The Role of Chromosome-Nuclear Envelope Attachments in 3D Genome Organization.

Chromosomes are intricately folded and packaged in the cell nucleus and interact with the nuclear envelope. This complex nuclear architecture has a profound effect on how the genome works and how the cells function. The main goal of review is to highlight recent studies on the effect of chromosome-nuclear envelope interactions on chromatin folding and function in the nucleus. The data obtained suggest that chromosome-nuclear envelope attachments are important for the organization of nuclear architecture in various organisms. A combination of experimental cell biology methods with computational modeling offers a unique opportunity to explore the fundamental relationships between different aspects of 3D genome organization in greater details. This powerful interdisciplinary approach could reveal how the organization and function of the genome in the nuclear space is affected by the chromosome-nuclear envelope attachments and will enable the development of novel approaches to regulate gene expression.

INFESTATION OF HONEYBEE (APIS MELLIFERA) FAMILIES BY MICROSPORIDIANS OF THE GENUS NOSEMA IN TOMSK PROVINCE

Infestation of bee colonies and apiaries by representatives of the genus Nosema, microsporidian protozoans of European honeybees (Apis mellifera L.), causing nosemosis, in Tomsk Province was investigated. In 2012­2015, nosemosis was detected in 32 out of 124 honeybee colonies (31.3 %) and in 20 out of 64 studied apiaries (25.8 %). The maximal infestation rate of bee colonies and apiaries constituted more than 40 % in 2014­2015. N. apis pathogen was registered in 84.4 % of infected bee colonies (16 apiaries); N. ceranae was identified in 9.4 % of infected bee colonies (2 apiaries); and co-infection (N. apis and N. ceranae) was detected in 6.3 % of infected bee colonies (2 apiaries). The reasons of the spreading of the nosemosis, such as climatic conditions, control of imported bee colonies on the presence of Nosema infection, and some others are discussed.

A standard photomap of ovarian nurse cell chromosomes in the European malaria vector Anopheles atroparvus.

Anopheles atroparvus (Diptera: Culicidae) is one of the main malaria vectors of the Maculipennis group in Europe. Cytogenetic analysis based on salivary gland chromosomes has been used in taxonomic and population genetic studies of mosquitoes from this group. However, a high-resolution cytogenetic map that could be used in physical genome mapping in An. atroparvus is still lacking. In the present study, a high-quality photomap of the polytene chromosomes from ovarian nurse cells of An. atroparvus was developed. Using fluorescent in situ hybridization, 10 genes from the five largest genomic supercontigs on the polytene chromosome were localized and 28% of the genome was anchored to the cytogenetic map. The study established chromosome arm homology between An. atroparvus and the major African malaria vector Anopheles gambiae, suggesting a whole-arm translocation between autosomes of these two species. The standard photomap constructed for ovarian nurse cell chromosomes of An. atroparvus will be useful for routine physical mapping. This map will assist in the development of a fine-scale chromosome-based genome assembly for this species and will also facilitate comparative and evolutionary genomics studies in the genus Anopheles.

A standard cytogenetic map for Anopheles sinensis and chromosome arm homology between the subgenera Anopheles and Cellia.

Anopheles sinensis (Diptera: Culicidae) is an important vector of Plasmodium vivax in Southeast Asia. To facilitate population genetic and genomic studies of An. sinensis, we developed a standard cytogenetic photomap for this species. The polytene chromosomes were straightened and divided into 39 numbered divisions and 116 lettered subdivisions. The chromosomal localizations of 13 DNA probes were determined by fluorescent in situ hybridization. A comparison of the physical map for An. sinensis with the genome map for Anopheles gambiae revealed a whole-arm autosomal translocation between the two species. Specifically, the 2R arm of An. gambiae corresponds to the 3R arm of An. sinensis and the pattern of correspondence of the other chromosome arms remains regular. We mapped the breakpoints of the polymorphic paracentric chromosomal inversion 3Ra to subdivisions 28A and 31A. The standard cytogenetic map developed in this study will be useful for detailed comparative genome mapping and population genetic studies of An. sinensis.

Polymorphic chromosomal inversions in Anopheles moucheti, a major malaria vector in Central Africa.

Anopheles moucheti Evans (Diptera: Culicidae) is a major vector of malaria in forested areas of Central Africa. However, few genetic tools are available for this species. The present study represents the first attempt to characterize chromosomes in An. moucheti females collected in Cameroon. Ovarian nurse cells contained polytene chromosomes, which were suitable for standard cytogenetic applications. The presence of three polymorphic chromosomal inversions in An. moucheti was revealed. Two of these inversions were located on the 2R chromosome arm. The homology between the 2R chromosome arms of An. moucheti and Anopheles gambiae Giles was established by fluorescent in situ hybridization of six An. gambiae genic sequences. Mapping of the probes on chromosomes of An. moucheti detected substantial gene order reshuffling between the two species. The presence of polytene chromosomes and polymorphic inversions in An. moucheti provides a new basis for further population genetic, taxonomic and ecological studies of this neglected malaria vector.

