RESUMO
The bantam gene encodes a vital microRNA and has a complex expression pattern in various tissues at different stages of Drosophila development. This microRNA is involved in the control of normal development of the ocular and wing imaginal discs, the central nervous system, and also in maintaining the undifferentiated state of stem cells in the ovaries of adult females. At the cellular level, bantam stimulates cell proliferation and prevents apoptosis. The bantam gene is a target of several conserved signaling cascades, in particular, Hippo. At the moment, at least ten proteins are known to directly regulate the expression of this gene in different tissues of Drosophila. In this study, we found that the bantam regulatory region contains motifs characteristic of binding sites for DREF, a transcription factor that regulates the expression of Hippo cascade genes. Using transgenic lines containing a full-length bantam lethality-rescuing deletion fragment and a fragment with a disrupted DREF binding site, we show that these motifs are functionally significant because their disruption at the bantam locus reduces expression levels in the larvae and ovaries of homozygous flies, which correlates with reduced vitality and fertility. The effect of DREF binding to the promoter region of the bantam gene on its expression level suggests an additional level of complexity in the regulation of expression of this microRNA. A decrease in the number of eggs laid and a shortening of the reproductive period in females when the DREF binding site in the regulatory region of the bantam gene is disrupted suggests that, through bantam, DREF is also involved in the regulation of Drosophila oogenesis.
RESUMO
Chromatin insulator proteins are one of the major components that determine the domain organization of the genome. According to the latest data, they can mark the boundaries of topological domains and prevent the spread of silent chromatin to adjacent areas. One approach to the analysis of the actions of these proteins is to use the ectopic involvement in the UAS>DBD(GAL4). The method allows to evaluate the effect of selected protein in chromatin organization, to establish its association with other insulator proteins and influence on the processes of transcription and replication. and influence the processes of transcription and replication. In this study, we have developed and tested the functionality of the system components in ectopic tethering of the Chromator (Chriz) to the region of intercalary heterochromatin 10A1-2. Preliminary data have been obtained showing that ectopically tethered Chromator to the band 10A1-2 can induce a partial decompactization of the band chromatin. Further use of this experimental model provides the opportunity to investigate the effect of insulator proteins on the chromatin structure.
Assuntos
Montagem e Desmontagem da Cromatina , Proteínas de Drosophila/metabolismo , Elementos Isolantes , Proteínas Associadas à Matriz Nuclear/metabolismo , Cromossomos Politênicos/metabolismo , Fatores de Transcrição/metabolismo , Animais , Proteínas de Drosophila/genética , Drosophila melanogaster , Proteínas Associadas à Matriz Nuclear/genética , Cromossomos Politênicos/genética , Fatores de Transcrição/genéticaRESUMO
For the first time we used a homologous recombination method to obtain complete and precise deletion of Drosophila dRNaseZ gene. In the founder line of flies in which the RNaseZ sequence was replaced by attP site, the full-length sequence of the gene was reintegrated, and its functionality was shown. This approach will allow us to generate further gene mutations in different domains of dRNaseZ protein and discover a broad spectrum and uncover functions outside of tRNA processing.
Assuntos
Proteínas de Drosophila/genética , Endorribonucleases/genética , Técnicas Genéticas , Recombinação Homóloga , Deleção de Sequência , Animais , Animais Geneticamente Modificados , Sítios de Ligação Microbiológicos , Northern Blotting , Western Blotting , Sistema Nervoso Central/metabolismo , Cromossomos Artificiais Bacterianos , Drosophila , Proteínas de Drosophila/metabolismo , Endorribonucleases/metabolismo , Feminino , Gônadas/metabolismo , Discos Imaginais/metabolismo , Larva , Masculino , Proteínas Mutantes Quiméricas/metabolismo , Reação em Cadeia da Polimerase , RNA/metabolismo , RNA Mitocondrial , RNA de Transferência/metabolismoRESUMO
Methods of physical DNA mapping and direct visualization of replication and transcription in specific regions of genome play crucial role in the researches of structural and functional organization of eukaryotic genomes. Since DNA strands in the cells are organized into high-fold structure and present as highly compacted chromosomes, the majority of these methods have lower resolution at chromosomal level. One of the approaches to enhance the resolution and mapping accuracy is the method of molecular combing. The method is based on the process of stretching and alignment of DNA molecules that are covalently attached with one of the ends to the cover glass surface. In this article we describe the major methodological steps of molecular combing and their adaptation for researches of DNA replication parameters in polyploidy and diploid tissues of Drosophyla larvae.
Assuntos
Mapeamento Cromossômico/métodos , Replicação do DNA , DNA/química , Drosophila melanogaster/genética , Larva/citologia , Animais , Mapeamento Cromossômico/instrumentação , Reagentes de Ligações Cruzadas/química , DNA/genética , Diploide , Drosophila melanogaster/citologia , Drosophila melanogaster/metabolismo , Vidro/química , Hibridização in Situ Fluorescente , Larva/genética , Larva/metabolismo , Sondas Moleculares/química , Desnaturação de Ácido Nucleico , Técnicas de Cultura de Órgãos , Reação em Cadeia da Polimerase , Poliploidia , Origem de Replicação , Sefarose/químicaRESUMO
The functional organization of particular chromosome regions is tightly associated with their function in eukaryotic cells. Details of this association are among the most topical problems of modem genetics. The paper characterizes the results of recent research of the specifics of the genetic organization and chromatin decondensation in interbands of Drosophila polytene chromosomes. Data on functional heterogeneity of interbands are considered. Experimental findings point to a lack of correlation between the decondensed chromatin state and the observed transcription level in particular interbands. The DNA sequences responsible for the interband formation are principally identifiable via site-specific homologous FRT/FLP recombination between two P transposons contained in chromosomes. The results allow a search for particular protein factors that are involved in the decondensed state of interbands and structural and functional differentiation of polytene chromosomes.
Assuntos
Cromatina/genética , Cromossomos Politênicos/genética , Animais , Cromatina/metabolismo , Bandeamento Cromossômico , Mapeamento Cromossômico , Elementos de DNA Transponíveis/fisiologia , Drosophila melanogaster , Cromossomos Politênicos/metabolismo , Recombinação Genética/fisiologia , Transcrição Gênica/fisiologiaRESUMO
Polymorphic sites in the genes encoding monoamine oxidase A (MAO-A), serotonin transporter (hSERT) and 5-HT2A receptor were typed in Khant and Komi ethnic groups with the purpose of revealing possible interpopulation differences in genotype and allele frequencies. No statistically significant differences in the hSERT and 5-HT2A gene frequencies were detected. At the same time, the populations examined had statistically significantly different MAO-A genotype and allele frequencies. These results obtained indicate the prevalence of the site gain alleles of the EcoRV and Fnu4HI RFLP loci at the MAO-A gene in Komis and the of the corresponding site loss alleles in Khants.
