Лаборатория структуры и функций генов человека

Отдел геномики и постгеномных технологий

Руководитель: Свердлов Евгений Давидович, академик
sverd@ibch.ru+7(495)330-65-29

Функциональная геномика бактерий и человека, молекулярные механизмы канцерогенеза, взаимодействие патоген-хозяин, диагностика и генная терапия онкологических заболеваний

Разработка подходов к диагностике и генной терапии онкологических заболеваний (Т. В. Виноградова, Е. П. Копанцев, И. П. Чернов)

Большое внимание в последние годы уделяется проблемам создания лекарств нового поколения. В рамках федеральной целевой программы «Исследования и разработки по приоритетным направлениям развития научно-технологического комплекса России на 2007–2012 годы» с участием лаборатории осуществляется крупный комплексный проект «Разработка и выпуск опытных партий новых эффективных направленно-модифицированных терапевтических и диагностических средств постгеномной генерации для использования в онкологической практике». Целью этого проекта является создание и выпуск в клинику первых генно-терапевтических препаратов для лечения рака легкого и пищевода. Используемые в проекте подходы базируются на фундаментальных исследованиях и методических разработках, выполненных в лаборатории. Проект объединяет крупнейшие научные центры страны: Институт молекулярной генетики РАН, Российский онкологический научный центр им. Н. Н. Блохина РАМН, Институт биоорганической химии им. академиков М. М. Шемякина и Ю. А. Овчинникова, Новосибирский государственный университет, Институт биологии гена РАН, Институт эпидемиологии и микробиологии им. Н. Ф. Гамалеи РАМН, а также биотехнологические компании ЗАО «Евроген» и ЗАО «Биннофарм». В рамках проекта созданы конструкции, позволяющие осуществлять специфическую экспрессию терапевтических генов в раковых клетках, а также системы доставки терапевтических генов в опухоли.

Таким образом, была создана основа для разработки и производства новых эффективных медицинских препаратов для онкологии: терапевтических средств на основе генно-терапевтического подхода, терапевтических средств на основе биологически активных белков и диагностических препаратов.

Рак и развитие (М. В. Зиновьева, Т. В. Виноградова, Г. С. Монастырская)

Одним из важнейших результатов, полученных в рамках этого проекта, является демонстрация того, что гены, которые включаются в эмбриогенезе при развитии легких и пищевода, выключаются при превращении нормальных клеток в раковые и наоборот. Тем самым подтверждается гипотеза о том, что возникновение и развитие раковых клеток можно сравнить с эмбриональным развитием или самовоспроизводством и дифференцировкой стволовых клеток тех или иных тканей. Гены, имеющие дифференциальную экспрессию и при раке и при развитии, возможно, являются ключевыми для опухолеобразования. Исследование таких генов позволяет осуществлять рациональный выбор мишеней терапевтического воздействия: наиболее перспективными мишенями являются системы, регулирующие эмбриональное развитие и нарушенные при канцерогенезе.

 

Модифицированные олигонуклеотиды (В. К. Потапов)

Главным направлением работы группы в последние 3 года было разработка методов получения и синтез модифицированных нуклеозидов с измененной структурой основания или сахаро-фосфатного остова. Наиболее перспективным направлением является разработка подходов к твердофазному синтезу морфолино-олигонуклеотидов — аналогов, несущих в своем составе вместо рибозы остаток морфолина. Аналоги сохраняют структуру ДНК, образуют прочные комплементарные комплесы с природными олигонуклеотидами и являются незаряженными молекулами, устойчивыми к действию нуклеаз.

Второе направление — синтез олигонуклеотидов, несущих в определенных местах спейсеры заданной длины с тиольной группой, которые используются группой проф. Влодовера из NCI (Frederic) для изучения комплексов с интегразами HIV—1 и ASV методами рентгеноструктурного анализа.

