Anton A. Buzdin

Selected publications

  1. Suntsova M., Gogvadze E.V., Salozhin S., Gaifullin N., Eroshkin F., Dmitriev S.E., Martynova N., Kulikov K., Malakhova G., Tukhbatova G., Bolshakov A.P., Ghilarov D., Garazha A., Aliper A., Cantor C.R., Solokhin Y., Roumiantsev S., Balaban P., Zhavoronkov A., Buzdin A. (2013). Human-specific endogenous retroviral insert serves as an enhancer for the schizophrenia-linked gene PRODH. Proc. Natl. Acad. Sci. U.S.A. , [+]

    Using a systematic, whole-genome analysis of enhancer activity of human-specific endogenous retroviral inserts (hsERVs), we identified an element, hsERVPRODH, that acts as a tissue-specific enhancer for the PRODH gene, which is required for proper CNS functioning. PRODH is one of the candidate genes for susceptibility to schizophrenia and other neurological disorders. It codes for a proline dehydrogenase enzyme, which catalyses the first step of proline catabolism and most likely is involved in neuromediator synthesis in the CNS. We investigated the mechanisms that regulate hsERVPRODH enhancer activity. We showed that the hsERVPRODH enhancer and the internal CpG island of PRODH synergistically activate its promoter. The enhancer activity of hsERVPRODH is regulated by methylation, and in an undermethylated state it can up-regulate PRODH expression in the hippocampus. The mechanism of hsERVPRODH enhancer activity involves the binding of the transcription factor SOX2, whch is preferentially expressed in hippocampus. We propose that the interaction of hsERVPRODH and PRODH may have contributed to human CNS evolution.

    ID:920
  2. Mityaev M.V., Kopantzev E.P., Buzdin A.A., Vinogradova T.V., Sverdlov E.D. (2010). Enhancer element potentially involved in human survivin gene promoter regulation in lung cancer cell lines. Biochemistry Mosc. 75 (2), 182–91 [+]

    We have revealed evolutionarily conserved regions in a 4500-bp DNA sequence 5'-adjacent to the survivin (BIRC5) gene. In the transcribed region of the BIRC5 gene we have detected and characterized in detail a 3'-fragment of the CpG island that stimulated in enhancer-like way the gene promoter activity in normal cells and in a number of cancer, in particular lung cancer, cell lines. When added to the initial 1498-bp survivin promoter region, a transcribed DNA fragment of a CpG island approximately twofold enhanced the promoter activity in cancer cells and in normal lung fibroblasts. The observed effect did not depend upon the orientation of the fragment and distances between the fragment and the transcription initiation site. In the case of a heterologous SV40 virus promoter, the effect was less pronounced. In addition to earlier reports, the results provide new information on the BIRC5 gene expression regulation and also demonstrate that this gene exon sequences can play a significant role in BIRC5 gene expression regulation. The data provide another possibility to increase survivin promoter activity without changing its cancer specificity for application in cancer (in particular, lung cancer) gene therapy.

    ID:489
  3. Schumann G.G., Gogvadze E.V., OsanaiFutahashi M., Kuroki A., Münk C., Fujiwara H., Ivics Z., Buzdin A.A. (2010). Unique functions of repetitive transcriptomes. Int Rev Cell Mol Biol 285, 115–88 [+]

    Repetitive sequences occupy a huge fraction of essentially every eukaryotic genome. Repetitive sequences cover more than 50% of mammalian genomic DNAs, whereas gene exons and protein-coding sequences occupy only ~3% and 1%, respectively. Numerous genomic repeats include genes themselves. They generally encode "selfish" proteins necessary for the proliferation of transposable elements (TEs) in the host genome. The major part of evolutionary "older" TEs accumulated mutations over time and fails to encode functional proteins. However, repeats have important functions also on the RNA level. Repetitive transcripts may serve as multifunctional RNAs by participating in the antisense regulation of gene activity and by competing with the host-encoded transcripts for cellular factors. In addition, genomic repeats include regulatory sequences like promoters, enhancers, splice sites, polyadenylation signals, and insulators, which actively reshape cellular transcriptomes. TE expression is tightly controlled by the host cells, and some mechanisms of this regulation were recently decoded. Finally, capacity of TEs to proliferate in the host genome led to the development of multiple biotechnological applications.

