Repetitive DNAs constitute a significant part of the genomes and are used as cytogenetic markers for the understanding of chromosome evolution. Bees of the genus Melipona presents conserved chromosome number but high variability concerning heterochromatin distribution, dividing the genus into two groups: (i) species with low quantity of heterochromatin and (ii) species with high amount of heterochromatin. Here aiming the understanding of repetitive DNAs chromosomal organization in Melipona we mapped through fluorescence in situ hybridization (FISH) some repetitive DNAs in M. scutellaris. For this, FISH were performed with the amplification products of the 18S rDNA, U2 snDNA and DOP-PCR, as well as the product of the C0t fraction and different microsatellites. The mitotic chromosomes used in FISH were obtained by squashing the brain ganglia, and in this material, being performed also the C-band and CMA/DAPI. The distribution of the C0t fraction in other species of Melipona was verified with membrane hybridization. Chromosome number of n = 9 and 2n = 18 (male and female, respectively) was confirmed and a large amount of heterochromatin was observed along the chromosomes, except at the terminals that were characterized as euchromatic and only rich in different microsatellites. The heterochromatic regions were observed with richness in A + T and C + G. The FISH experiments showed signals from the 18S rDNA and U2 snDNA probes on distinct chromosomes. This demonstrates that these multigenic families followed different evolutionary paths in the chromosome evolution of the species. The DOP-PCR and C0t fraction showed signs of hybridization in the heterochromatic regions, and the C0t fraction of M. scutellaris was also observed in the genomes of M. rufiventris, M. rufiventris rufiventris and M. seminigra, evidencing the dispersion of similar repetitive elements in genomes of these three species. Thus, it was possible to characterize the karyotype of M. scutellaris with different cytogenetic markers. In addition, the results with membrane hybridization reinforce the hypothesis of the repetitive DNA sharing among nearby species, suggesting that there are different repetitive elements present in the genome of the genus Melipona.
B chromosomes are dispensable elements that occur in addition to normal chromosomes (A) and exhibit non-Mendelian inheritance. These supernumerary chromosomes occur in about 15% of the eukaryotes, being composed basically of repetitive and heterochromatic DNA more specifically the multigenic families, transposition elements and satellite DNAs (satDNAs). Several studies using animals and plants have been performed in order to obtain new information about the evolution and behavior of satDNAs and besides evidences for transcription have been emerged, suggesting functional roles. In the present work we checked the transcription of five satDNAs previously isolated from grasshopper Abracris flavolineata genome, a species with B chromosome. RNA was extracted from two tissues (saltatory leg and testis) from two individuals 0B and one 1B and the cDNA was obtained to test satDNA transcription. In both tissues all satDNAs were transcribed, but they revealed variability depending of the individual for two of them. For satDNAs 204 and 220 it was noticed transcription in salutatory legs of only one individual 0B, as for satDNA 204 in which transcription in testis was restrict for one individual 0B. The other satDNAs were transcribed in all individuals. The observation of transcription in 1B individuals indicate that this chromosome is not supressing satDNAs. Finally, transcription of all satDNAs in a somatic and germinative tissues suggests functional role of these sequences independent of the cell line. The next steps are to test the differences in transcript quantity for distict tissues of 0B and 1B individuals, trying to understand the influence of B chromosome presence in satDNA transcrition and to check the possible occurrence of secundary functional structures for the satDNAs sequences.
B chromosomes are additional elements in a normal karyotype that has been considered frequently as parasitic and dispensable. These elements do not recombine with A chromosomes, could be accumulated and present non-mendelian patterns of inheritance. In most of the cases they comprise an enrichment of heterochromatic repetitive DNA, such as Satellite DNAs (satDNA). To contribute to understanding about origin, composition and evolutionary patterns of B chromosome in the grasshopper Abracris flavolineata, in this work we characterized through Illumina sequencing and graph-based clustering the most abundant tandem repeats of A. flavolineata, karyotype 2n = 22,X0+B?.The analyses revealed the occurrence of ten families of satDNAs comprising about 6.279% of the male genome harboring one B chromosome. For the X chromosome and autosomes, distinct patterns of hybridization was observed between the satDNAs showing variability among them, like scattered blocks on euchromatic areas or specific ones on heterochromatic regions of the chromosomes. Five of the satDNAs recovered were distributed on centromeric and both terminal regions of the B chromosome, reinforcing the hypothesis of B chromosome origin through isochromosome formation. Two satDNA families were located only in centromere, while three families were not present in B chromosome. CL220 satDNA was the unique family shared between the B chromosome and only one A element, the chromosome 1, supporting strongly the hypothesis of its origin from this autosomal pair, also noticed previously by U2 snDNA mapping. In addition, this analysis provides new information regarding the composition and evolution of B chromosome of A. flavolineata. Furthermore, the non-spreading and specific location of satDNAs and other repetitive sequences restrict to telomeres and centromere reinforces the possible idea that this B chromosome could harbor unknown euchromatic sequences along chromosomal arms and do not present intense amplification of heterochromatic ones, as have been commonly described for other species.