Микроокружение опухоли и экспрессия генов (Плешкан Виктор, Антонова Дина)

По современным представлениям опухоль представляет собой сложную структуру, состоящую из собственно опухолевых клеток и клеток опухолевого микроокружения. Микроокружение, или строма, представляет собой опухоль ассоциированные фибробласты (ОАФ), иммунные клетки, цитокины и другие элементы. Строма необходима для поддержания опухолевой прогрессии, защиты и пролиферации опухолевых клеток. Многообразное взаимодействие опухолевых и стромальных клеток обеспечивает эти процессы. Различные паттерны экспрессии генов характерны для опухолевых и стромальных клеток. Многие из таких дифференциально экспрессирующихся генов являются ключевыми для взаимодействия «опухоль-строма».

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Опухолевое микроокружение (опухолевая строма) – это окружающие опухолевые клетки компоненты внеклеточного матрикса и неопухолевые клетки: эндотелиальные клетки, клетки иммунной системы, опухоль-ассоциированные фибробласты (ОАФ).

Нами исследуются ОАФ, их основные маркеры и способы воздействия на них, как на потенциальную мишень при противоопухолевой терапии. В Лаборатории были идентифицированы несколько генов, обладающих повышенной экспрессией в ОАФ. Мы исследуем промоторы этих генов на предмет тканеспецифичной экспрессии в клетках опухолевого микроокружения для использования в генной терапии. Также мы исследуем продукты генов опухолевого микроокружения и их роли при прогрессии опухоли и возможностях их использования для создания новых противоопухолевых подходов. Так, мы исследуем влияние фактора роста соединительной ткани CTGF на миграцию опухолевых клеток и его взаимосвязь c другими патологическими процессами при опухолевой прогрессии. В нашей группе проводятся эксперименты по выявлению потенциала опухоль-ассоциированных фибробластов, которые выступают в качестве мишени при противоопухолевой терапии. Мы готовы работать с новыми студентами, которые обладают возможностью и желанием усердно работать по данной тематике.

Ф.И.О.ДолжностьКонтакты
Свердлов Евгений Давидович, академикрук. подр.sverd@ibch.ru+7(495)330-65-29
Монастырская Галина Сергеевна, к. х. н., старший научный сотрудникв.н.с.gal@ibch.ru+7(495)330-65-38
Потапов Виктор Кузьмич, д. х. н.в.н.с.vk@ibch.ru
Николаев Лев Григорьевич, д. б. н.в.н.с.lev@ibch.ru+7(495)3307029
Чернов Игорь Павлович, к. х. н.с.н.с.igor_ch@ibch.ru+7(495)330-70-29
Акопов Сергей Борисович, к. б. н.с.н.с.akser@ibch.ru+7(495)330-70-29
Виноградова Татьяна Викторовна, к. б. н.с.н.с.tv@ibch.ru+7(495)330-65-47
Копанцев Евгений Павлович, к. б. н.с.н.с.kopantzev@ibch.ru+7(495)3307029
Снежков Евгений Валерьевич, к. х. н.с.н.с.eugene@ibch.ru+7(495)330-70-29
Успенская Наталия Яковлевна, к. б. н.с.н.с.usp@ibch.ru+7(495)330-58-10
Копанцева Марина Робертовнас.н.с.
Коробко Игорь Викторовичс.н.с.
Дидыч Дмитрий Александрович, к. б. н.н.с.dmitry_d@inbox.ru+7(495)330-70-29
Буланенкова Светлана Сергеевна, к. б. н.н.с.
Кашкин Кирилл Никитич, к. б. н.н.с.kaschkin@ibch.ru+7(495)3306992
Филюкова Ольга Борисовна, к. б. н.н.с.+7(495)330-69-92
Плешкан Виктор Викторович, к. б. н.н.с.vpleshkan@gmail.com+7(903)297-43-09
Алексеенко Ирина Васильевна, к. б. н.н.с.irina.alekseenko@mail.ru
Кузьмич Алексей Ивановичн.с.
Сасс Александр Викторовичм.н.с.+7(495)330-70-29
Зиновьева Марина Валерьевна, к. б. н.м.н.с.mzinov@ibch.ru+7(495)330-65-47
Кондратьева Лия Германовнаасп.liakondratyeva@yandex.ru+7(495)3307029
Гнатенко Дмитрий Александровичасп.gnatenkodmitrij@gmail.com
Аверина Людмила Викторовнаинженер+7(495)330-63-29
Мелехина Ольга Вячеславовнаинженер
Полтавцева Римма Алексеевнаинженер
Красько Анастасия Марковнаинженер
Кравченко Зоя Борисовнаинж.-иссл.+7(495)3306329
Антонова Дина Васильевнаинж.-иссл.tyulkina.dina@mail.ru
Погосян Рудольф Андреевичвед. инж.r.j.f@mail.ru+7(495)3306547
Левитан Татьяна Львовнаст. инж.levitanibh@mail.ru+7(495)3306529
Медведева Наталия Игоревнаст. инж.+7(495)3306329

