Michael Nevels

Publications

• 7.45

  Impact points

  How to control an infectious bead string: nucleosome-based regulation and targeting of herpesvirus chromatin.

  Michael Nevels, Alexandra Nitzsche, Christina Paulus

  Reviews in medical virology. 05/2011; 21(3):154-80.

  Herpesvirus infections of humans can cause a broad variety of symptoms ranging from mild afflictions to life-threatening disease. During infection, the large double-stranded DNA genomes of all herpesviruses are transcribed, replicated and encapsidated in the host cell nucleus, where DNA is typically... [more] Herpesvirus infections of humans can cause a broad variety of symptoms ranging from mild afflictions to life-threatening disease. During infection, the large double-stranded DNA genomes of all herpesviruses are transcribed, replicated and encapsidated in the host cell nucleus, where DNA is typically structured and manoeuvred through nucleosomes. Nucleosomes individually assemble DNA around core histone octamers to form 'beads-on-a-string' chromatin fibres. Herpesviruses have responded to the advantages and challenges of chromatin formation in biologically unique ways. Although herpesvirus DNA is devoid of histones within nucleocapsids, nuclear viral genomes most likely form irregularly arranged or unstable nucleosomes during productive infection, and regular nucleosomal arrays resembling host cell chromatin in latently infected cells. Besides variations in nucleosome density, herpesvirus chromatin 'bead strings' undergo dynamic changes in histone composition and modification during the different stages of productive replication, latent infection and reactivation from latency, raising the likely possibility that epigenetic processes may dictate, at least in part, the outcome of infection and ensuing pathogenesis. Here, we summarise and discuss several new and important aspects regarding the nucleosome-based mechanisms that regulate herpesvirus chromatin structure and function in infected cells. Special emphasis is given to processes of histone deposition, histone variant exchange and covalent histone modification in relation to the transcription from the viral genome during productive and latent infections by human cytomegalovirus and herpes simplex virus type 1. We also present an overview on emerging histone-directed antiviral strategies that may be developed into 'epigenetic therapies' to improve current prevention and treatment options targeting herpesvirus infection and disease.
• 8.98

  Impact points

  Human cytomegalovirus IE1 protein elicits a type II interferon-like host cell response that depends on activated STAT1 but not interferon-γ.

  Theresa Knoblach, Benedikt Grandel, Jana Seiler, Michael Nevels, Christina Paulus

  PLoS pathogens. 04/2011; 7(4):e1002016.

  Human cytomegalovirus (hCMV) is a highly prevalent pathogen that, upon primary infection, establishes life-long persistence in all infected individuals. Acute hCMV infections cause a variety of diseases in humans with developmental or acquired immune deficits. In addition, persistent hCMV infection ... [more] Human cytomegalovirus (hCMV) is a highly prevalent pathogen that, upon primary infection, establishes life-long persistence in all infected individuals. Acute hCMV infections cause a variety of diseases in humans with developmental or acquired immune deficits. In addition, persistent hCMV infection may contribute to various chronic disease conditions even in immunologically normal people. The pathogenesis of hCMV disease has been frequently linked to inflammatory host immune responses triggered by virus-infected cells. Moreover, hCMV infection activates numerous host genes many of which encode pro-inflammatory proteins. However, little is known about the relative contributions of individual viral gene products to these changes in cellular transcription. We systematically analyzed the effects of the hCMV 72-kDa immediate-early 1 (IE1) protein, a major transcriptional activator and antagonist of type I interferon (IFN) signaling, on the human transcriptome. Following expression under conditions closely mimicking the situation during productive infection, IE1 elicits a global type II IFN-like host cell response. This response is dominated by the selective up-regulation of immune stimulatory genes normally controlled by IFN-γ and includes the synthesis and secretion of pro-inflammatory chemokines. IE1-mediated induction of IFN-stimulated genes strictly depends on tyrosine-phosphorylated signal transducer and activator of transcription 1 (STAT1) and correlates with the nuclear accumulation and sequence-specific binding of STAT1 to IFN-γ-responsive promoters. However, neither synthesis nor secretion of IFN-γ or other IFNs seems to be required for the IE1-dependent effects on cellular gene expression. Our results demonstrate that a single hCMV protein can trigger a pro-inflammatory host transcriptional response via an unexpected STAT1-dependent but IFN-independent mechanism and identify IE1 as a candidate determinant of hCMV pathogenicity.

