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Current Genomics Apr 2018This mini-review is devoted to the problem genetic meaning of main polytene chromosome structures - bands and interbands. Generally, densely packed chromatin forms black... (Review)
Review
This mini-review is devoted to the problem genetic meaning of main polytene chromosome structures - bands and interbands. Generally, densely packed chromatin forms black bands, moderately condensed regions form grey loose bands, whereas decondensed regions of the genome appear as interbands. Recent progress in the annotation of the Drosophila genome and epigenome has made it possible to compare the banding pattern and the structural organization of genes, as well as their activity. This was greatly aided by our ability to establish the borders of bands and interbands on the physical map, which allowed to perform comprehensive side-by-side comparisons of cytology, genetic and epigenetic maps and to uncover the association between the morphological structures and the functional domains of the genome. These studies largely conclude that interbands 5'-ends of housekeeping genes that are active across all cell types. Interbands are enriched with proteins involved in transcription and nucleosome remodeling, as well as with active histone modifications. Notably, most of the replication origins map to interband regions. As for grey loose bands adjacent to interbands, they typically host the bodies of house-keeping genes. Thus, the bipartite structure composed of an interband and an adjacent grey band functions as a standalone genetic unit. Finally, black bands harbor tissue-specific genes with narrow temporal and tissue expression profiles. Thus, the uniform and permanent activity of interbands combined with the inactivity of genes in bands forms the basis of the universal banding pattern observed in various Drosophila tissues.
PubMed: 29606905
DOI: 10.2174/1389202918666171016123830 -
Genes Feb 2020underreplicate the DNA of thoracic nuclei, stalling during S phase at a point that is proportional to the total genome size in each species. In polytene tissues, such...
underreplicate the DNA of thoracic nuclei, stalling during S phase at a point that is proportional to the total genome size in each species. In polytene tissues, such as the salivary glands, all of the nuclei initiate multiple rounds of DNA synthesis and underreplicate. Yet, only half of the nuclei isolated from the thorax stall; the other half do not initiate S phase. Our question was, why half? To address this question, we use flow cytometry to compare underreplication phenotypes between thoracic tissues. When individual thoracic tissues are dissected and the proportion of stalled DNA synthesis is scored in each tissue type, we find that underreplication occurs in the indirect flight muscle, with the majority of underreplicated nuclei in the dorsal longitudinal muscles (DLM). Half of the DNA in the DLM nuclei stall at S phase between the unreplicated G0 and fully replicated G1. The dorsal ventral flight muscle provides the other source of underreplication, and yet, there, the replication stall point is earlier (less DNA replicated), and the endocycle is initiated. The differences in underreplication and ploidy in the indirect flight muscles provide a new tool to study heterochromatin, underreplication and endocycle control.
Topics: Animals; Cell Nucleus; DNA; DNA Replication; Drosophila melanogaster; Flight, Animal; Flow Cytometry; G1 Phase; Muscle, Skeletal; Polytene Chromosomes; Resting Phase, Cell Cycle; S Phase; Salivary Glands; Thorax
PubMed: 32111003
DOI: 10.3390/genes11030246 -
Malaria Journal Mar 2024Anopheles vagus (subgenus Cellia) has been identified as a vector for malaria, filariasis, and Japanese encephalitis in Asia. Sporozoites of Plasmodium falciparum and... (Review)
Review
BACKGROUND
Anopheles vagus (subgenus Cellia) has been identified as a vector for malaria, filariasis, and Japanese encephalitis in Asia. Sporozoites of Plasmodium falciparum and Plasmodium vivax have been found in this zoophilic mosquito in Asia and Indonesia. This study systematically reviews publications regarding An. vagus species, variation, bio-ecology, and malaria transmission in various localities in Asia, especially Indonesia, to determine whether the current data support An. vagus as a species complex.
METHODS
The databases Pubmed, Scopus, Europe PMC, and Proquest were searched to identify information regarding the morphology, karyotypes, polytene chromosome, cross-mating, ecology, and molecular identification of An. vagus was then evaluated to determine whether there were possible species complexes.