Ранее здесь работали:

Ажикина Татьяна Леодоровна, д. б. н.в.н.с.tatazhik@ibch.ru
Бони Ирина Венедиктовна, к. х. н.с.н.с.irina_boni@ibch.ru
Завалова Людмила Львовна, к. б. н.с.н.с.lz@ibch.ru
Скворцов Тимофей Алексеевич, к. б. н.н.с.timofey@ibch.ru
Беляева Нина Николаевна, к. б. н.н.с.
Скапцова Надежда Васильевнан.с.
Быченко Оксана Станиславовна, к. б. н.н.с.
Гайнетдинов Ильдар Ваисович, к. б. н.н.с.ildargv@gmail.com
Колединская Людмила Сергеевнам.н.с.
Скворцова Юлия Валентиновна, к. б. н.м.н.с.
Билтуева Юлия асп.bjuliya87@mail.ru
Кондратьева София Алексеевнаасп.sofia.a.kondr@gmail.com
Мазурова Арина Сергеевнаасп.mazarina@ibch.ru
Кротов Сергей ВикторовичинженерSerg07@pochta.ru

Избранные публикации

  1. Gainetdinov I.V., Kondratieva S.A., Skvortsova Y.V., Zinovyeva M.V., Stukacheva E.A., Klimov A., Tryakin A.A., Azhikina T.L. (2016). Distinguishing epigenetic features of preneoplastic testis tissues adjacent to seminomas and nonseminomas. Oncotarget , [+]

    PIWI pathway proteins are expressed during spermatogenesis where they play a key role in germ cell development. Epigenetic loss of PIWI proteins expression was previously demonstrated in testicular germ cell tumors (TGCTs), implying their involvement in TGCT development. In this work, apart from studying only normal testis and TGCT samples, we also analyzed an intermediate stage, i.e. preneoplastic testis tissues adjacent to TGCTs. Importantly, in this study, we minimized the contribution of patient-to-patient heterogeneity by using matched preneoplastic/TGCT samples. Surprisingly, expression of germ cell marker DDX4 suggests that spermatogenesis is retained in premalignant testis tissues adjacent to nonseminoma, but not those adjacent to seminoma. Moreover, this pattern is followed by expression of PIWI pathway genes, which impacts one of their functions: DNA methylation level over LINE-1 promoters is higher in preneoplastic testis tissues adjacent to nonseminomas than those adjacent to seminomas. This finding might imply distinct routes for development of the two types of TGCTs and could be used as a novel diagnostic marker, possibly, noninvasively. Finally, we studied the role of CpG island methylation in expression of PIWI genes in patient samples and using in vitro experiments in cell line models: a more complex interrelation between DNA methylation and expression of the corresponding genes was revealed.

    ID:1361
  2. Aseev L.V., Chugunov A.O., Efremov R.G., Boni I.V. (2013). A single missense mutation in a coiled-coil domain of Escherichia coli ribosomal protein S2 confers a thermosensitive phenotype that can be suppressed by ribosomal protein S1. J. Bacteriol. 195 (1), 95–104 [+]