    ID:486
  4. Kuzmin D., Gogvadze E., Kholodenko R., Grzela D.P., Mityaev M., Vinogradova T., Kopantzev E., Malakhova G., Suntsova M., Sokov D., Ivics Z., Buzdin A. (2010). Novel strong tissue specific promoter for gene expression in human germ cells. BMC Biotechnol. 10, 58 [+]

    Tissue specific promoters may be utilized for a variety of applications, including programmed gene expression in cell types, tissues and organs of interest, for developing different cell culture models or for use in gene therapy. We report a novel, tissue-specific promoter that was identified and engineered from the native upstream regulatory region of the human gene NDUFV1 containing an endogenous retroviral sequence.

    ID:487
  5. Bantysh O.B., Buzdin A.A. (2009). Novel family of human transposable elements formed due to fusion of the first exon of gene MAST2 with retrotransposon SVA. Biochemistry Mosc. 74 (12), 1393–9 [+]

    We identified a novel human-specific family of transposable elements that consists of fused copies of the CpG-island containing the first exon of gene MAST2 and retrotransposon SVA. We propose a mechanism for the formation of this family termed CpG-SVA, comprising 5'-transduction by an SVA insert. After the divergence of human and chimpanzee ancestor lineages, retrotransposon SVA has inserted into the first intron of gene MAST2 in the sense orientation. Due to splicing of an aberrant RNA driven by MAST2 promoter, but terminally processed using SVA polyadenylation signal, the first exon of MAST2 has fused to a spliced 3'-terminal fragment of SVA retrotransposon. The above ancestor CpG-SVA element due to retrotranspositions of its own copies has formed a novel family represented in the human genome by 76 members. Recruitment of a MAST2 CpG island was most likely beneficial to the hybrid retrotransposons because it could significantly increase retrotransposition frequency. Also, we show that human L1 reverse transcriptase adds an extra cytosine residue to the 3' terminus of the nascent first strand of cDNA.

    ID:490
  6. Gogvadze E., Buzdin A. (2009). Retroelements and their impact on genome evolution and functioning. Cell. Mol. Life Sci. 66 (23), 3727–42 [+]

    Retroelements comprise a considerable fraction of eukaryotic genomes. Since their initial discovery by Barbara McClintock in maize DNA, retroelements have been found in genomes of almost all organisms. First considered as a "junk DNA" or genomic parasites, they were shown to influence genome functioning and to promote genetic innovations. For this reason, they were suggested as an important creative force in the genome evolution and adaptation of an organism to altered environmental conditions. In this review, we summarize the up-to-date knowledge of different ways of retroelement involvement in structural and functional evolution of genes and genomes, as well as the mechanisms generated by cells to control their retrotransposition.

    ID:491
  7. Gogvadze E., Stukacheva E., Buzdin A., Sverdlov E. (2009). Human-specific modulation of transcriptional activity provided by endogenous retroviral insertions. J. Virol. 83 (12), 6098–105 [+]

    Many phenotypic differences exist between Homo sapiens and its closest relatives, chimpanzees, and these differences can arise as a result of variations in the regulation of certain genes common to these closely related species. Human-specific endogenous retroviruses (HERVs) and their solitary long terminal repeats (LTRs) are probable candidates for such a role due to the presence of regulatory elements, such as enhancers, promoters, splice sites, and polyadenylation signals. In this study we show for the first time that HERVs can participate in the specific antisense regulation of human gene expression owing to their LTR promoter activity. We found that two HERV LTRs situated in the introns of genes SLC4A8 (for sodium bicarbonate cotransporter) and IFT172 (for intraflagellar transport protein 172) in the antisense orientation serve in vivo as promoters for generating RNAs complementary to the exons of enclosing genes. The antisense transcripts formed from LTR promoter were shown to decrease the mRNA level of the corresponding genes. The human-specific regulation of these genes suggests their involvement in the evolutionary process.

