Eugeny D. Sverdlov

Personal information

Career summary

1962–1965: graduate of Chemical Department of Lomonosov Moscow State University
1965–1988: researcher, leader of scientific group, head of laboratory in Shemyakin Institute of Bioorganic Chemistry of Russian Academy of Sciences
1988–2006: director of Institute of Molecular Genetics of Russian Academy of Sciences
2006–present: scientific councilor of Russian Academy of Sciences; scientific councilor of Institute of Molecular Genetics, Russian Academy of Sciences
1996–1998: supernumerary professor in chemistry, Boston University, USA

Teaching activity

1984–1987: professor in chemistry, Department of Molecular and Biological Physics, Moscow Institute of Physics and Technology, Moscow
1993–present: professor in molecular genetics, Biological department, Lomonosov Moscow State University

Other

  • Supervisor of more than 70 PhD and 6 DSc theses;
  • Editor-in-chief of “Molecular genetics, microbiology and virology” journal, Moscow;
  • Member of editorial boards: “Bioorganic chemistry”, Moscow; Doklady Akademii Nauk (Proceedings of Russian Academy of Sciences), Moscow; GENE, Netherlands (till 2003); International Journal of Genome Research, USA;
  • Member of Bureau, Branch of Physical-Chemical Biology, Russian Academy of Sciences;
  • Member of European Society of Human Genetics;
  • Member of Human Genome Organization (HUGO).

Book “Organic Chemistry of Nucleic Acids” was translated into English under edition of Nobel Prize winner Lord A. Todd and became a reference book for laboratories working in the field of nucleic acids chemistry.

Education

PeriodCountry, cityEducation institutionAdditional info
1956–1961 Russia, Moscow M.V. Lomonosov Moscow State University, Chemical Department Ms in Chemistry
1962–1965 Russia, Moscow M.V. Lomonosov Moscow State University, Chemical Department PhD in Chemistry
1980 Russia, Moscow Shemyakin Institute of Bioorganic Chemistry, USSR Academy of Sciences DSc in Chemistry

Awards & honors

1981 — USSR State Prize
1984 — USSR Lenin Prize
1984 — Order “Red Banner of Labor”
1999 — Order of Honor
2009 — Order “For merits to the State” of 4th degree

Main scientific results

Main research is devoted to development of methods of study of genetic material, to analysis of structure and functions of genes, to development of theoretical basis of modern biotechnology and production of biotechnological materials for medicine and agriculture. Author of more than 300 publications.

1967—1985. Series of investigations devoted to development of methods of nucleic acids modifications and chemical approaches to determine their sequence, secondary structure and functional properties were performed. Many chemical processes that were developed in that time are used routinely in laboratory practice of the world. Fundamental principle of nucleic acids sequencing was introduced and experimentally proved in 1972—1973 (Sverdlovet al. FEBS Letters, 28, 231, 1972; FEBS Letters 33, 15, 1973). This principle is lying now in the basis of modern methods of nucleic acids sequencing.

1980—1991. Series of research works on structure and functions of bacterial RNA polymerases were performed in collaboration with laboratories of Yu.A. Ovchinnikov and R.B. Hesin. Primary structures of Е. coli, S. typhimurium и Pseudomonas putida RNA polymerases were determined for the first time. Mutations giving rise to resistance of microorganisms to rifampicin and its derivatives were discovered frist. So, nature of M. tuberculosis resistance to this antibiotic that cause severe difficulties in treatment of tuberculosis was revealed. Methods of affin modification of protein complexes with nucleic acids were developed during these researches. These methods are acknowledged all over the world and are widely used up to now, they are referred to in principal papers and reviews of the field. These investigations were awarded to State award of the USSR. RNA polymerases were determined for the first time. Mutations giving rise to resistance of microorganisms to rifampicin and its derivatives were discovered frist. So, nature of M. tuberculosis resistance to this antibiotic that cause severe difficulties in treatment of tuberculosis was revealed. Methods of affin modification of protein complexes with nucleic acids were developed during these researches. These methods are acknowledged all over the world and are widely used up to now, they are referred to in principal papers and reviews of the field. These investigations were awarded to State award of the USSR.