    Ribosomal protein S2 is an essential component of translation machinery, and its viable mutated variants conferring distinct phenotypes serve as a valuable tool in studying the role of S2 in translation regulation. One of a few available rpsB mutants, rpsB1, shows thermosensitivity and ensures enhanced expression of leaderless mRNAs. In this study, we identified the nature of the rpsB1 mutation. Sequencing of the rpsB1 allele revealed a G-to-A transition in the part of the rpsB gene which encodes a coiled-coil domain of S2. The resulting E132K substitution resides in a highly conserved site, TKKE, a so-called N-terminal capping box, at the beginning of the second alpha helix. The protruding coiled-coil domain of S2 is known to provide binding with 16S rRNA in the head of the 30S subunit and, in addition, to interact with a key mRNA binding protein, S1. Molecular dynamics simulations revealed a detrimental impact of the E132K mutation on the coiled-coil structure and thereby on the interactions between S2 and 16S rRNA, providing a clue for the thermosensitivity of the rpsB1 mutant. Using a strain producing a leaderless lacZ transcript from the chromosomal lac promoter, we demonstrated that not only the rpsB1 mutation generating S2/S1-deficient ribosomes but also the rpsA::IS10 mutation leading to partial deficiency in S1 alone increased translation efficiency of the leaderless mRNA by about 10-fold. Moderate overexpression of S1 relieved all these effects and, moreover, suppressed the thermosensitive phenotype of rpsB1, indicating the role of S1 as an extragenic suppressor of the E132K mutation.

    ID:812
  3. Ignatov D., Malakho S., Majorov K., Skvortsov T., Apt A., Azhikina T. (2013). RNA-Seq Analysis of Mycobacterium avium Non-Coding Transcriptome. PLoS ONE 8 (9), e74209 [+]

    Deep sequencing was implemented to study the transcriptional landscape of Mycobacterium avium. High-resolution transcriptome analysis identified the transcription start points for 652 genes. One third of these genes represented leaderless transcripts, whereas the rest of the transcripts had 5' UTRs with the mean length of 83 nt. In addition, the 5' UTRs of 6 genes contained SAM-IV and Ykok types of riboswitches. 87 antisense RNAs and 10 intergenic small RNAs were mapped. 6 intergenic small RNAs, including 4.5S RNA and rnpB, were transcribed at extremely high levels. Although several intergenic sRNAs are conserved in M. avium and M. tuberculosis, both of these species have unique intergenic sRNAs. Moreover, we demonstrated that even conserved small RNAs are regulated differently in these species. Different sets of intergenic sRNAs may underlie differences in physiology between conditionally pathogenic M. avium and highly specialized pathogen M. tuberculosis.

    ID:866
  4. Ignatov D., Kondratieva E., Azhikina T., Apt A. (2012). Mycobacterium avium-triggered diseases: pathogenomics. Cell. Microbiol. 14 (6), 808–18 [+]

    The species Mycobacterium avium includes several subspecies representing highly specialized avian and mammalian pathogens, non-obligatory pathogens of immune compromised humans and saprophitic organisms. Recently obtained information concerning the diversity of M. avium genomic structures not only clarified phylogenic relationships within this species, but began to shed light on the question of how such closely related microorganisms adapt to the occupation of distinct ecological niches. In this review we discuss specific features of M. avium genetic composition, as well as genetic and molecular aspects of M. avium hominissuis (MAH)-triggered disease pathogenesis, including virulence, penetration, immune response manipulation and host genetic control.

    ID:868
  5. Azhikina T., Kozlova A., Skvortsov T., Sverdlov E. (2011). Heterogeneity and degree of TIMP4, GATA4, SOX18, and EGFL7 gene promoter methylation in non-small cell lung cancer and surrounding tissues. Cancer Genet 204 (9), 492–500 [+]

    We used methylation-sensitive high resolution melting analysis to assess methylation of CpG islands within the promoters of the TIMP4, GATA4, SOX18, and EGFL7 genes in samples of non-small cell lung cancer and surrounding apparently normal tissue and noncancerous lung tissues. We found that the promoter methylation was heterogeneous in both tumor and surrounding normal tissue. This is in contrast to healthy lung tissue, where the promoters were normally either non- or hypomethylated, and the heterogeneity of methylation was low. An increased heterogeneity of methylation in the normal tissues surrounding the tumor may suggest an early start of epigenetic processes preceding genetic and morphologic changes and can be used as a biomarker of early cancerization events. This analysis is an easy and sensitive tool for studying epigenetic heterogeneity and could be used in clinical practice.