RESULTS
Of the 1326 articles identified, 15 studies were considered for synthesis. The Anopheles spp. samples for this study came from Asia. Eleven studies used morphology to identify An. vagus, with singular studies using each of karyotype identification, chromosomal polytene identification, and cross-breeding experiments. Ten studies used molecular techniques to identify Anopheles spp., including An. vagus. Most studies discovered morphological variations of An. vagus either in the same or different areas and ecological settings. In this review, the members of An. vagus sensu lato grouped based on morphology (An. vagus, An. vagus vagus, An. vagus limosus, and An. limosus), karyotyping (form A and B), and molecular (An. vagus genotype A and B, An. vagus AN4 and AN5). Genetic analysis revealed a high conservation of the ITS2 fragment among members except for the An. vagus genotype B, which was, in fact, Anopheles sundaicus. This review also identified that An. vagus limosus and An. vagus vagus were nearly identical to the ITS2 sequence.
CONCLUSION
Literature review studies revealed that An. vagus is conspecific despite the distinct morphological characteristic of An. vagus and An. limosus. Further information using another barcoding tool, such as mitochondrial COI and ND6 and experimental cross-mating between the An. vagus and An. limosus may provide additional evidence for the status of An. vagus as a species complex.
Topics: Animals; Phylogeny; Anopheles; Genotype; Mosquito Vectors; Malaria
PubMed: 38539155
DOI: 10.1186/s12936-024-04888-0 -
Genes Nov 2019In the ciliate somatic macronuclei differentiate from germline micronuclei during sexual reproduction, accompanied by developmental sequence reduction. Concomitantly,... (Review)
Review
In the ciliate somatic macronuclei differentiate from germline micronuclei during sexual reproduction, accompanied by developmental sequence reduction. Concomitantly, over 95% of micronuclear sequences adopt a heterochromatin structure characterized by the histone variant H3.4 and H3K27me3. RNAi-related genes and histone variants dominate the list of developmentally expressed genes. Simultaneously, 27nt-ncRNAs that match sequences retained in new macronuclei are synthesized and bound by PIWI1. Recently, we proposed a mechanistic model for 'RNA-induced DNA replication interference' (RIRI): during polytene chromosome formation PIWI1/27nt-RNA-complexes target macronucleus-destined sequences (MDS) by base-pairing and temporarily cause locally stalled replication. At polytene chromosomal segments with ongoing replication, H3.4K27me3-nucleosomes become selectively deposited, thus dictating the prospective heterochromatin structure of these areas. Consequently, these micronucleus-specific sequences become degraded, whereas 27nt-RNA-covered sites remain protected. However, the biogenesis of the 27nt-RNAs remains unclear. It was proposed earlier that in stichotrichous ciliates 27nt-RNA precursors could derive from telomere-primed bidirectional transcription of nanochromosomes and subsequent Dicer-like (DCL) activity. As a minimalistic explanation, we propose here that the 27nt-RNA precursor could rather be mRNA or pre-mRNA and that the transition of coding RNA from parental macronuclei to non-coding RNAs, which act in premature developing macronuclei, could involve RNA-dependent RNA polymerase (RDRP) activity creating dsRNA intermediates prior to a DCL-dependent pathway. Interestingly, by such mechanism the partition of a parental somatic genome and possibly also the specific nanochromosome copy numbers could be vertically transmitted to the differentiating nuclei of the offspring.
Topics: Ciliophora; DNA Replication; Gene Expression Regulation, Developmental; Genome, Protozoan; Histones; Micronucleus, Germline; Nucleosomes; RNA Interference; RNA Precursors; RNA, Messenger; RNA, Small Nuclear; Telomere
PubMed: 31752243
DOI: 10.3390/genes10110940 -
Experimental Biology and Medicine... Nov 2019Repairing DNA double-strand breaks is particularly challenging in pericentromeric heterochromatin, where the abundance of repeated sequences exacerbates the risk of...
UNLABELLED
Repairing DNA double-strand breaks is particularly challenging in pericentromeric heterochromatin, where the abundance of repeated sequences exacerbates the risk of ectopic recombination. In Kc cells, accurate homologous recombination repair of heterochromatic double-strand breaks relies on the relocalization of repair sites to the nuclear periphery before Rad51 recruitment and strand invasion. This movement is driven by Arp2/3-dependent nuclear actin filaments and myosins’ ability to walk along them. Conserved mechanisms enable the relocalization of heterochromatic repair sites in mouse cells, and defects in these pathways lead to massive ectopic recombination in heterochromatin and chromosome rearrangements. In polytene chromosomes, extensive DNA movement is blocked by a stiff structure of chromosome bundles. Repair pathways in this context are poorly characterized, and whether heterochromatic double-strand breaks relocalize in these cells is unknown. Here, we show that damage in heterochromatin results in relaxation of the heterochromatic chromocenter, consistent with a dynamic response. Arp2/3, the Arp2/3 activator Scar, and the myosin activator Unc45, are required for heterochromatin stability in polytene cells, suggesting that relocalization enables heterochromatin repair also in this tissue. Together, these studies reveal critical roles for actin polymerization and myosin motors in heterochromatin repair and genome stability across different organisms and tissue types.