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• 6.09

  Impact points

  The multi-targeted kinase inhibitor sorafenib inhibits human cytomegalovirus replication.

  Martin Michaelis, Christina Paulus, Nadine Löschmann, Stephanie Dauth, Elisabeth Stange, Hans Wilhelm Doerr, Michael Nevels, Jindrich Cinatl

  Cellular and molecular life sciences : CMLS. 03/2011; 68(6):1079-90.

  Human cytomegalovirus (HCMV) is a major pathogen in immunocompromised individuals. Here, non-toxic concentrations of the anti-cancer kinase inhibitor sorafenib were shown to inhibit replication of different HCMV strains (including a ganciclovir-resistant strain) in different cell types. In contrast ... [more] Human cytomegalovirus (HCMV) is a major pathogen in immunocompromised individuals. Here, non-toxic concentrations of the anti-cancer kinase inhibitor sorafenib were shown to inhibit replication of different HCMV strains (including a ganciclovir-resistant strain) in different cell types. In contrast to established anti-HCMV drugs, sorafenib inhibited HCMV major immediate early promoter activity and HCMV immediate early antigen (IEA) expression. Sorafenib is known to inhibit Raf. Comparison of sorafenib with the MEK inhibitor U0126 suggested that sorafenib inhibits HCMV IEA expression through inhibition of Raf but independently of signaling through the Raf downstream kinase MEK 1/2. In concordance, siRNA-mediated depletion of Raf but not of MEK-reduced IEA expression. In conclusion, sorafenib diminished HCMV replication in clinically relevant concentrations and inhibited HCMV IEA expression, a pathophysiologically relevant event that is not affected by established anti-HCMV drugs. Moreover, we demonstrated for the first time that Raf activation is involved in HCMV IEA expression.

• Adenoviren - Grundlagen

  Timo Sieber, Michael Nevels, Thomas Dobner

  01/2010: pages 639-646;

  ISBN: 978-3-13-113962-7

• The human cytomegalovirus major immediate-early proteins as antagonists of intrinsic and innate antiviral host responses.

  Christina Paulus, Michael Nevels

  Viruses. 12/2009; 1(3):760-79.

  The major immediate-early (IE) gene of human cytomegalovirus (CMV) is believed to have a decisive role in acute infection and its activity is an important indicator of viral reactivation from latency. Although a variety of gene products are expressed from this region, the 72-kDa IE1 and the 86-kDa I... [more] The major immediate-early (IE) gene of human cytomegalovirus (CMV) is believed to have a decisive role in acute infection and its activity is an important indicator of viral reactivation from latency. Although a variety of gene products are expressed from this region, the 72-kDa IE1 and the 86-kDa IE2 nuclear phosphoproteins are the most abundant and important. Both proteins have long been recognized as promiscuous transcriptional regulators. More recently, a critical role of the IE1 and IE2 proteins in counteracting non-adaptive host cell defense mechanisms has been revealed. In this review we will briefly summarize the available literature on IE1- and IE2-dependent mechanisms contributing to CMV evasion from intrinsic and innate immune responses.

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• 7.45

  Impact points

  Chromatinisation of herpesvirus genomes.