    ID:704
  6. Azhikina T., Skvortsov T., Radaeva T., Mardanov A., Ravin N., Apt A., Sverdlov E. (2010). A new technique for obtaining whole pathogen transcriptomes from infected host tissues. BioTechniques 48 (2), 139–44 [+]

    We propose a novel experimental approach based on coincidence cloning for analyzing sequences of bacterial intracellular pathogens specifically transcribed in affected tissues. Co-denaturation and co-renaturation of excess bacterial genomic DNA with the cDNA prepared on total RNA of the infected tissue allows one to select the bacterial fraction of the cDNA sample. We used this technique for preparing and characterizing the Mycobacterium tuberculosis cDNA pool, representing the transcriptome of infected mouse lungs in the chronic phase of infection. A cDNA pool enriched in fragments of mycobacterial cDNA was analyzed by the high-throughput 454 sequencing procedure. We demonstrated that its composition corresponded to what can be expected in the chronic phase of infection and, after the adaptation of M. tuberculosis to the host immune system, was characterized by an active lipid metabolism and switched from aerobic to anaerobic respiration. The technique is universal and requires no prior knowledge of the pathogen genome sequence. Pools of transcribed sequences obtained by this technique retain the main characteristics of the genome-wide gene transcription pattern within infected tissue, and can be used for in vivo analysis of gene expression of a wide spectrum of infection agents, such as viruses, bacteria, and protista.

    ID:701
  7. Kopantzev E.P., Monastyrskaya G.S., Vinogradova T.V., Zinovyeva M.V., Kostina M.B., Filyukova O.B., Tonevitsky A.G., Sukhikh G.T., Sverdlov E.D. (2008). Differences in gene expression levels between early and later stages of human lung development are opposite to those between normal lung tissue and non-small lung cell carcinoma. Lung Cancer 62 (1), 23–34 [+]

    We, for the first time, directly compared gene expression profiles in human non-small cell lung carcinomas (NSCLCs) and in human fetal lung development. Previously reported correlations of gene expression profiles between lung cancer and lung development, deduced from matching data on mouse development and human cancer, have brought important information, but suffered from different timing of mouse and human gene expression during fetal development and fundamental differences in tumorigenesis in mice and humans. We used the suppression subtractive hybridization technique to subtract cDNAs prepared from human fetal lung samples at weeks 10-12 and 22-24 and obtained a cDNA library enriched in the transcripts more abundant at the later stage. cDNAs sequencing and RT-PCR analysis of RNAs from human fetal and adult lungs revealed 12 differentially transcribed genes: ADH1B, AQP1, FOLR1, SLC34A2, CAV1, INMT, TXNIP, TPM4, ICAM-1, HLA-DRA, EFNA1 and HLA-E. Most of these genes were found up-regulated in mice and rats at later stages than in human lung development. In surgical samples of NSCLC, these genes were down-regulated as compared to surrounding normal tissues and normal lungs, thus demonstrating opposite expression profiles for the genes up-regulated during fetal lung development.

    ID:4
  8. Chernov I.P., Timchenko K.A., Akopov S.B., Nikolaev L.G., Sverdlov E.D. (2007). Identification of tissue-specific DNA-protein binding sites by means of two-dimensional electrophoretic mobility shift assay display. Anal. Biochem. 364 (1), 60–6 [+]

    We developed a technique of differential electrophoretic mobility shift assay (EMSA) display allowing identification of tissue-specific protein-binding sites within long genomic sequences. Using this approach, we identified 10 cell type-specific protein-binding sites (protein target sites [PTSs]) within a 137-kb human chromosome 19 region. In general, tissue-specific binding of proteins from different nuclear extracts by individual PTSs did not follow the all-or-nothing principle. Most often, PTS-protein complexes were formed in all cases, but they were different for different nuclear extracts used.