    ID:493
  8. Buzdin A.A. (2009). [Functional analysis of retroviral endogenous inserts in the human genome evolution]. Bioorg. Khim. 36 (1), 38–46 [+]

    Retroelements, mobile elements produced in DNA by reverse transcription, comprise about 40% of the human genome. A small part of these elements appeared in the genome quite recently after the divergence of humans and chimpanzees had occurred. Evolutionarily young retroelements are represented by the members of four groups, SVA, Alu, L1, and the endogenous HERV-K (HML-2) virus. These retroelements could play a functional role in the course of the molecular evolution of human DNA. We comprehensively studied the contribution of human-specific endogenous viruses (hsERV) to the structural modifications and regulation of the human genome. We found that hsERV presented in 134 copies occupied about 330 000 bp of human DNA. They added to genomic sequences the copies of 50 functional retroviral genes as well as 134 potential promoters and enhancers, 50% of which are located in the regions adjacent to known genes, and 22% in gene introns. At least 67% of these elements are human-specific promoters in vivo. hsERV viruses regulate the activity of known protein-encoding genes by means of RNA interference, function as enhancers, and provide new polyadenylation signals for mRNA.

    ID:488
  9. Giliarov D.A., Sakharova T.A., Buzdin A.A. (2009). [Molecular receptors of taste agents]. Bioorg. Khim. 35 (1), 5–14 [+]

    All representatives of higher eukaryotes can probably differentially perceive nutrients and poisonous substances. Molecular mechanisms of transduction of taste information have been best studied for mammals and for the fruit fly Drosophila. Here, we consider receptor mechanisms and conjugated primary signal processes of stimulation of taste receptor cells by stimuli of various taste modalities.

    ID:492
  10. Gogvadze E.V., Buzdin A.A. (2009). [New mechanism of retrogene formation in mammalian genomes: in vivo recombination during RNA reverse transcription]. Mol. Biol. (Mosk.) 39 (3), 364–73 [+]

    L1 LINE retrotransposons play an important role in the shaping and permanent evolution of mammalian genomes. In particular, occupying about 20% of genomic DNA, they transduce their 3' flanking sequences to new genomic loci and create pseudogenes by reverse transcribing different kinds of cellular RNAs. Recently we discovered in mammalian genomes several families of chimeric pseudogenes, consisting of fused copies of various cellular transcripts. The characteristic peculiarities of such chimeric inserts suggest the involvement of L1 enzymatic machinery in their formation. The detailed sequence analysis revealed that 5' terminal parts of the chimeras are copies of nuclear RNAs, while 3' terminal sequences were formed on the templates of transcripts having cytoplasmic orientation. These data enabled us to propose the mechanism for the chimeras formation, comprising the switch of templates during RNA reverse transcription by L1 reverse transcriptase. The further identification of not only "double", but also "triple" chimeric retrogenes evidences the possibility of the double template switches as well. Some of the found chimeras are transcriptionally active, thus allowing to consider the discovered phenomenon as the new mechanism of the gene formation by "shuffling" of pre-existing transcribed sequences. Being active in mammals now, the mechanism appeared at least 75 million years ago and is evolutionary conserved.