  Christina Paulus, Alexandra Nitzsche, Michael Nevels

  Reviews in medical virology. 11/2009;

  The double-stranded DNA genomes of herpesviruses exist in at least three alternative global chromatin states characterised by distinct nucleosome content. When encapsidated in virus particles, the viral DNA is devoid of any nucleosomes. In contrast, within latently infected nuclei herpesvirus genome... [more] The double-stranded DNA genomes of herpesviruses exist in at least three alternative global chromatin states characterised by distinct nucleosome content. When encapsidated in virus particles, the viral DNA is devoid of any nucleosomes. In contrast, within latently infected nuclei herpesvirus genomes are believed to form regular nucleosomal structures resembling cellular chromatin. Finally, during productive infection nuclear viral DNA appears to adopt a state of intermediate chromatin formation with irregularly spaced nucleosomes. Nucleosome occupancy coupled with posttranslational histone modifications and other epigenetic marks may contribute significantly to the extent and timing of transcription from the viral genome and, consequently, to the outcome of infection. Recent research has provided first insights into the viral and cellular mechanisms that either maintain individual herpesvirus chromatin states or mediate transition between them. Here, we summarise and discuss both early work and new developments pointing towards common principles pertinent to the dynamic structure and epigenetic regulation of herpesvirus chromatin. Special emphasis is given to the emerging similarities in nucleosome assembly and disassembly processes on herpes simplex virus type 1 and human cytomegalovirus genomes over the course of the viral productive replication cycle and during the switch between latent and lytic infectious stages. Copyright (c) 2009 John Wiley & Sons, Ltd.

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• 5.15

  Impact points

  Physical Requirements and Functional Consequences of Complex Formation Between the Cytomegalovirus IE1 Protein and Human STAT2.

  Steffen Krauss, Julia Kaps, Nathalie Czech, Christina Paulus, Michael Nevels

  Journal of virology. 10/2009;

  Our previous work has shown that efficient evasion from type I interferon reponses by human Cytomegalovirus (hCMV) requires expression of the 72-kDa immediate-early 1 (IE1) protein. It has been suggested that IE1 inhibits interferon signaling through intranuclear sequestration of the signal transduc... [more] Our previous work has shown that efficient evasion from type I interferon reponses by human Cytomegalovirus (hCMV) requires expression of the 72-kDa immediate-early 1 (IE1) protein. It has been suggested that IE1 inhibits interferon signaling through intranuclear sequestration of the signal transducer and activator of transcription 2 (STAT2) protein. Here we show that physical association and subnuclear co-localization between IE1 and STAT2 depend on short acidic and serine/proline-rich low complexity motifs in the carboxy-terminal region of the 491 amino acid viral polypeptide. These motifs compose an essential core (amino acids 373-420) and an adjacent ancillary site (amino acids 421 to 445) for STAT2 interaction that are predicted to form part of a natively unstructured domain. The presence of presumably "disordered" carboxy-terminal domains enriched in low complexity motifs is evolutionary highly conserved across all examined mammalian IE1 orthologs, and the murine cytomegalovirus IE1 protein appears to interact with STAT2 just like the human counterpart. A recombinant hCMV specifically mutated in the IE1 core STAT2 binding site displays hypersensitivity to interferon-alpha, delayed early viral protein accumulation, and attenuated growth in fibroblasts. However, replication of this mutant virus is specifically restored by knock-down of STAT2 expression. Interestingly, complex formation with STAT2 proved to be entirely separable from disruption of nuclear domain 10 (ND10), another key activity of IE1. Finally, our results demonstrate that IE1 counteracts the anti-viral interferon reponse and promotes viral replication by at least two distinct mechanisms, one depending on sequestration of STAT2 and the other one likely involving ND10 interaction.

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• 14.07

  Impact points

  Activation of telomerase in glioma cells by human cytomegalovirus: another piece of the puzzle.

  Jindrich Cinatl, Michael Nevels, Christina Paulus, Martin Michaelis

  Journal of the National Cancer Institute. 05/2009; 101(7):441-3.
• 5.15

  Impact points

  Temporal Dynamics of Cytomegalovirus Chromatin Assembly in Productively Infected Human Cells.