Chromosome phylogenies of malaria mosquitoes.

Malaria mosquitoes often belong to complexes of sibling species, members of which are morphologically and genetically similar to each other. However, members within these complexes can vary significantly in their ecological adaptations and abilities to transmit the malaria parasite. The high degree of genetics similarity among sibling species makes the reconstruction of phylogenetic relationships within species complexes difficult. This paper reviews studies that infer the ancestral--descendant relationships among sibling species using molecular markers and chromosomal inversions. A methodology based on analyzing breakpoints of fixed overlapping inversions is shown to be useful for rooting phylogenies in complexes of sibling species, if the chromosomal arrangements in outgroup species are known. The construction of detailed phylogenies for malaria vectors will help to identify the association of evolutionary genomic changes with the origin of human blood choice and specific ecological adaptations.

Progress in mapping the yellow fever mosquito genome.

Mosquito-borne diseases cause significant problems for the human health. For this reason, the genomes of three most dangerous species of mosquitoes, including the yellow fever mosquito Aedes aegypti, were sequenced in last decade. The efficient vector of arboviruses. Ae. aegypti, is also a convenient model for laboratory research. The intensive genetic mapping of morphological and molecular markers conducted for this mosquito in the past was very successful. This mapping was also used as a tool to localize a number of quantitative trait loci related to the mosquito's ability to transmit various pathogens. However, physical mapping of the Ae. aegypti genome is difficult due to the lack of high-quality polytene chromosomes. Here, we review different mapping approaches that help improving genome sequence assembly and also integrate linkage, chromocome and genome maps the yellow fever mosquito.

Chromosomal localization of actin genes in the malaria mosquito Anopheles darlingi.

Physical and genetic maps have been used for chromosomal localization of genes in vectors of infectious diseases. The availability of polytene chromosomes in malaria mosquitoes provides a unique opportunity to precisely map genes of interest. We report the physical mapping of two actin genes on polytene chromosomes of the major malaria vector in the Amazon, Anopheles darlingi (Diptera: Culicidae). Clones with actin gene sequences were obtained from a cDNA library constructed from RNA isolated from adult females and males of An. darlingi. Each of the two clones was mapped to a unique site on chromosomal arm 2L in subdivisions 21A (clone pl05-A04) and 23B (clone pl17-G06). The obtained results, together with previous mapping data, provide a suitable basis for comparative genomics and for establishing chromosomal homologies among major malaria vectors.

Chromosome evolution in malaria mosquitoes.

The genomic era offers excellent opportunities to improve our understanding of the genetic basis of mosquito adaptation, evolution, and competence to a pathogen. The availability of polytene chromosomes in anopheline mosquitoes makes them an excellent model system for studying genome organization, evolution, and function. Physical mapping facilitated the whole genome sequence assembly for the major malaria vector Anopheles gambiae and comparative genome mapping has determined types, patterns, and rates of chromosomal rearrangements in mosquito evolution. Together with sequencing projects, high-resolution physical mapping can shed light on mechanisms of chromosomal rearrangements and phylogenetic relationships among species.

High-resolution cytogenetic map for the African malaria vector Anopheles gambiae.

Cytogenetic and physical maps are indispensible for precise assembly of genome sequences, functional characterization of chromosomal regions, and population genetic and taxonomic studies. We have created a new cytogenetic map for Anopheles gambiae by using a high-pressure squash technique that increases overall band clarity. To link chromosomal regions to the genome sequence, we attached genome coordinates, based on 302 markers of bacterial artificial chromosome, cDNA clones, and PCR-amplified gene fragments, to the chromosomal bands and interbands at approximately a 0.5-1 Mb interval. In addition, we placed the breakpoints of seven common polymorphic inversions on the map and described the chromosomal landmarks for the arm and inversion identification. The map's increased resolution can be used to further enhance physical mapping, improve genome assembly, and stimulate epigenomic studies of malaria vectors.

Organization and evolution of heterochromatin in malaria mosquitoes.

The African malaria mosquito Anopheles gambiae was the first disease vector chosen for genome sequencing. Although its genome assembly has been facilitated by physical mapping, large gaps still pose a serious problem for accurate annotation and genome analysis. The majority of the gaps are located in regions of pericentromeric and intercalary heterochromatin. Genomic analysis has identified protein-coding genes and various classes of repetitive elements in the Anopheles heterochromatin. Molecular and cytogenetic studies have demonstrated that heterochromatin is a structurally heterogeneous and rapidly evolving part of the malaria mosquito genome.