    ID:500
  11. Gogvadze E.V., Buzdin A.A., Sverdlov E.D. (2009). [Multiple template switches on LINE-directed reverse transcription: the most probable formation mechanism for the double and triple chimeric retroelements in mammals]. Bioorg. Khim. 31 (1), 82–9 [+]

    It was shown that the shuffling mechanism for transcribed genome components, which involves a template switch on the RNA reverse transcription using the L1 retroelement enzymatic machinery, is common in mammals. The occurrence frequency of the resulting chimeric retroelements in the genomes of rodents is twice as high as in the DNA of primates. Moreover, we proved that not only single but also double switches may occur in vivo, which result in the fusion of copies of three different transcripts. Many of the identified chimeras are transcribed in mammals.

    ID:501
  12. Buzdin A.A., Lebedev Iu.B., Sverdlov E.D. (2009). [Human genome-specific HERV-K intron LTR genes have a random orientation relative to the direction of transcription, and, possibly, participated in antisense gene expression regulation]. Bioorg. Khim. 29 (1), 103–6 [+]

    A consensus nucleotide sequence of long terminal repeats (LTRs) of endogenous human-specific retroviruses of the K family (HERV-K) was constructed and used for the genome-wide search for homologies in international databases. There were revealed 142 LTRs, 12 of which were localized in introns of unique human genes. It was found for the first time that ten intron LTRs are absent in the orthologic loci of the chimpanzee genome and the orientation of nine of them is opposite to the transcription direction of the corresponding human genes. A hypothesis was propounded that the found LTRs affect the gene expression by initiation of the antisense RNA synthesis.

    ID:504
  13. Mityaev M.V., Kopantzev E.P., Buzdin A.A., Vinogradova T.V., Sverdlov E.D. (2008). Functional significance of a putative sp1 transcription factor binding site in the survivin gene promoter. Biochemistry Mosc. 73 (11), 1183–91 [+]

    We sequenced 1500-bp genomic DNA regions upstream from the survivin gene (BIRC5). DNA was isolated from human placenta and tumors of patients with diagnosed squamous cancer of the lung that showed high-level BIRC5 gene expression. We have revealed four new promoter allelic variants differing in single nucleotide substitutions, one variant with two nucleotide substitutions, and a variant with a TAAA tetranucleotide insertion. All promoter variants displayed low activity in cells with functionally active p53 protein and high activity in cell lines characterized by low level or absence of p53 protein function. The activity of the promoters with single nucleotide substitutions was comparable to that of the wild-type promoter, whereas two nucleotide substitutions markedly reduced the activity. We also demonstrated the functional significance of a putative Sp1 transcription factor-binding site at (-63...-54) upstream from the transcription initiation site. Mutation within this sequence led to a sharp decrease of promoter activity. The functional architecture of the survivin promoter is discussed based on results known from the literature and those obtained here.

    ID:494
  14. Buzdin A., Gogvadze E., Lebrun M.H. (2007). Chimeric retrogenes suggest a role for the nucleolus in LINE amplification. FEBS Lett. 581 (16), 2877–82 [+]

    Chimeric retrogenes, found in mammalian and fungal genomes, are bipartite elements composed of DNA copies of cellular transcripts either directly fused to each other or fused to the 3' part of a LINE retrotransposon. These cellular transcripts correspond to messenger RNAs, ribosomal RNAs, small nuclear RNAs and 7SL RNA. The chimeras are likely formed by RNA template switches during reverse transcription of LINE elements by their retrotranspositional machinery. The 5' part of chimeras are copies of nucleolar RNAs, suggesting that the nucleolus plays a significant role in LINE retrotransposition. RNAs from the nucleolus might have protective function against retroelement invasion or, alternatively, the nucleolus may be required for retrotranspositional complex assembly and maturation. These hypotheses will be discussed in this review.