1986—1991. Series of investigations on structure and functions of Na, K-ATPases were performed. These enzymes are most important components of ions transport systems in animal and human cells, their damage lead to cardiovascular disorders. Structures of subunits and genes of ATPases from pig and human were determined for the first time, as well as existence of gene families coding these subunits were revealed and features of their expression in different tissues were described first. Models of functional organization of ATPase in cells were constructed.

Several works in biotechnology resulted in particular in isolation of human interferons genes and creation of recombinant bacterial strains producting these bioregulators that are important for medicine and agriculture were carried out during the same period. These works were performed in collaboration with Institute of Genetics and Selection of Industrial Microorganisms (GosNIIGenetika) and were awarded to Lenin Prize. Preparation of recombinant human interferon is used in clinics up to now.

1992—2009. Methods of wide-ranging comparative analysis of genomes of human and animals as well as their microbial pathogenes were developed. These methods are of great importance for understanding of genetic basis of pathologies and of principles of evolutional and populational genetics. A technology of suppressive subtractive hybridization is most acknowledged of them in the world. The technology allows revealing of differences between expression profiles of different genomes and tissues including tumour and normal tissues. Researches on endogenous proviruses and peculiarities of their structures and probable pathological consequences of their activity in genomes of host cells are performed at the same time. Ways of simultaneous analysis of all genomic retroviruses were designed. Wide-ranging investigation of their expression in different tissues, in tumor and normal cells is performed in present time. Series of methods allowing identification of most important functional elements of genomes — regulators of transcription, enhancers, insulators, binding sites of transcriptional factors have been developed. Identification of these elements is necessary for understanding of function of genome as integral system in processes of vital activity, of new functions arising in evolution and of their damage in different pathologies.

Scientific societies’ membership

Academies membership

1984 — corresponding member of USSR Academy of Sciences
1991 — member of Russian Academy of Agriculture Sciences
1997 — member of Russian Academy of Sciences
2001 — member of German Academy of Sciences Leopoldina
2001 — member of European Academy of Sciences

Selected publications

  1. Kashkin K.N., Chernov I.P., Didych D.A., Sverdlov E.D. (2017). Construction of a combinatorial library of chimeric tumor-specific promoters. BioTechniques 63 (3), 107–116 [+]

    Gene therapy is a fast-developing field of molecular medicine. New, effective, and cancer-specific promoters are in high demand by researchers seeking to treat cancer through expression of therapeutic genes. Here, we created a combinatorial library of tumor-specific chimeric promoter modules for identifying new promoters with desired functions. The library was constructed by randomly combining promoter fragments from eight human genes involved in cell proliferation control. The pool of chimeric promoters was inserted into a lentiviral expression vector upstream of the CopGFP reporter gene, transduced into A431 cells, and enriched for active promoters by cell sorting. The enriched library contained a remarkably high proportion of active and tumor-specific promoters. This approach to generating combinatorial libraries of chimeric promoters may serve as a useful tool for selecting highly specific and effective promoters for cancer research and gene therapy.

    ID:2001
  2. Kashkin K., Chernov I., Stukacheva E., Monastyrskaya G., Uspenskaya N., Kopantzev E., Sverdlov E. (2015). Cancer specificity of promoters of the genes controlling cell proliferation. J. Cell. Biochem. 116 (2), 299–309 [+]

    Violation of proliferation control is a common feature of cancer cells. We put forward the hypothesis that promoters of genes involved in the control of cell proliferation should possess intrinsic cancer specific activity. We cloned promoter regions of CDC6, POLD1, CKS1B, MCM2, and PLK1 genes into pGL3 reporter vector and studied their ability to drive heterologous gene expression in transfected cancer cells of different origin and in normal human fibroblasts. Each promoter was cloned in short (335-800 bp) and long (up to 2.3 kb) variants to cover probable location of core and whole promoter regulatory elements. Cloned promoters were significantly more active in cancer cells than in normal fibroblasts that may indicate their cancer specificity. Both versions of CDC6 promoters were shown to be most active while the activities of others were close to that of BIRC5 gene (survivin) gene promoter. Long and short variants of each cloned promoter demonstrated very similar cancer specificity with the exception of PLK1-long promoter that was substantially more specific than its short variant and other promoters under study. The data indicate that most of the important cis-regulatory transcription elements responsible for intrinsic cancer specificity are located in short variants of the promoters under study. CDC6 short promoter may serve as a promising candidate for transcription targeted cancer gene therapy.