  Alexandra Nitzsche, Christina Paulus, Michael Nevels

  Journal of virology. 10/2008;

  The genomes of herpesviruses including human cytomegalovirus (CMV) are double-stranded DNA molecules maintained as episomes during infection. The viral DNA lacks histones when encapsidated in the virion. However, it has been found histone-associated inside infected cells implying unidentified chroma... [more] The genomes of herpesviruses including human cytomegalovirus (CMV) are double-stranded DNA molecules maintained as episomes during infection. The viral DNA lacks histones when encapsidated in the virion. However, it has been found histone-associated inside infected cells implying unidentified chromatin assembly mechanisms. Our results indicate that components of the host cell nucleosome deposition machinery target intranuclear CMV DNA resulting in stepwise viral chromatin assembly. CMV genomes undergo limited histone association and nucleosome assembly as early as 30 minutes after infection via DNA replication-independent mechanisms. Low average viral genome chromatinization is maintained throughout the early stages of infection. The late phase of infection is characterized by a striking increase in average histone occupancy coupled with the process of viral DNA replication. While the initial chromatinization affected all analyzed parts of the CMV chromosome, a subset of viral genomic regions, including the major immediate-early promoter, proved to be largely resistant to replication-dependent histone deposition. Finally, our results predict the likely requirement of an unanticipated chromatin disassembly process that enables packaging of histone-free DNA into progeny capsids.

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• Erreger-induzierte Tumore

  Michael Nevels, Thomas Dobner

  01/2008: pages 53-66;

  ISBN: 978-3-437-21741-8

• Determination of the transforming activities of adenovirus oncogenes.

  Michael Nevels, Thomas Dobner

  Methods in molecular medicine. 02/2007; 131:187-95.

  The last 50 yr of molecular biological investigations into human adenoviruses (Ads) have contributed enormously to our understanding of the basic principles of normal and malignant cell growth. Much of this knowledge stems from analyses of the Ad productive infection cycle in permissive host cells. ... [more] The last 50 yr of molecular biological investigations into human adenoviruses (Ads) have contributed enormously to our understanding of the basic principles of normal and malignant cell growth. Much of this knowledge stems from analyses of the Ad productive infection cycle in permissive host cells. Also, initial observations concerning the transforming potential of human Ads subsequently revealed decisive insights into the molecular mechanisms of the origins of cancer and established Ads as a model system for explaining virus-mediated transformation processes. Today it is well established that cell transformation by human Ads is a multistep process involving several gene products encoded in early transcription units 1A (E1A) and 1B (E1B). Moreover, a large body of evidence now indicates that alternative or additional mechanisms are engaged in Ad-mediated oncogenic transformation involving gene products encoded in early region 4 (E4) as well as epigenetic changes resulting from viral DNA integration. In particular, studies on the transforming potential of several E4 gene products have now revealed new pathways that point to novel general mechanisms of virus-mediated oncogenesis. In this chapter we describe in vitro and in vivo assays to determine the transforming and oncogenic activities of the E1A, E1B, and E4 oncoproteins in primary baby rat kidney cells and athymic nude mice.
• 9.43

  Impact points

  A human cytomegalovirus antagonist of type I IFN-dependent signal transducer and activator of transcription signaling.

  Christina Paulus, Steffen Krauss, Michael Nevels

  Proceedings of the National Academy of Sciences of the United States of America. 04/2006; 103(10):3840-5.

  Type I IFNs are crucial components of the innate immune response to viral attack. They are rapidly synthesized and secreted after infection with human cytomegalovirus (CMV) and trigger a signal transduction pathway that involves successive activation and nuclear translocation of signal transducer an... [more] Type I IFNs are crucial components of the innate immune response to viral attack. They are rapidly synthesized and secreted after infection with human cytomegalovirus (CMV) and trigger a signal transduction pathway that involves successive activation and nuclear translocation of signal transducer and activator of transcription 1 (STAT1) and STAT2. The activated STATs, together with the IFN regulatory factor 9 protein, form a trimeric transcription complex (IFN-stimulated gene factor 3) that stimulates expression of numerous IFN-responsive genes, many of which exhibit antiviral activity. Here we demonstrate that the viral 72-kDa IE1 protein (IE1-72kDa) confers partial resistance to the antiviral activity of type I IFNs upon CMV. Accordingly, IFN-responsive transcripts accumulate to substantially increased levels after infection with an IE1-deficient mutant as compared with wild-type virus, and ectopic expression of the viral protein in stably transfected cells is sufficient to block their induction. We further show that IE1-72kDa forms a physical complex with STAT1 and STAT2 in nuclei of infected cells and in vitro and prevents association of STAT1, STAT2, and IFN regulatory factor 9 with promoters of IFN-responsive genes in vivo. Our results indicate that the viral protein blocks an intranuclear step after nuclear translocation and before DNA binding of IFN-stimulated gene factor 3, presumably by interfering with the integrity and/or correct subnuclear localization of the protein complex. This study identifies the CMV IE1-72kDa protein as a viral antagonist of the cellular innate immune response, inhibiting IFN-dependent STAT signaling by means of an unprecedented molecular mechanism.
• 9.43