[Localization of repetitive DNA sequences in the pericentromeric heterochromatin of malarial mosquitoes of the "Anopheles maculipennis" complex].

Distribution of eight fragments of conserved repetitive DNA from pericentromeric heterochromatin of chromosome 2 of Anopheles atroparvus has been investigated by in situ hybridization on polytene chromosomes of An. atroparvus and An. messeae. We have shown that heterochromatic regions of all chromosomes both in An. atroparvus and An. messeae vary in combinations of, at least, conserved repeats. Some repeats have been found only in pericentromeric heterochromatic regions of chromosomes 2 (clones Atr2R-46a, Atr2R-73, Atr2R-85a in An. atroparvus and Atr2R-25 in An. messeae). Others have been found in two (clones Atr2R-25a and Atr2R-90 in An. atroparvus, Atr2R-25a in An. messeae) and more (clones Atr2R-118, Atr2R-136 in An. atroparvus, Atr2R-73 in An. messeae) pericentromeric heterochromatic regions of chromosomes. DNA comparison of pericentromeric heterochromatic regions of chromosomes in species of the "Anopheles maculipennis" complex is species- and chromosome-specific, due, in particular, to different maintenance of conserved repeates.

[Specific DNA sequences are associated with nuclear envelopes of pseudonurse cells in Drosophila melanogaster otu 11].

Nuclei of ovarian pseudonurse cells from the mutant strain of Drosophila melanogaster otu 11 are suitable for mapping the attachment of chromosomes to the nuclear envelope (NE). Loci in contact with the NE included region 20CD of the X chromosome, region 41 of chromosome 2, the proximal end of region 81 of chromosome 3, and region 101 of chromosome 4. In situ hybridization revealed that all 4 regions contained sequences homologous to clone lambda20p1.4. DNA of clone lambda20p1.4 was previously found to bind specifically to purified D. melanogaster lamins. These results suggest that specific DNA sequences are involved in attachment of chromosomes to NE in vivo.

[A combined approach to mapping the proximal border of the right arm of polytene chromosome 2 in Drosophila melanogaster otu 11].

A combined approach based on cytological observations in situ hybridization, and qualitative Southern-blot analyses were used to localize the proximal border of the right arm of polytene chromosome 2 in Drosophila melanogaster otu 11 strain. A genetically functional chromosome 2 is bounded by "deletions" C', C, D, B, A and ms2-10. Using in situ hybridization in conjunction with comparative quantitative Southern-blot hybridization to deletions in centromeric heterochromatin, DNA of specific centromeric clone lambda20p1.4 was localized with respect to "deletions" and on otu 11 polytene chromosomes. Comparison of hybridization sites of lambda20p1.4 on polytene chromosomes, and its amount in mutant lines of D. melanogaster carrying known "deletions" in the centromeric heterochromatin enabled us to localize the proximal border of the right arm of chromosome 2 in D. melanogaster otu 11 strain between the 39/40 region and hybridization site of the k20p1.4 DNA fragment.

A microsatellite map of the African human malaria vector Anopheles funestus.

Microsatellite markers and chromosomal inversion polymorphisms are useful genetic markers for determining population structure in Anopheline mosquitoes. In Anopheles funestus (2N = 6), only chromosome arms 2R, 3R, and 3L are known to carry polymorphic inversions. The physical location of microsatellite markers with respect to polymorphic inversions is potentially important information for interpreting population genetic structure, yet none of the available marker sets have been physically mapped in this species. Accordingly, we mapped 32 polymorphic A. funestus microsatellite markers to the polytene chromosomes using fluorescent in situ hybridization (FISH) and identified 16 markers outside of known polymorphic inversions. Here we provide an integrated polytene chromosome map for A. funestus that includes the breakpoints of all known polymorphic inversions as well as the physical locations of microsatellite loci developed to date. Based on this map, we suggest a standard set of 16 polymorphic microsatellite markers that are distributed evenly across the chromosome complement, occur predominantly outside of inversions, and amplify reliably. Adoption of this set by researchers working in different regions of Africa will facilitate metapopulation analyses of this primary malaria vector.

[Chromosome localization of the lambda20p1.4 clone of the Drosophila melanogaster nuclear lamina DNA in the melanogaster species subgroup of the genus Drosophila (Sophophora)]. / Khromosomnaia lokalizatsiia klona lambda20p1.4 DNK iadernoi laminy Drosophila melanogaster u vidov podgruppy melanogaster (rod Drosophila (Sophophora)).

