RESUMEN
We developed a procedure for locating genes on Drosophila melanogaster polytene chromosomes and described three types of chromosome structures (gray bands, black bands, and interbands), which differed markedly in morphological and genetic properties. This was reached through the use of our original methods of molecular and genetic analysis, electron microscopy, and bioinformatics data processing. Analysis of the genome-wide distribution of these properties led us to a bioinformatics model of the Drosophila genome organization, in which the genome was divided into two groups of genes. One was constituted by 65, in which the genome was divided into two groups, 62 genes that are expressed in most cell types during life cycle and perform basic cellular functions (the so-called "housekeeping genes"). The other one was made up of 3162 genes that are expressed only at particular stages of development ("developmental genes"). These two groups of genes are so different that we may state that the genome has two types of genetic organization. Different are the timings of their expression, chromatin packaging levels, the composition of activating and deactivating proteins, the sizes of these genes, the lengths of their introns, the organization of the promoter regions of the genes, the locations of origin recognition complexes (ORCs), and DNA replication timings.
Asunto(s)
Drosophila , Genes Esenciales , Animales , Drosophila/genética , Drosophila melanogaster/genética , Cromatina , IntronesRESUMEN
BACKGROUND: Despite many efforts, little is known about distribution and interactions of chromatin proteins which contribute to the specificity of chromomeric organization of interphase chromosomes. To address this issue, we used publicly available datasets from several recent Drosophila genome-wide mapping and annotation projects, in particular, those from modENCODE project, and compared molecular organization of 13 interband regions which were accurately mapped previously. RESULTS: Here we demonstrate that in interphase chromosomes of Drosophila cell lines, the interband regions are enriched for a specific set of proteins generally characteristic of the "open" chromatin (RNA polymerase II, CHRIZ (CHRO), BEAF-32, BRE1, dMI-2, GAF, NURF301, WDS and TRX). These regions also display reduced nucleosome density, histone H1 depletion and pronounced enrichment for ORC2, a pre-replication complex component. Within the 13 interband regions analyzed, most were around 3-4 kb long, particularly those where many of said protein features were present. We estimate there are about 3500 regions with similar properties in chromosomes of D. melanogaster cell lines, which fits quite well the number of cytologically observed interbands in salivary gland polytene chromosomes. CONCLUSIONS: Our observations suggest strikingly similar organization of interband chromatin in polytene chromosomes and in chromosomes from cell lines thereby reflecting the existence of a universal principle of interphase chromosome organization.
Asunto(s)
Cromatina/genética , Proteínas Cromosómicas no Histona/genética , Drosophila melanogaster/genética , Cromosomas Politénicos/genética , Animales , Histonas/genética , InterfaseRESUMEN
Salivary gland polytene chromosomes demonstrate banding pattern, genetic meaning of which is an enigma for decades. Till now it is not known how to mark the band/interband borders on physical map of DNA and structures of polytene chromosomes are not characterized in molecular and genetic terms. It is not known either similar banding pattern exists in chromosomes of regular diploid mitotically dividing nonpolytene cells. Using the newly developed approach permitting to identify the interband material and localization data of interband-specific proteins from modENCODE and other genome-wide projects, we identify physical limits of bands and interbands in small cytological region 9F13-10B3 of the X chromosome in D. melanogaster, as well as characterize their general molecular features. Our results suggests that the polytene and interphase cell line chromosomes have practically the same patterns of bands and interbands reflecting, probably, the basic principle of interphase chromosome organization. Two types of bands have been described in chromosomes, early and late-replicating, which differ in many aspects of their protein and genetic content. As appeared, origin recognition complexes are located almost totally in the interbands of chromosomes.
Asunto(s)
Drosophila melanogaster/metabolismo , Cromosomas Politénicos/metabolismo , Animales , ADN/metabolismo , Sondas de ADN/metabolismo , Bases de Datos Genéticas , Drosophila melanogaster/ultraestructura , Genoma de los Insectos/genética , Hibridación Fluorescente in Situ , Proteínas de Insectos/metabolismo , Mapeo Físico de Cromosoma , Cromosomas Politénicos/ultraestructuraRESUMEN
We studied whether interbands can be ectopically formed in Drosophila melanogaster polytene chromosomes. For comparative purposes, two types of P-element constructs were used. The first type was represented by P-element based insertions into compact bands. Sequences of these insertions or adjacent genomic sequences could be activated ectopically either by GAL4 or by dosage compensation machinery. In the second type, the DNA from transcriptionally silent interbands was positioned between the FRT sites, and was flanked by DNA sequences of genes that were also inactive in salivary glands. Electron microscopy analysis of salivary gland polytene chromosomes demonstrated that both types of constructs formed distinct, yet morphologically similar interbands. Notably, the second class of transposon insertions appeared in polytene chromosomes as two bands separated by one interband. Excision of interband material from such insertions resulted in fusion of newly appeared bands into a single band. We were able to confirm by molecular means that the DNA sequences in integrated constructs were intact, that chromatin organization of this DNA mimicked that of native interbands, and that it was accurately excised from the constructs by FLP. Thus, we demonstrate that transfer of interband DNA into a silent genetic environment does not compromise interband formation. Our results do not support the idea of the existence of distinct cytogenetic "band + interband" units, furthermore, they suggest the autonomy of the decompacted state of interbands.
Asunto(s)
Bandeo Cromosómico , Cromosomas/genética , Drosophila melanogaster/genética , Animales , Animales Modificados Genéticamente , Cromatina/genética , Cromosomas/ultraestructura , ADN/análisis , ADN/genética , Elementos Transponibles de ADN , ADN Mitocondrial , Diploidia , Evolución Molecular , Genes de Insecto , Heterocromatina , Hibridación in Situ , Masculino , Filogenia , Glándulas Salivales/citología , Análisis de Secuencia de ADNRESUMEN
The evolution of 2 tandemly repeated sequences Spelt1 and Spelt52 was studied in Triticum species representing 2 evolutionary lineages of wheat and in Aegilops sect. Sitopsis, putative donors of their B/G genomes. Using fluorescence in situ hybridization we observed considerable polymorphisms in the hybridization patterns of Spelt1 and Spelt52 repeats between and within Triticum and Aegilops species. Between 2 and 28 subtelomeric sites of Spelt1 probe were detected in Ae. speltoidies, depending on accession. From 8 to 12 Spelt1 subtelomeric sites were observed in species of Timopheevi group (GAt genome), whereas the number of signals in emmer/aestivum accessions was significantly less (from 0 to 6). Hybridization patterns of Spelt52 in Ae. speltoides, Ae. longissima, and Ae. sharonensis were species specific. Subtelomeric sites of Spelt52 repeat were detected only in T. araraticum (T. timopheevii), and their number and chromosomal location varied between accessions. Superimposing copy number data onto our phylogenetic scheme constructed from RAPD data suggests 2 major independent amplifications of Spelt52 and 1 of Spelt1 repeats in Aegilops divergence. It is likely that the Spelt1 amplification took place in the ancient Ae. speltoides before the divergence of polyploid wheats. The Spelt52 repeat was probably amplified in the lineage of Ae. speltoides prior to divergence of the allopolyploid T. timopheevii but after the divergence of T. durum. In a separate amplification event, Spelt52 copy number expanded in the common ancestor of Ae. longissima and Ae. sharonensis.