    ID:5
  9. Bulanenkova S., Snezhkov E., Nikolaev L., Sverdlov E. (2007). Identification and mapping of open chromatin regions within a 140 kb polygenic locus of human chromosome 19 using E. coli Dam methylase. Genetica 130 (1), 83–92 [+]

    Using transient expression of the E. coli Dam methylase gene and analysis of the distribution of methylated GATC sites, we studied the distribution of open chromatin regions within a 140 kb long human genome segment in HEK-293 cells. Dam methylated sites were found in gene introns, exons, and intergenic regions, and their distribution along DNA was uneven. There were regions of high and low density of Dam methylated GATC sites, presumably corresponding to "open" and "closed" chromatin regions, respectively, and to the functional profile of the genomic locus under study. The Dam methylation profile was also generally in agreement with transcriptional activity of genes in the locus. Moreover, DNA regions accessible to Dam methylase apparently coincided with those hypersensitive to DNase I.

    ID:11
  10. Illarionova A.E., Vinogradova T.V., Sverdlov E.D. (2007). Only those genes of the KIAA1245 gene subfamily that contain HERV(K) LTRs in their introns are transcriptionally active. Virology 358 (1), 39–47 [+]

    Insertion of LTRs into some genome locations might seriously affect regulation of the neighboring genes expression. This hypothesis is widely accepted but, however, not confirmed directly. Earlier, we have identified a family of closely related genes highly similar to the KIAA1245 mRNA counterpart. This family included a subfamily of genes some of which contained and the others lacked an LTR in their structure. We compared transcription of several closely related genes of the subfamily differing in the presence or absence of LTRs. Only LTR-containing genes were transcribed in transformed cell lines, tumorous and embryonic human tissues, whereas LTR-lacking genes remained silent. Since the genes were in the same intracellular microenvironment, we suggested that this effect was most probably due to intrinsic cis-characteristics of integrated LTRs and confirmed this by demonstrating high enhancer activity of KIAA1245 LTRs. The expression of the LTR-containing genes in embryonic tissues might suggest their involvement in evolutionary events during primate speciation.

    ID:6
  11. Buzdin A., Kovalskaya-Alexandrova E., Gogvadze E., Sverdlov E. (2006). At least 50% of human-specific HERV-K (HML-2) long terminal repeats serve in vivo as active promoters for host nonrepetitive DNA transcription. J. Virol. 80 (21), 10752–62 [+]

    В статье приведены результаты полногеномного исследования, выявившего, что более 50% всех специфичных для ДНК человека вставок эндогенных ретровирусов активны как промоторы для соседних уникальных участков генома

    ID:16
  12. Akopov S.B., Ruda V.M., Batrak V.V., Vetchinova A.S., Chernov I.P., Nikolaev L.G., Bode J., Sverdlov E.D. (2006). Identification, genome mapping, and CTCF binding of potential insulators within the FXYD5-COX7A1 locus of human chromosome 19q13.12. Mamm. Genome 17 (10), 1042–9 [+]

    Identification of insulators is one of the most difficult problems in functional mapping of genomes. For this reason, up to now only a few insulators have been described. In this article we suggest an approach that allows direct isolation of insulators by a simple positive-negative selection based on blocking enhancer effects by insulators. The approach allows selection of fragments capable of blocking enhancers from mixtures of genomic fragments prepared from up to 1-Mb genomic regions. Using this approach, a 1-Mb human genome locus was analyzed and eight potential insulators were selected. Five of the eight sequences were positioned in intergenic regions and two were within introns. The genes of the alpha-polypeptide H+/K+ exchanging ATPase (ATP4A) and amyloid beta (A4) precursor-like protein 1 (APLP1) within the locus studied were found to be flanked by insulators on both sides. Both genes are characterized by distinct tissue-specific expression that differs from the tissue specificity of the surrounding genes. The data obtained are consistent with the conception that insulators subdivide genomic DNA into loop domains that comprise genes characterized by similar expression profiles. Using chromatin immunoprecipitation assay, we demonstrated also that at least six of the putative insulators revealed in this work could bind the CTCF transcription factor in vivo. We believe that the proposed approach could be a useful instrument for functional analysis of genomes.