Chromosome localization of sequences homologous to lambda 20p1.4 of the Drosophila melanogaster nuclear lamina DNA (nlDNA) was established by in situ hybridization in species of the melanogaster subgroup. DNA of the lambda 20p1.4 clone was shown to be located in the chromocenter in all the species examined. Laboratory strains of D. simulans, D. mauritiana, and D. sechellia exhibited interspecific differences in localization of lambda 20p1.4 nlDNA on chromosome arms. In eight natural populations, intraspecific polymorphism of lambda 20p1.4 nlDNA chromosome localization was shown to be present in D. simulans but absent in D. melanogaster. The possible participation of transposable elements in nlDNA relocation is discussed.

Linear and spatial organization of polytene chromosomes of the African malaria mosquito Anopheles funestus.

Anopheles funestus Giles is one of the major malaria vectors in Africa, but little is known about its genetics. Lack of a cytogenetic map characterized by regions has hindered the progress of genetic research with this important species. This study developed a cytogenetic map of An. funestus using ovarian nurse cell polytene chromosomes. We demonstrate an important application with the cytogenetic map for characterizing various chromosomal inversions for specimens collected from coastal Kenya. The linear and spatial organization of An. funestus polytene chromosomes was compared with the best-studied malaria mosquito, An. gambiae Giles. Comparisons of chromosome morphology between the two species have revealed that the most extensive chromosomal rearrangement occurs in pericentromeric heterochromatin of autosomes. Differences in pericentromeric heterochromatin types correlate with nuclear organization differences between An. funestus and An. gambiae. Attachments of chromosomes to the nuclear envelope strongly depend on the presence of diffusive beta-heterochromatin. Thus, An. funestus and An. gambiae exhibit species-specific characteristics in chromosome-linear and -spatial organizations.

Release of mitotic descendants by giant cells from irradiated Burkitt's lymphoma cell line..

Polyploid giant cells are produced as part of the response of p53 mutant Burkitt's lymphoma cell lines to high doses of irradiation. Polyploid giant cells arise by endo-reduplication in the first week after a single 10 Gray dose of irradiation. Within the giant cells a sub-nuclear structure is apparent and within this, sub-nuclear autonomy is evident, as displayed by independent nuclear structure and DNA replication in different parts of the nucleus. The majority of these cells soon die as apoptotic polykaryons. However, approximately 10-20% of giant cells remain viable into the second week after irradiation and begin vigorous extrusion of large degraded chromatin masses. During the second week, the giant cells begin to reconstruct their nuclei into polyploid 'bouquets', where chromosome double-loops are formed. Subsequently, the bouquets return to an interphase state and separate into several secondary nuclei. The individual sub-nuclei then resume DNA synthesis with mitotic divisions and sequester cytoplasmic territories around themselves, giving rise to the secondary cells, which continue mitotic propagation. This process of giant cell formation, reorganization and breakdown appears to provide an additional mechanism for repairing double-strand DNA breaks within tumour cells.

[Pericentromeric and intercalating alpha-heterochromatin of polytene chromosomes of the malaria mosquito]. / Pritsentromernyi i interkaliarnyi alpha-geterokhromatin politennykh khromosom maliariinykh komarov.

The properties of heterochromatin from polytene chromosomes of the malaria mosquito Anopheles messeae Fal. and A. atroparvus V. Tiel. were studied by various methods of differential staining and by hybridization in situ with two repetitive DNA sequences of Drosophila melanogaster. In malaria mosquito, the heterochromatin was heterogeneous. Two forms of alpha-heterochromatin were revealed: pericentromeric and intercalary heterochromatin, which is localized within the internal chromosome regions.

[Variability of pericentromeric chromatin in chromosome 2 of ovarian nurse cells during inbreeding in Anopheles atroparvus V.Tiel]. / Izmenchivost' pritsentromernogo geterokhromatina khromosomy 2 pitaiushchikh kletok iaichnikov pri inbridinge u Anopheles atroparvus V. Tiel.

We investigated the variability of pericentromeric chromatin of chromosome 2 in ovarian nurse cells (trophocytes) in two laboratory lines of malaria mosquito Anopheles atroparvus V. Tiel and in their hybrids. One line had been raised by means of sib inbreeding, the other kept at constantly high population density. The inbreeding was shown to result in an increased percentage of chromosomes bearing an achromatinic zone in the centromeric region, which resulted in chromosome breakage. Toxicological tests demonstrated an increase in the sensitivity of the progeny of females with abnormal morphotypes of chromosome 2 to the entomopathogenic bacterium Bacillus thuringiensis israelensis. The appearance of the achromatinic zone is attributed to local chromatin underreplication accompanying chromosome polytenization. Possible reasons for this phenomenon and its implication for adaptation are discussed.