    ID:497
  15. Kholodenko R., Kholodenko I., Sorokin V., Tolmazova A., Sazonova O., Buzdin A. (2007). Anti-apoptotic effect of retinoic acid on retinal progenitor cells mediated by a protein kinase A-dependent mechanism. Cell Res. 17 (2), 151–62 [+]

    Retinal progenitor cells (RPCs) are neural stem cells able to differentiate into any normal adult retinal cell type, except for pigment epithelial cells. Retinoic acid (RA) is a powerful growth/differentiation factor that generally causes growth inhibition, differentiation and/or apoptosis. In this study, we demonstrate that RA not only affects mouse RPC differentiation but also improves cell survival by reducing spontaneous apoptotic rate without affecting RPC proliferation. The enhanced cell survival was accompanied by a significant upregulation of the expression of protein kinase A (PKA) and several protein kinase C (PKC) isoforms. Treatment of cells grown in RA-free media with 8-bromoadenosine3',5'-cyclic monophosphate, a known activator of PKA, resulted in an anti-apoptotic effect similar to that caused by RA; whereas the PKA inhibitor N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride led to a significant ( approximately 32%) increase in apoptosis. In contrast, treatment of RPCs with any of two PKC selective inhibitors, 2,2',3,3',4,4'-hexahydroxy-1,1'-biphenyl-6,6'-dimethanol dimethyl ether and bisindolylmaleimide XI, led to diminished apoptosis; while a PKC activator, phorbol 12-myristate 13-acetate, increased apoptosis. These and other data suggest that the effect of RA on RPC survival is mostly due to the increased anti-apoptotic activity elicited by PKA, which might in turn be antagonized by PKC. Such a mechanism is a new example of tight regulation of important biological processes triggered by RA. Although the detailed mechanisms remain to be elucidated, we provide evidence that the pro-survival effect of RA on RPCs is not mediated by changed expression of p53 or bcl-2, and appears to be independent of beta-amyloid, Fas ligand, TNF-alpha, ganglioside GM1 and ceramide C16-induced apoptotic pathways.

    ID:21
  16. Buzdin A. (2007). Human-specific endogenous retroviruses. ScientificWorldJournal 7, 1848–68 [+]

    This review focuses on a small family of human-specific genomic repetitive elements, presented by 134 members that shaped approximately 330 kb of the human DNA. Although modest in terms of its copy number, this group appeared to modify the human genome activity by endogenizing approximately 50 functional copies of viral genes that may have important implications in the immune response, cancer progression, and antiretroviral host defense. A total of 134 potential promoters and enhancers have been added to the human DNA, about 50% of them in the close gene vicinity and 22% in gene introns. For 60 such human-specific promoters, their activity was confirmed by in vivo assays, with the transcriptional level varying approximately 1000-fold from hardly detectable to as high as approximately 3% of â-actin transcript level. New polyadenylation signals have been provided to four human RNAs, and a number of potential antisense regulators of known human genes appeared due to human-specific retroviral insertional activity. This information is given here in the context of other major genomic changes underlining differences between human and chimpanzee DNAs. Finally, a comprehensive database, is available for download, of human-specific and polymorphic endogenous retroviruses is presented, which encompasses the data on their genomic localization, primary structure, encoded viral genes, human gene neighborhood, transcriptional activity, and methylation status.

    ID:495
  17. Gogvadze E., Barbisan C., Lebrun M.H., Buzdin A. (2007). Tripartite chimeric pseudogene from the genome of rice blast fungus Magnaporthe grisea suggests double template jumps during long interspersed nuclear element (LINE) reverse transcription. BMC Genomics 8, 360 [+]

    A systematic survey of loci carrying retrotransposons in the genome of the rice blast fungus Magnaporthe grisea allowed the identification of novel non-canonical retropseudogenes. These elements are chimeric retrogenes composed of DNA copies from different cellular transcripts directly fused to each other. Their components are copies of a non protein-coding highly expressed RNA of unknown function termed WEIRD and of two fungal retrotransposons: MGL and Mg-SINE. Many of these chimeras are transcribed in various M. grisea tissues and during plant infection. Chimeric retroelements with a similar structure were recently found in three mammalian genomes. All these chimeras are likely formed by RNA template switches during the reverse transcription of diverse LINE elements.