    ID:2000
  3. Alekseenko I.V., Pleshkan V.V., Kopantzev E.P., Stukacheva E.A., Chernov I.P., Vinogradova T.V., Sverdlov E.D. (2012). Activity of the upstream component of tandem TERT/survivin promoters depends on features of the downstream component. PLoS ONE 7 (10), e46474 [+]

    We spliced the promoters of the human telomerase and human survivin genes (PhTERT and PhSurv, respectively) widely used for gene therapy and known to have the broadest cancer type spectrum of activity. Two head-to-tail constructs were obtained: the PhTERT-PhSurv and PhSurv-PhTERT tandems. The splicing caused quantitative and qualitative changes in the promoter features. In both constructs, only the promoter proximal to the transcribed gene retained its ability to initiate transcription, whereas the distal promoter was silent, the phenomenon never reported before. However, the distal promoter modulated the activity of the proximal one by increasing its strength and causing an appearance of additional transcription start sites. We suggested that this suppression might be due to the presence of Sp1 transcription factor binding sites in both promoters and Sp1-bridges between these sites. Such Sp1-bridges might convert the tandem promoter linear DNA into a stem-loop structure. If localized inside the formed loop, the distal promoter could lose its ability to initiate transcription. To test this hypothesis, we constructed two modified double promoters, where the proximal PhSurv promoter was replaced either by a shortened variant of the survivin promoter (PhSurv269) or by the mouse survivin promoter. Both PhSurv substitutes were considerably shorter than PhSurv and had different numbers and/or positions of Sp1 sites. In modified tandems, transcription was initiated from both promoters. We also prepared two mutant forms of the PhSurv-PhTERT tandem with two or four Sp1 sites removed from the distal "long" PhSurv promoter. In the first case, the distal PhSurv promoter remained silent, whereas the removal of four Sp1 binding sites restored its activity. In the majority of studied cancer cell lines the efficiency of transcription from the hTERT-(shortened hSurv269) promoter tandem was markedly higher than from each constituent promoter. In normal lung fibroblast cells, the tandem promoter activity was considerably lower.

    ID:2002
  4. 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
  5. 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
  6. 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
  7. Lebedev Y.B., Amosova A.L., Mamedov I.Z., Fisunov G.Y., Sverdlov E.D. (2007). Most recent AluY insertions in human gene introns reduce the content of the primary transcripts in a cell type specific manner. Gene 390 (1-2), 122–9 [+]

    New strategy for large-scale functional analysis of polymorphic retroelements is described. Inhibitory effect of dimorphic Alu on human gene expression was discovered. Cell type specificity of the inhibitory effect was observed in heterozygous cell lines.

    ID:100
  8. 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
  9. 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 [+]

    We report the first genome-wide comparison of in vivo promoter activities of a group of human-specific endogenous retroviruses in healthy and cancerous germ line tissues. To this end, we employed a recently developed technique termed genomic repeat expression monitoring. We found that at least 50% of human-specific long terminal repeats (LTRs) possessed promoter activity, and many of them were up- or downregulated in a seminoma. Individual LTRs were expressed at markedly different levels, ranging from approximately 0.001 to approximately 3% of the housekeeping beta-actin gene transcript level. We demonstrated that the main factors affecting the LTR promoter activity were the LTR type (5'-proviral, 3' proviral, or solitary) and position with regard to genes. The averaged promoter strengths of solitary and 3'-proviral LTRs were almost identical in both tissues, whereas 5'-proviral LTRs displayed two- to fivefold higher promoter activities. The relative content of promoter-active LTRs in gene-rich regions was significantly higher than that in gene-poor loci. This content was maximal in those regions where LTRs "overlapped" readthrough transcripts. Although many promoter-active LTRs were mapped near known genes, no clear-cut correlation was observed between transcriptional activities of genes and neighboring LTRs. Our data also suggest a selective suppression of transcription for LTRs located in gene introns.