  Impact points

  Human cytomegalovirus immediate-early 1 protein facilitates viral replication by antagonizing histone deacetylation.

  Michael Nevels, Christina Paulus, Thomas Shenk

  Proceedings of the National Academy of Sciences of the United States of America. 01/2005; 101(49):17234-9.

  The human cytomegalovirus 72-kDa immediate-early (IE)1 and 86-kDa IE2 proteins are expressed at the start of infection, and they are believed to exert much of their function through promiscuous transcriptional activation of viral and cellular gene expression. Here, we show that the impaired growth o... [more] The human cytomegalovirus 72-kDa immediate-early (IE)1 and 86-kDa IE2 proteins are expressed at the start of infection, and they are believed to exert much of their function through promiscuous transcriptional activation of viral and cellular gene expression. Here, we show that the impaired growth of an IE1-deficient mutant virus in human fibroblasts is efficiently rescued by histone deacetylase (HDAC) inhibitors of three distinct chemical classes. In the absence of IE1 expression, the viral major IE and UL44 early promoters exhibited decreased de novo acetylation of histone H4 during the early phase of infection, and the hypoacetylation correlated with reduced transcription and accumulation of the respective gene products. Consistent with these findings, IE1 interacts specifically with HDAC3 within infected cells. We also demonstrate an interaction between IE2 and HDAC3. We propose that the ability to modify chromatin is fundamental to transcriptional activation by IE1 and, likely, IE2 as well.
• 5.15

  Impact points

  SUMOylation of the human cytomegalovirus 72-kilodalton IE1 protein facilitates expression of the 86-kilodalton IE2 protein and promotes viral replication.

  Michael Nevels, Wolfram Brune, Thomas Shenk

  Journal of virology. 08/2004; 78(14):7803-12.

  The 72-kDa immediate-early 1 protein (IE1-72kDa) of human cytomegalovirus has been previously shown to be posttranslationally modified by covalent conjugation to the ubiquitin-related protein SUMO-1. Using an infectious bacterial artificial chromosome clone of human cytomegalovirus, we constructed a... [more] The 72-kDa immediate-early 1 protein (IE1-72kDa) of human cytomegalovirus has been previously shown to be posttranslationally modified by covalent conjugation to the ubiquitin-related protein SUMO-1. Using an infectious bacterial artificial chromosome clone of human cytomegalovirus, we constructed a mutant virus (BADpmIE1-K450R) that is deficient for SUMOylation of IE1-72 kDa due to a single amino acid exchange in the SUMO-1 attachment site. Compared to wild-type virus, this mutant grew more slowly and generated a reduced yield in infected human fibroblasts, indicating that SUMO modification is required for the full activity of IE1-72 kDa. The lack of SUMOylation did not affect the intranuclear localization of IE1-72 kDa, including its ability to target to and disrupt PML bodies and to bind to mitotic chromatin. Likewise, SUMOylation-deficient IE1-72 kDa activated several viral promoters as efficiently as the wild-type protein. However, the failure to modify IE1-72 kDa resulted in substantially reduced levels of the IE2 transcript and its 86-kDa protein (IE2-86 kDa). These observations suggest that SUMO modification of IE1-72 kDa contributes to efficient HCMV replication by promoting the accumulation of IE2-86 kDa.

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• 5.15

  Impact points

  Murine cytomegalovirus m41 open reading frame encodes a Golgi-localized antiapoptotic protein.