    ID:496
  18. 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
  19. Kholodenko I.V., Buzdin A.A., Kholodenko R.V., Baibikova J.A., Sorokin V.F., Yarygin V.N., Sverdlov E.D. (2006). Mouse retinal progenitor cell (RPC) cocultivation with retinal pigment epithelial cell culture affects features of RPC differentiation. Biochemistry Mosc. 71 (7), 767–74 [+]

    We provide evidence that coculturing of retinal progenitor cells (RPC) with retinal pigment epithelial cells significantly biases the standard in vitro RPC differentiation patterns. In particular, in cocultivation experiments RPCs lost the ability to differentiate spontaneously and displayed approximately 2.1-2.4-fold increase in immunoreactivity to the neural stem cell marker nestin and approximately 1.6-1.7-fold increase in rod photoreceptor cell rhodopsin marker immunoreactivity. The data suggest the influence of the intercellular interaction networks on RPC differentiation.

    ID:498
  20. 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
  21. 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
  22. Buzdin A., Vinogradova T., Lebedev Y., Sverdlov E. (2005). Genome-wide experimental identification and functional analysis of human specific retroelements. Cytogenet. Genome Res. 110 (1-4), 468–74 [+]

    Retroelements (REs) actively reshape genomes through genomic rearrangements, creation of new genes and modulation of the regulatory machinery of existing genes, thus introducing genomic novelties which potentially may be subject to natural selection. Thousands of RE integrations, presumably distinguishing the human and chimpanzee genomes, might well be involved in modern human speciation. In this self-review we describe our recent results on genome-wide identification of human specific RE integrations and their transcriptional activity obtained with three new experimental techniques (TGDA, DiffIR and SDDIR) developed by us for such studies. A new mechanism of the formation of retroelements involving template switches during L1-mediated mRNA reverse transcription, revealed in this research, will also be described in the review.

    ID:499
  23. Buzdin A.A. (2004). Retroelements and formation of chimeric retrogenes. Cell. Mol. Life Sci. 61 (16), 2046–59 [+]

    It is very likely that formation of new genes is the main pathway of molecular evolution in living organisms. Many such genes are products of preexisting reshuffling of genetic material. In these processes a very important role is played by mutations associated with the activity of transposable elements, mostly retroelements (REs) for higher eukaryotes. The life cycle of REs involves a stage of so-called reverse transcription of their RNA intermediates, i.e. synthesis of complementary DNA on an RNA template. Transcriptionally active sequences of RE origin are referred to as retrogenes. REs create chimeric genes by a variety of mechanisms: new RE insertions, recombinations between RE sequences, formation of functional gene active pseudogenes and template switches during reverse transcription of messenger RNA. The abovementioned events are also able to give rise to new RE families. These mechanisms are reviewed here along with the description of major RE groups.

    ID:503
  24. Chalaya T., Gogvadze E., Buzdin A., Kovalskaya E., Sverdlov E.D. (2004). Improving specificity of DNA hybridization-based methods. Nucleic Acids Res. 32 (16), e130 [+]

    Methods based on DNA reassociation in solution with the subsequent PCR amplification of certain hybrid molecules, such as coincidence cloning and subtractive hybridization, all suffer from a common imperfection: cross-hybridization between various types of paralogous repetitive DNA fragments. Although the situation can be slightly improved by the addition of repeat-specific competitor DNA into the hybridization mixture, the cross-hybridization outcome is a significant number of background chimeric clones in resulting DNA libraries. In order to overcome this challenge, we developed a technique called mispaired DNA rejection (MDR), which utilizes a treatment of resulting reassociated DNA with mismatch-specific nucleases. We examined the MDR efficiency using cross-hybridization of complex, whole genomic mixtures derived from human and chimpanzee genomes, digested with frequent-cutter restriction enzyme. We show here that both single-stranded DNA-specific and mismatched double-stranded DNA-specific nucleases can be used for MDR separately or in combination, reducing the background level from 60 to 4% or lower. The technique presented here is of universal usefulness and can be applied to both cDNA and genomic DNA subtractions of very complex DNA mixtures. MDR is also useful for the genome-wide recovery of highly conserved DNA sequences, as we demonstrate by comparing human and pygmy marmoset genomes.