    ID:13
  13. Fushan A., Monastyrskaya G., Abaev I., Sverdlov E. (2006). Genomic fingerprinting of Burkholderia pseudomallei and B. mallei pathogens with DNA array based on interspecies sequence differences obtained by subtractive hybridization. Res. Microbiol. 157 (7), 684–92 [+]

    The ability to rapidly and efficiently identify causative agents of dangerous human and animal diseases is a prerequisite to diagnosis, prophylaxis and therapy. Such identification systems can be developed based on DNA markers enabling differentiation between various bacterial strains. One source of these markers is genetic polymorphism. An efficient method for detecting the most stable polymorphisms without knowledge of genomic sequences is subtractive hybridization. In this work we report an approach to typing of Burkholderia pseudomallei and B. mallei that cause melioidosis and glanders, respectively. Typing is based on hybridization of bacterial genomes with a DNA array of genomic markers obtained using subtractive hybridization. The array comprised 55 DNA fragments which distinguished the genomes of B. pseudomallei C-141 and B. mallei C-5 strains, and it was used to test 28 radioactively labeled B. pseudomallei strains and 8 B. mallei strains. Each strain was characterized by a specific hybridization pattern, and the results were analyzed using cluster analysis. 18 patterns specific to B. pseudomallei and 6 patterns specific to B. mallei were found to be unique. The data allowed us to differentiate most studied B. pseudomallei variants from one another and from B. mallei strains. It was concluded that DNA markers obtained by subtractive hybridization can be potentially useful for molecular typing of B. pseudomallei and B. mallei strains, as well as for their molecular diagnosis. The method reported can be easily adapted for use both with DNA arrays and DNA microarrays with fluorescent probes.

    ID:12
  14. Vetchinova A.S., Akopov S.B., Chernov I.P., Nikolaev L.G., Sverdlov E.D. (2006). Two-dimensional electrophoretic mobility shift assay: identification and mapping of transcription factor CTCF target sequences within an FXYD5-COX7A1 region of human chromosome 19. Anal. Biochem. 354 (1), 85–93 [+]

    An approach for fast identification and mapping of transcription factor binding sites within long genomic sequences is proposed. Using this approach, 10 CCCTC-binding factor (CTCF) binding sites were identified within a 1-Mb FXYD5-COX7A1 human chromosome 19 region. In vivo binding of CTCF to these sites was verified by chromatin immunoprecipitation assay. CTCF binding sites were mapped within gene introns and intergenic regions, and some of them contained Alu-like repeated elements.

    ID:9
  15. Azhikina T., Gainetdinov I., Skvortsova Y., Sverdlov E. (2006). Methylation-free site patterns along a 1-Mb locus on Chr19 in cancerous and normal cells are similar. A new fast approach for analyzing unmethylated CCGG sites distribution. Mol. Genet. Genomics 275 (6), 615–22 [+]

    We describe a newly developed technique for rapid identification of positions of genomic DNA breaks, preexisting or introduced by specific digestion, in particular, by restriction endonucleases (RIDGES). We applied RIDGES in analyzing unmethylated CCGG sites distribution along a 1-Mb long genome region (D19S208-COX7A1 on chromosome 19) in cancerous and normal lung tissues. Both tissues were characterized by a profoundly uneven density of unmethylated sites along the fragment. Interestingly, the distribution of hypomethylated regions did not correlate with gene locations within the fragment, and one of the most hypomethylated areas contained practically no genes. We also demonstrated that the methylation pattern of a long genome DNA fragment was rather stable and practically unchanged in human lung cancer tissue as compared with its normal counterpart, in accordance with the suggestion (Ross et al. in Nat Genet 24:227-235, 2000) that cell lines of common origin have typically similar transcription profiles. An analogous suggestion might probably be made for global methylation patterns of genomic DNA.

    ID:10
  16. Azhikina T., Gvozdevsky N., Botvinnik A., Fushan A., Shemyakin I., Stepanshina V., Lipin M., Barry C. 3rd, Sverdlov E. (2006). A genome-wide sequence-independent comparative analysis of insertion-deletion polymorphisms in multiple Mycobacterium tuberculosis strains. Res. Microbiol. 157 (3), 282–90 [+]

    We applied an enhanced version of subtractive hybridization for comparative analyses of indel differences between genomes of several Mycobacterium tuberculosis strains widespread in Russian regions, and the H37Rv reference strain. A number of differences were detected and partially analyzed, thus demonstrating the practicality of the approach. A majority of the insertions found were shared by all Russian strains, except for strain 1540 that revealed the highest virulence in animal tests. This strain possesses a number of genes absent from other clinical strains. Two of the differential genes were found to encode putative membrane proteins and are presumed to affect mycobacterial interaction with the host cell, thus enhancing virulent properties of the isolate. The method used is of general application, and enables the elaboration of a catalogue of indel polymorphic genomic differences between closely related strains.