  Wolfram Brune, Michael Nevels, Thomas Shenk

  Journal of virology. 12/2003; 77(21):11633-43.

  Viruses have evolved various strategies to prevent premature apoptosis of infected host cells. Some of the viral genes mediating antiapoptotic functions have been identified by their homology to cellular genes, but others are structurally unrelated to genes of known function. In this study, we used ... [more] Viruses have evolved various strategies to prevent premature apoptosis of infected host cells. Some of the viral genes mediating antiapoptotic functions have been identified by their homology to cellular genes, but others are structurally unrelated to genes of known function. In this study, we used a random, unbiased approach to identify such genes in the murine cytomegalovirus genome. From a library of random transposon insertion mutants, a mutant virus that caused premature cell death was isolated. The transposon was inserted within open reading frame m41. An independently constructed m41 deletion mutant showed the same phenotype, whereas deletion mutants lacking the adjacent genes m40 and M42 did not. Apoptosis occurred in different cell types, could be blocked by caspase inhibitors, and did not require p53. Within the murine cytomegalovirus genome, m41, m40, and m39 form a small cluster of genes of unknown function. They are homologous to r41, r40, and r39 of rat cytomegalovirus, but lack sequence homology to UL41, UL40, and UL37 exon 1 (UL37x1) which are located at the corresponding positions of the human cytomegalovirus genome. Unlike UL37x1 of human cytomegalovirus, which encodes a mitochondrion-localized inhibitor of apoptosis that is essential for virus replication, m41 encodes a protein that localizes to the Golgi apparatus. The murine cytomegalovirus m41 product is the first example of a Golgi-localized protein that prevents premature apoptosis and thus extends the life span of infected cells.

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• 5.15

  Impact points

  "Hit-and-run" transformation by adenovirus oncogenes.

  M Nevels, B Täuber, T Spruss, H Wolf, T Dobner

  Journal of virology. 05/2001; 75(7):3089-94.

  According to classical concepts of viral oncogenesis, the persistence of virus-specific oncogenes is required to maintain the transformed cellular phenotype. In contrast, the "hit-and-run" hypothesis claims that viruses can mediate cellular transformation through an initial "hit,"... [more] According to classical concepts of viral oncogenesis, the persistence of virus-specific oncogenes is required to maintain the transformed cellular phenotype. In contrast, the "hit-and-run" hypothesis claims that viruses can mediate cellular transformation through an initial "hit," while maintenance of the transformed state is compatible with the loss ("run") of viral molecules. It is well established that the adenovirus E1A and E1B gene products can cooperatively transform primary human and rodent cells to a tumorigenic phenotype and that these cells permanently express the viral oncogenes. Additionally, recent studies have shown that the adenovirus E4 region encodes two novel oncoproteins, the products of E4orf6 and E4orf3, which cooperate with the viral E1A proteins to transform primary rat cells in an E1B-like fashion. Unexpectedly, however, cells transformed by E1A and either E4orf6 or E4orf3 fail to express the viral E4 gene products, and only a subset contain E1A proteins. In fact, the majority of these cells lack E4- and E1A-specific DNA sequences, indicating that transformation occurred through a hit-and-run mechanism. We provide evidence that the unusual transforming activities of the adenoviral oncoproteins may be due to their mutagenic potential. Our results strongly support the possibility that even tumors that lack any detectable virus-specific molecules can be of viral origin, which could have a significant impact on the use of adenoviral vectors for gene therapy.

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• 5.15

  Impact points

  Two distinct activities contribute to the oncogenic potential of the adenovirus type 5 E4orf6 protein.

  M Nevels, S Rubenwolf, T Spruss, H Wolf, T Dobner

  Journal of virology. 07/2000; 74(11):5168-81.