    ID:502
  25. 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
  26. 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
  27. 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
  28. Buzdin A., Ustyugova S., Gogvadze E., Vinogradova T., Lebedev Y., Sverdlov E. (2002). A new family of chimeric retrotranscripts formed by a full copy of U6 small nuclear RNA fused to the 3' terminus of l1. Genomics 80 (4), 402–6 [+]

    Long interspersed nuclear elements (LINE-1, L1) constitute a large family of mammalian retrotransposons that have been replicating and evolving in mammals for more than 100 million years and now compose 17% of the human genome. They have an important creative role in human genomic evolution through mechanisms such as new integrations, generation of processed pseudogenes, and transfer of non-L1 DNA flanking their 3' ends to new genomic locations. Here we present evidence that the L1 integration machinery was used for the creation of a new family of chimeric retrotranscripts, which contain a full copy of U6 small nuclear RNA and a 3' part of L1 at their 5' and 3' ends, respectively. There are at least 56 members of this family in the human genome. The integrations of such fused retrotranscripts into the human genome took place until recently. Here we report one U6-L1 insertion that is polymorphic in humans. We also propose a mechanism used to generate chimeric retrotranscripts.

    ID:505
  29. 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
  30. Buzdin A.A., Revina L.P., Kostina L.I., Zalunin I.A., Chestukhina G.G. (2002). Interaction of 65- and 62-kD proteins from the apical membranes of the Aedes aegypti larvae midgut epithelium with Cry4B and Cry11A endotoxins of Bacillus thuringiensis. Biochemistry Mosc. 67 (5), 540–6 [+]

    A protein with the molecular weight of 65 kD is the only component of Aedes aegypti larvae BBM capable to specifically bind mosquitocidal toxins Cry4B and Cry11A of Bacillus thuringiensis. This protein lacks the leucine aminopeptidase activity which is characteristic for the toxin-binding proteins from the membranes of caterpillars. Cry-toxins inactive against A. aegypti larvae either fail to bind to the 65-kD protein and to a putative product of its proteolysis with the molecular weight of 62 kD (Cry1Ab), or bind but do not compete for this binding with mosquitocidal proteins (Cry9A). The proteolytic splitting out of the first five alpha-helices in the Cry4B toxin molecule does not affect its binding to the 65- and 62-kD proteins, but an additional removal of 20-30 amino acids from the C-terminal of the molecule sharply spoils this binding. Monosaccharide residues are not involved in the binding of the 65- and 62-kD proteins with Cry4B, Cry11A, and Cry9A.

    ID:506
  31. 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
  32. Krieger I.V., Revina L.P., Kostina L.I., Buzdin A.A., Zalunin I.A., Chestukhina G.G., Stepanov V.M. (1999). Membrane proteins of Aedes aegypti larvae bind toxins Cry4B and Cry11A of Bacillus thuringiensis ssp. israelensis. Biochemistry Mosc. 64 (10), 1163–8 [+]

    Proteins of molecular weight 65 and 62 kD and having affinity for toxins Cry4B and Cry11A produced by Bacillus thuringiensis ssp. israelensis have been isolated from brush border membranes of Aedes aegypti larvae using affinity chromatography. Using a ligand blotting technique, we show that the binding of these proteins to the biotinylated toxins is reversible and that the two toxins compete for binding to the two proteins. These proteins are likely to be Cry4B and Cry11A toxin receptors in gut epithelial cells of Aedes aegypti larvae.

    ID:507