr21a and Gr63a, GPRGR22 and GPRGR24, and show that these are co-expressed in A. gambiae maxillary palps. We show that Gr21a and Gr63a together are sufficient for olfactory CO2-chemosensation in Drosophila. Ectopic expression of Gr21a and Gr63a together confers CO2 sensitivity on CO2-insensitive olfactory neurons, but neither gustatory receptor alone has this function. Mutant flies lacking Gr63a lose both electrophysiological and behavioural responses to CO2. Knowledge of the molecular identity of the insect olfactory CO2 receptors may spur the development of novel mosquito control strategies designed to take advantage of this unique and critical olfactory pathway. This in turn could bolster the worldwide fight against malaria and other insect-borne diseases.
Asunto(s)
Dióxido de Carbono/análisis , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Neuronas Aferentes/metabolismo , Animales , Anopheles/metabolismo , Conducta Animal/efectos de los fármacos , Dióxido de Carbono/metabolismo , Dióxido de Carbono/farmacología , Proteínas de Drosophila/genética , Drosophila melanogaster/efectos de los fármacos , Drosophila melanogaster/genética , Drosophila melanogaster/crecimiento & desarrollo , Electrofisiología , Humanos , Proteínas de Insectos/genética , Proteínas de Insectos/metabolismo , Larva/genética , Larva/metabolismo , Datos de Secuencia Molecular , Mutación/genética , Neuronas Aferentes/efectos de los fármacosRESUMEN
Fibroblast growth factors (FGFs) are important regulators of retinal development and survival. We examined the expression and distribution of FGF9 and its preferred receptors FGFR2IIIc and FGFR3IIIc in this tissue. FGF9 transcripts in whole rat retina were detected by RT-PCR but were not present in purified cultured Muller glia. Transcripts appeared as 3.2-kb and 4.0-kb bands on Northern blots, and Western blotting of whole retina revealed FGF9-immunoreactive bands at 30 and 55 kDa. FGF9 mRNA demonstrated a biphasic expression profile, elevated at birth and adulthood, but relatively decreased during terminal retinal differentiation (4-14 days postnatal). Antibody labeling broadly reflected these findings: staining in vivo was observed mainly in the inner retina (and outer plexiform layer in adults) whereas FGF9 was not detectable in cultured Muller glia. In adults, FGF9 in situ hybridization also showed a detectable signal in inner retina. FGFR2IIIc and FGFR3IIIc were detected by RT-PCR, and Western blotting showed both FGFRs existed as multiple forms between approximately 100-200 kDa. FGFR2 and FGFR3 antibodies showed prominent labeling in the inner retina, especially in proliferating cultured Muller glia. Exogenous FGF9 elicited a dose-dependent increase in Muller glial proliferation in vitro. These data suggest a role for FGF9 in retinal differentiation and maturation, possibly representing a neuronally derived factor acting upon glial (and other) cells.
Asunto(s)
Diferenciación Celular/fisiología , Factores de Crecimiento de Fibroblastos/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Tirosina Quinasas Receptoras/metabolismo , Receptores de Factores de Crecimiento de Fibroblastos/metabolismo , Retina/metabolismo , Envejecimiento/metabolismo , Animales , Animales Recién Nacidos , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Factor 9 de Crecimiento de Fibroblastos , Factores de Crecimiento de Fibroblastos/genética , Factores de Crecimiento de Fibroblastos/farmacología , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/fisiología , Neuroglía/efectos de los fármacos , Neuroglía/metabolismo , Neuronas/metabolismo , Isoformas de Proteínas/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos , Receptor Tipo 3 de Factor de Crecimiento de Fibroblastos , Retina/crecimiento & desarrolloRESUMEN
Individual members of the E2F/DP protein family control cell cycle progression by acting predominantly as an activator or repressor of transcription. In Drosophila melanogaster the E2f1, E2f2, Dp, and Rbf1 genes all contribute to replication control in ovarian follicle cells, which become 16C polyploid and subsequently undergo chorion gene amplification late in oogenesis. Mutation of E2f2, Dp, or Rbf1 causes ectopic DNA replication throughout the follicle cell genome during gene amplification cycles. Here we show by both reverse transcription-PCR and DNA microarray analysis that the transcripts of prereplication complex (pre-RC) genes are elevated compared to the wild type in E2f2, Dp, and Rbf1 mutant follicle cells. For some genes the magnitude of this transcriptional derepression is greater in Rbf1 than in E2f2 mutants. These differences correlate with differences in the magnitude of the replication defects in follicle cells, which attain an inappropriate 32C DNA content in both Rbf1 and Dp mutants but not in E2f2 mutants. The ectopic genomic replication of E2f2 mutant follicle cells can be suppressed by reducing the Orc2, Orc5, or Mcm2 gene dose by half, indicating that small changes in pre-RC gene expression can affect DNA synthesis in these cells. We conclude that RBF1 forms complexes with both E2F1/DP and E2F2/DP that cooperate to repress the expression of pre-RC genes, which helps confine DNA synthesis to sites of gene amplification. In contrast, E2F1 and E2F2 repressors function redundantly for some genes in the embryo. Thus, the relative functional contributions of E2F1 and E2F2 to gene expression and cell cycle control depends on the developmental context.