IMPACT STATEMENT
Heterochromatin relies on dedicated pathways for ‘safe’ recombinational repair. In mouse and fly cultured cells, DNA double-strand break repair requires the movement of damaged sites away from the heterochromatin ‘domain’ nuclear actin filaments and myosins. Here, we explore the importance of these pathways in salivary gland cells, which feature a stiff bundle of endoreduplicated polytene chromosomes. Repair pathways in polytene chromosomes are largely obscure and how nuclear dynamics operate in this context is unknown. We show that heterochromatin relaxes in response to damage, and relocalization pathways are necessary to prevent abnormal repair and promote the stability of heterochromatic sequences. These results deepen our understanding of DNA damage response mechanisms in polytene chromosomes, revealing unexpected dynamics. It also provides a first understanding of nuclear dynamics responding to replication damage and rDNA breaks, providing a new understanding of the importance of nuclear architecture in genome stability. We expect these discoveries will shed light on tumorigenic processes, including therapy-induced cancer relapses.
Topics: Actin Cytoskeleton; Actin-Related Protein 2-3 Complex; Animals; DNA Breaks, Double-Stranded; DNA Repair; Drosophila Proteins; Drosophila melanogaster; Genomic Instability; Heterochromatin; Molecular Chaperones; Myosins; Polytene Chromosomes
PubMed: 31364400
DOI: 10.1177/1535370219862282 -
PLoS Genetics Oct 2022Meiosis in males of higher dipterans is achiasmate. In their spermatocytes, pairing of homologs into bivalent chromosomes does not include synaptonemal complex and...
Meiosis in males of higher dipterans is achiasmate. In their spermatocytes, pairing of homologs into bivalent chromosomes does not include synaptonemal complex and crossover formation. While crossovers preserve homolog conjunction until anaphase I during canonical meiosis, an alternative system is used in dipteran males. Mutant screening in Drosophila melanogaster has identified teflon (tef) as being required specifically for alternative homolog conjunction (AHC) of autosomal bivalents. The additional known AHC genes, snm, uno and mnm, are needed for the conjunction of autosomal homologs and of sex chromosomes. Here, we have analyzed the pattern of TEF protein expression. TEF is present in early spermatocytes but cannot be detected on bivalents at the onset of the first meiotic division, in contrast to SNM, UNO and MNM (SUM). TEF binds to polytene chromosomes in larval salivary glands, recruits MNM by direct interaction and thereby, indirectly, also SNM and UNO. However, chromosomal SUM association is not entirely dependent on TEF, and residual autosome conjunction occurs in tef null mutant spermatocytes. The higher tef requirement for autosomal conjunction is likely linked to the quantitative difference in the amount of SUM protein that provides conjunction of autosomes and sex chromosomes, respectively. During normal meiosis, SUM proteins are far more abundant on sex chromosomes compared to autosomes. Beyond promoting SUM recruitment, TEF has a stabilizing effect on SUM proteins. Increased SUM causes excess conjunction and consequential chromosome missegregation during meiosis I after co-overexpression. Similarly, expression of SUM without TEF, and even more potently with TEF, interferes with chromosome segregation during anaphase of mitotic divisions in somatic cells, suggesting that the known AHC proteins are sufficient for establishment of ectopic chromosome conjunction. Overall, our findings suggest that TEF promotes alternative homolog conjunction during male meiosis without being part of the final physical linkage between chromosomes.
Topics: Animals; Male; Drosophila melanogaster; Drosophila; Drosophila Proteins; Polytetrafluoroethylene; Chromosome Segregation; Meiosis; Sex Chromosomes; Chromosome Pairing
PubMed: 36251690
DOI: 10.1371/journal.pgen.1010469 -
Cells Feb 2020Spatial organization of chromosome territories and interactions between interphase chromosomes themselves, as well as with the nuclear periphery, play important roles in...