Euchroma é um gênero considerado monotípico para E. gigantea, com descrição de quatro subespécies e um alto polimorfismo cromossômico, que decorre de rearranjos que podem estar relacionados a diferentes tipos de DNA repetitivo. Com o objetivo de investigar o papel de sequências repetitivas na diversificação cariotípica desta espécie, duas sequências de DNA satélite (DNAsat) foram caracterizadas, analisadas filogeneticamente entre diferentes cariótipos e mapeadas através de hibridização in situ fluorescente. Os espécimes analisados foram coletados em Belém (PA), Ribeirão Preto (SP), Brasília (DF), Recife (PE) e Maceió (AL). As sequências foram isoladas a partir de sequenciamento Illumina de baixa cobertura do genoma de um espécime com 2n=34(PE) e caracterizadas através de ferramentas de bioinformática. Adicionalmente, a sequência de DNAr 18S foi mapeada. Os espécimes analisados exibiram cariótipos com 2n=22(DF), 26(SP), 32(PA), 34(PE) e 35(AL), todos com mecanismo sexual múltiplo formado por cinco, seis ou oito cromossomos em cadeia. Apenas os espécimes com 2n=22 não apresentaram cromossomos B, nos demais houve variação de três a 21. As sequências EgiSat1-172 e EgiSat2-202 apresentaram riqueza em pares de base A+T de 40,1% e 49,5%, respectivamente e proporções relativas no genoma de 0,299% para EgiSat1 e 0,09% para EgiSat2. A análise filogenética evidenciou um alto grau de identidade das sequências, o que pode ser decorrente do surgimento dessas sequências em um ancestral comum. A partir do mapeamento foram observados de dois a quatro sítios de EgiSat1 e EgiSat2 co-localizados na região pericentromérica de cromossomos autossomos nos cariótipos com 2n=22, 26, 32, 34 ou em autossomos e no cromossomo X3 no cariótipo com 2n=35. Estes padrões de distribuição foram similares aos do DNAr 18S. EgiSat1 e EgiSat2 não apresentaram similaridades com os genes do DNAr 45S, possivelmente estas sequências estão relacionadas a sequências próximas, como o espaçador intergênico. A variabilidade na distribuição de sítios de EgiSat1 e EgiSat2 pode decorrer de recombinação ectópica, reinserção de DNA circular extracromossômico ou atividade de transposons. Visando entender melhor a relação entre as sequências EgiSat1, EgiSat2 e o DNAr, sequências relacionadas estão sendo analisadas. Outros marcadores citogenéticos estão sendo utilizados a fim de elucidar a evolução cromossômica de E. gigantea.Financiadores: CAPES, CNPq, FACEPE, FAPESP
Satellite DNA (satDNA) is a class of non-coding repetitive DNA abundant in most eukaryotic genomes. Mostly satDNAs constitute clustered arrays of tandemly repeated sequences located in the gene-poor heterochromatin of centromeres and telomeres. Moreover in some cases, they are also dispersed in eu/heterochromatin of sex chromosomes and as single or short arrays nearby protein-coding genes within euchromatin. Here we take advantages of DNA- and RNA-seq data from cricket’s species in order to perform a comparative analysis of content and expression of satDNAs. By graph-based clustering analysis of DNA-seq reads using RepeatExplorer software, dotplots analysis and FISH mapping we found that ~ 4% of the G. assimilis genome is represented by 13 A+T-rich satDNAs consisting of 11 well-defined families mainly located in the heterochromatic areas of chromosomes, and some of them able to form high-order repeats (HORs). In order to determine transcription profiles the raw RNA-seq reads from different tissues library of Gryllus species were mapped to each of the G. assimilis satDNAs using Bowtie2 and the method FPKM (fragments per kilo-base of transcript per million mapped reads) was used. The in silico transcriptional analysis of RNA-seq reads in G. assimilis, G. bimaculatus, G, firmus and G. rubens showed that some satDNAs are conserved in Gryllus species but differentially expressed in distinct tissues, sexes and besides tissue- and species-specific. In concordances with the transcriptional activity we found that G. assimilis satDNAs are also capable to adopt RNA secondary structures with well-defined helices ranging from 2 bp to 7 bp lengths. The folding possibility forming secondary structure helps to the satDNA dispersion along the genome by rolling-circle replication mechanism, in which circular monomer result from secondary structure RNA processing into linear monomers and subsequently circularization by a host-specific RNA ligase. The conservation of expression for different satDNAs in Gryllus species suggests a functional role for these sequences, as observed in other insects. Our data suggests functional roles of satDNAs for sexual differentiation at the chromatin levels, heterochromatin formation and centromeric function.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.