    ID:16
  10. 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
  11. 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
  12. 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
  13. Chernov I.P., Akopov S.B., Nikolaev L.G., Sverdlov E.D. (2006). Identification and mapping of DNA binding proteins target sequences in long genomic regions by two-dimensional EMSA. BioTechniques 41 (1), 91–6 [+]

    Specific binding of nuclear proteins, in particular transcription factors, to target DNA sequences is a major mechanism of genome functioning and gene expression regulation in eukaryotes. Therefore, identification and mapping specific protein target sites (PTS) is necessary for understanding genomic regulation. Here we used a novel two-dimensional electrophoretic mobility shift assay (2D-EMSA) procedure for identification and mapping of 52 PTS within a 563-kb human genome region located between the FXYD5 and TZFP genes. The PTS occurred with approximately equal frequency within unique and repetitive genomic regions. PTS belonging to unique sequences tended to group together within gene introns and close to their 5' and 3' ends, whereas PTS located within repeats were evenly distributed between transcribed and intragenic regions.

    ID:14
  14. 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
  15. 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
  16. 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
  17. 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 [+]

    We developed a technique called GREM (Genomic Repeat Expression Monitor) that can be applied to genome-wide isolation and quantitative analysis of any kind of transcriptionally active repetitive elements. Briefly, the technique includes three major stages: (i) generation of a transcriptome wide library of cDNA 5' terminal fragments, (ii) selective amplification of repeat-flanking genomic loci and (iii) hybridization of the cDNA library (i) to the amplicon (ii) with subsequent selective amplification and cloning of the cDNA-genome hybrids. The sequences obtained serve as 'tags' for promoter active repetitive elements. The advantage of GREM is an unambiguous mapping of individual promoter active repeats at a genome-wide level. We applied GREM for genome-wide experimental identification of human-specific endogenous retroviruses and their solitary long terminal repeats (LTRs) acting in vivo as promoters. Importantly, GREM tag frequencies linearly correlated with the corresponding LTR-driven transcript levels found using RT-PCR. The GREM technique enabled us to identify 54 new functional human promoters created by retroviral LTRs.

    ID:15
  18. Mamedov I.Z., Arzumanyan E.S., Amosova A.L., Lebedev Y.B., Sverdlov E.D. (2005). Whole-genome experimental identification of insertion/deletion polymorphisms of interspersed repeats by a new general approach. Nucleic Acids Res. 33 (2), e16 [+]

    A new experimental technique for genome-wide detection of integration sites of polymorphic retroelements (REs) is described. Efficiency of the technique was demonstrated by discovering large subset of dimorphic AluY elements undetectable by available bioinformatic approaches.

    ID:99
  19. Buzdin A., Gogvadze E., Kovalskaya E., Volchkov P., Ustyugova S., Illarionova A., Fushan A., Vinogradova T., Sverdlov E. (2003). The human genome contains many types of chimeric retrogenes generated through in vivo RNA recombination. Nucleic Acids Res. 31 (15), 4385–90 [+]

    L1 retrotransposons play an important role in mammalian genome shaping. In particular, they can transduce their 3'-flanking regions to new genomic loci or produce pseudogenes or retrotranscripts through reverse transcription of different kinds of cellular RNAs. Recently, we found in the human genome an unusual family of chimeric retrotranscripts composed of full-sized copies of U6 small nuclear RNAs fused at their 3' termini with 5'-truncated, 3'-poly(A)-tailed L1s. The chimeras were flanked by 11-21 bp long direct repeats, and contained near their 5' ends T2A4 hexanucleotide motifs, preferably recognized by L1 nicking endonuclease. These features suggest that the chimeras were formed using the L1 integration machinery. Here we report the identification of 81 chimeras consisting of fused DNA copies of different RNAs, including mRNAs of known human genes. Based on their structural features, the chimeras were subdivided into nine distinct families. 5' Parts of the chimeras usually originated from different nuclear RNAs, whereas their 3' parts represented cytoplasmic RNAs: mRNAs, including L1 mRNA and Alu RNA. Some of these chimeric retrotranscripts are expressed in a variety of human tissues. These findings suggest that RNA-RNA recombination during L1 reverse transcription followed by the integration of the recombinants into the host genome is a general event in genome evolution.