    ID:8
  17. Kovalskaya E., Buzdin A., Gogvadze E., Vinogradova T., Sverdlov E. (2006). Functional human endogenous retroviral LTR transcription start sites are located between the R and U5 regions. Virology 346 (2), 373–8 [+]

    Human endogenous retroviruses (HERVs) occupy about 5% of human DNA and are thought to be remnants of ancient retroviral infections of human ancestors' germ cells. HERVs can modify expression of host cell genes through their cis-regulatory elements concentrated in their long terminal repeats (LTRs). Although numerous HERV-related RNAs were identified in the human transcriptome, for most of them, it remains unclear whether they are LTR-promoted or read-through products initiated from neighboring genomic promoters. Here, we describe mapping of transcriptional start sites within solitary and proviral LTRs of the HERV-K (HML-2) human-specific subfamily of endogenous retroviruses. Surprisingly, the transcription was initiated predominantly from the very 3' termini of the LTR R regions. The data presented here may shed light on adaptive coevolution of human endogenous retroviruses with their host cells.

    ID:7
  18. Buzdin A., Kovalskaya-Alexandrova E., Gogvadze E., Sverdlov E. (2006). GREM, a technique for genome-wide isolation and quantitative analysis of promoter active repeats. Nucleic Acids Res. 34 (9), e67 [+]

    В статье опубликован новый метод, позволяющий проводить мониторинг транскрипционной активности повторяющихся элементов генома на полногеномном уровне

    ID:15
  19. Mamedov I., Batrak A., Buzdin A., Arzumanyan E., Lebedev Y., Sverdlov E.D. (2002). Genome-wide comparison of differences in the integration sites of interspersed repeats between closely related genomes. Nucleic Acids Res. 30 (14), e71 [+]

    Представлен окончательный вариант экспериментального подхода к полногеномному сравнительному анализу распределения участков интеграции LTR-элементов в геномах близкородственных видов. Впервые идентифицировано и предварительно охарактеризовано 11 инсерций LTR-элементов, отличающих геном человека от генома шимпанзе.

    ID:18
  20. Buzdin A., Khodosevich K., Mamedov I., Vinogradova T., Lebedev Y., Hunsmann G., Sverdlov E. (2002). A technique for genome-wide identification of differences in the interspersed repeats integrations between closely related genomes and its application to detection of human-specific integrations of HERV-K LTRs. Genomics 79 (3), 413–22 [+]

    В статье опубликован новый метод, позволяющий сравнивать распределение геномных повторов между близкородственными геномами при помощи метода вычитающей гибридизации

    ID:17
  21. Sverdlov E.D. (2000). Retroviruses and primate evolution. BioEssays 22 (2), 161–171 [+]

    Human endogenous retroviruses (HERVs), probably representing footprints of ancient germ-cell retroviral infections, occupy about 1% of the human genome. HERVs can influence genome regulation through expression of retroviral genes, either via genomic rearrangements following HERV integrations or through the involvement of HERV LTRs in the regulation of gene expression. Some HERVs emerged in the genome over 30 MYr ago, while others have appeared rather recently, at about the time of hominid and ape lineages divergence. HERVs might have conferred antiviral resistance on early human ancestors, thus helping them to survive. Furthermore, newly integrated HERVs could have changed the pattern of gene expression and therefore played a significant role in the evolution and divergence of Hominoidea superfamily. Comparative analysis of HERVs, HERV LTRs, neighboring genes, and their regulatory interplay in the human and ape genomes will help us to understand the possible impact of HERVs on evolution and genome regulation in the primates.

    ID:19

Свердлов Евгений Давидович

  • Москва, ул. Миклухо-Маклая, 16/10 — На карте
  • ИБХ РАН, корп. 52, комн. 657
  • Тел.: +7(495)330-65-29
  • Эл. почта: sverd@ibch.ru