Spatial organization of chromosome territories and interactions between interphase chromosomes themselves, as well as with the nuclear periphery, play important roles in epigenetic regulation of the genome function. However, the interplay between inter-chromosomal contacts and chromosome-nuclear envelope attachments in an organism's development is not well-understood. To address this question, we conducted microscopic analyses of the three-dimensional chromosome organization in malaria mosquitoes. We employed multi-colored oligonucleotide painting probes, spaced 1 Mb apart along the euchromatin, to quantitatively study chromosome territories in larval salivary gland cells and adult ovarian nurse cells of , , and . We found that the X chromosome territory has a significantly smaller volume and is more compact than the autosomal arm territories. The number of inter-chromosomal, and the percentage of the chromosome-nuclear envelope, contacts were conserved among the species within the same cell type. However, the percentage of chromosome regions located at the nuclear periphery was typically higher, while the number of inter-chromosomal contacts was lower, in salivary gland cells than in ovarian nurse cells. The inverse correlation was considerably stronger for the autosomes. Consistent with previous theoretical arguments, our data indicate that, at the genome-wide level, there is an inverse relationship between chromosome-nuclear envelope attachments and chromosome-chromosome interactions, which is a key feature of the cell type-specific nuclear architecture.
Topics: Animals; Anopheles; Female; Germ Cells; Malaria; Nuclear Envelope; Ovary; Polytene Chromosomes; Salivary Glands; X Chromosome
PubMed: 32024176
DOI: 10.3390/cells9020339 -
Cells Nov 2018Polynucleotide chains obeying Watson-Crick pairing are apt to form non-canonical complexes such as triple-helical nucleic acids. From early characterization in vitro,...
Polynucleotide chains obeying Watson-Crick pairing are apt to form non-canonical complexes such as triple-helical nucleic acids. From early characterization in vitro, their occurrence in vivo has been strengthened by increasing evidence, although most remain circumstantial particularly for triplex DNA. Here, different approaches were employed to specify triple-stranded DNA sequences in the chromosomes. Antibodies to triplex nucleic acids, previously characterized, bind to centromeric regions of mitotic chromosomes and also to the polytene section 59E of mutant strains carrying the allele, indicating that AAGAG tandem satellite repeats are triplex-forming sequences. The satellite probe hybridized to AAGAG-containing regions omitting chromosomal DNA denaturation, as expected, for the intra-molecular triplex DNA formation model in which single-stranded DNA coexists with triplexes. In addition, Thiazole Orange, previously described as capable of reproducing results obtained by antibodies to triple-helical DNA, binds to AAGAG repeats in situ thus validating both detection methods. Unusual phenotype and nuclear structure exhibited by correlate with the non-canonical conformation of tandem satellite arrays. From the approaches that lead to the identification of triple-helical DNA in chromosomes, facilities particularly provided by Thiazole Orange use may broaden the investigation on the occurrence of triplex DNA in eukaryotic genomes.
PubMed: 30477098
DOI: 10.3390/cells7120227 -
Biochemistry. Biokhimiia Apr 2018Drosophila polytene chromosomes are widely used as a model of eukaryotic interphase chromosomes. The most noticeable feature of polytene chromosome is transverse banding... (Review)
Review
Drosophila polytene chromosomes are widely used as a model of eukaryotic interphase chromosomes. The most noticeable feature of polytene chromosome is transverse banding associated with alternation of dense stripes (dark or black bands) and light diffuse areas that encompass alternating less compact gray bands and interbands visible with an electron microscope. In recent years, several approaches have been developed to predict location of morphological structures of polytene chromosomes based on the distribution of proteins on the molecular map of Drosophila genome. Comparison of these structures with the results of analysis of the three-dimensional chromatin organization by the Hi-C method indicates that the morphology of polytene chromosomes represents direct visualization of the interphase nucleus spatial organization into topological domains. Compact black bands correspond to the extended topological domains of inactive chromatin, while interbands are the barriers between the adjacent domains. Here, we discuss the prospects of using polytene chromosomes to study mechanisms of spatial organization of interphase chromosomes, as well as their dynamics and evolution.
Topics: Animals; Chromatin; Chromatin Assembly and Disassembly; Drosophila; Polytene Chromosomes
PubMed: 29626921
DOI: 10.1134/S0006297918040053 -
Cell Cycle (Georgetown, Tex.) Jan 2017
Topics: Amyloid; Animals; Chromosomal Puffs; Mitochondria; Models, Biological; Oocytes; Oogenesis; Xenopus; Xenopus Proteins
PubMed: 27736303
DOI: 10.1080/15384101.2016.1241605