    ID:20
  20. Buzdin A., Ustyugova S., Gogvadze E., Lebedev Y., Hunsmann G., Sverdlov E. (2003). Genome-wide targeted search for human specific and polymorphic L1 integrations. Hum. Genet. 112 (5-6), 527–33 [+]

    An efficient application of original method of subtractive genomic DNA hybridization to identification of species-specific LINE insertions.

    ID:97
  21. Buzdin A., Ustyugova S., Khodosevich K., Mamedov I., Lebedev Y., Hunsmann G., Sverdlov E. (2003). Human-specific subfamilies of HERV-K (HML-2) long terminal repeats: three master genes were active simultaneously during branching of hominoid lineages. Genomics 81 (2), 149–56 [+]

    Complete structural analysis of human specific LTRs. Multiple endogenous retroviral retropositions in course of human speciation were confirmed at the very first time.

    ID:96
  22. 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 [+]

    New whole-genome approach to comparison of LTR distribution in genomes of related species was presented. The eleven LTR insertions distinguishing human genome from chimpanzee ones were described at the first time.

    ID:18
  23. 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 [+]

    We have developed a method of targeted genomic difference analysis (TGDA) for genomewide detection of interspersed repeat integration site differences between closely related genomes. The method includes a whole-genome amplification of the flanks adjacent to target interspersed repetitive elements in both genomic DNAs under comparison, and subtractive hybridization (SH) of the selected amplicons. The potential of TGDA was demonstrated by the detection of differences in the integration sites of human endogenous retroviruses K (HERV-K) and related solitary long terminal repeats (LTRs) between the human and chimpanzee genomes. Of 55 randomly sequenced clones from a library enriched with human-specific integration (HSI) sites, 33 (60%) represented HSIs. All the human-specific (Hs) LTRs belong to two related evolutionarily young groups, suggesting simultaneous activity of two master genes in the hominid lineage. No deletion/insertion polymorphism was detected for the LTR HSIs for 25 unrelated caucasoid individuals. We also discuss the possible research applications for TGDA research.

    ID:17
  24. 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
  25. Lebedev Y.B., Belonovitch O.S., Zybrova N.V., Khil P.P., Kurdyukov S.G., Vinogradova T.V., Hunsmann G., Sverdlov E.D. (2000). Differences in HERV-K LTR insertions in orthologous loci of humans and great apes. Gene 247 (1-2), 265–77 [+]

    The classification of the long terminal repeats (LTRs) of the human endogenous retrovirus HERV-K (HML-2) family was refined according to diagnostic differences between the LTR sequences. The mutation rate was estimated to be approximately equal for LTRs belonging to different families and branches of human endogenous retroviruses (HERVs). An average mutation rate value was calculated based on differences between LTRs of the same HERV and was found to be 0.13% per million years (Myr). Using this value, the ages of different LTR groups belonging to the LTR HML-2 subfamily were found to vary from 3 to 50Myr. Orthologous potential LTR-containing loci from different primate species were PCR amplified using primers corresponding to the genomic sequences flanking LTR integration sites. This allowed us to calculate the phylogenetic times of LTR integrations in primate lineages in the course of the evolution and to demonstrate that they are in good agreement with the LTR ages calculated from the mutation rates. Human-specific integrations for some very young LTRs were demonstrated. The possibility of LTRs and HERVs involvement in the evolution of primates is discussed.

    ID:98
  26. Ovchinnikov Yu.A., Monastyrskaya G.S., Gubanov V.V., Guryev S.O., Salomatina I.S., Shuvaeva T.M., Lipkin V.M., Sverdlov E.D. (1982). The primary structure of E. coli RNA polymerase, Nucleotide sequence of the rpoC gene and amino acid sequence of the beta'-subunit. Nucleic Acids Res. 10 (13), 4035–44 [+]

    The primary structure of the E. coli rpoC gene (5321 base pairs) coding the beta'-subunit of RNA polymerase as well as its adjacent segment have been determined. The structure analysis of the peptides obtained by cleavage of the protein with cyanogen bromide and trypsin has confirmed the amino acid sequence of the beta'-subunit deduced from the nucleotide sequence analysis. The beta'-subunit of E. coli RNA polymerase contains 1407 amino acid residues. Its translation is initiated by codon GUG and terminated by codon TAA. It has been detected that the sequence following the terminating codon is strikingly homologous to known sequences of rho-independent terminators.

    ID:54