  Previous studies have shown that the adenovirus type 5 (Ad5) E4orf6 gene product displays features of a viral oncoprotein. It initiates focal transformation of primary rat cells in cooperation with Ad5 E1 genes and confers multiple additional transformed properties on E1-expressing cells, including ... [more] Previous studies have shown that the adenovirus type 5 (Ad5) E4orf6 gene product displays features of a viral oncoprotein. It initiates focal transformation of primary rat cells in cooperation with Ad5 E1 genes and confers multiple additional transformed properties on E1-expressing cells, including profound morphological alterations and dramatically accelerated tumor growth in nude mice. It has been reported that E4orf6 binds to p53 and, in the presence of the Ad5 E1B-55kDa protein, antagonizes p53 stability by targeting the tumor suppressor protein for active degradation. In the present study, we performed a comprehensive mutant analysis to assign transforming functions of E4orf6 to distinct regions within the viral polypeptide and to analyze a possible correlation between E4orf6-dependent p53 degradation and oncogenesis. Our results show that p53 destabilization maps to multiple regions within both amino- and carboxy-terminal parts of the viral protein and widely cosegregates with E4orf6-dependent acceleration of tumor growth, indicating that both effects are related. In contrast, promotion of focus formation and morphological transformation require only a carboxy-terminal segment of the E4 protein. Thus, these effects are completely independent of p53 stability, but may involve other interactions with the tumor suppressor. Our results demonstrate that at least two distinct activities contribute to the oncogenic potential of Ad5 E4orf6. Although genetically separable, both activities are largely mediated through a novel highly conserved, cysteine-rich motif and a recently described arginine-faced amphipathic alpha helix, which resides within a carboxy-terminal "oncodomain" of the viral protein.

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• 3.04

  Impact points

  An ovine adenovirus vector lacks transforming ability in cells that are transformed by AD5 E1A/B sequences.

  Z Z Xu, M Nevels, E S MacAvoy, L J Lockett, D Curiel, T Dobner, G W Both

  Virology. 05/2000; 270(1):162-72.

  Adenoviruses of the Mastadenovirus and Aviadenovirus genera are able to transform certain cell types and induce tumor formation in susceptible animals. For the mastadenoviruses the E1A/B sequences are largely responsible for these properties but E4 sequences may also be involved. The transforming se... [more] Adenoviruses of the Mastadenovirus and Aviadenovirus genera are able to transform certain cell types and induce tumor formation in susceptible animals. For the mastadenoviruses the E1A/B sequences are largely responsible for these properties but E4 sequences may also be involved. The transforming sequences of the aviadenoviruses, which lack E1A/B and E4 homologues, have not yet been fully identified. The recent proposal for a third genus of adenoviruses, which apparently lack an E1A homologue and have weak E1B homology, prompted an examination of the transforming properties of ovine adenovirus OAV287 (OAV), the prototype member of the new group. When OAV and human adenovirus type 5 (Ad5) were used to infect primary rat embryo cells, transformed foci developed in Ad5- but not in OAV-infected cultures. Similarly, after plasmid transfection, baby rat kidney cells were transformed by Ad5 E1A/B but not by OAV sequences. When CSL503 cells, an ovine cell line that is permissive for OAV, were transfected with Ad5 E1A/B sequences, transformed foci again appeared. However, plasmids or fragments containing complete or partial OAV genome sequences did not detectably transform CSL503 cells under the same conditions. When Ad5 E1A/B sequences were incorporated into the complete OAV genome and transfected, transformed clones were again obtained, showing that the gene dosage and transfection conditions were not limiting for transformation. The provision of Ad5 E1A and OAV sequences in combination marginally increased the number of morphologically altered foci in baby rat kidney cells but failed to induce multilayered focus formation. The data suggest that OAV lacks transforming functions in the cell types examined. Additional information suggesting that OAV may have a fundamentally distinct strategy for replication compared with other Ads is discussed.
• 5.15

  Impact points

  Transforming potential of the adenovirus type 5 E4orf3 protein.

  M Nevels, B Täuber, E Kremmer, T Spruss, H Wolf, T Dobner

  Journal of virology. 03/1999; 73(2):1591-600.

  Previous observations that the adenovirus type 5 (Ad5) E4orf6 and E4orf3 gene products have redundant effects in viral lytic infection together with the recent findings that E4orf6 possesses transforming potential prompted us to investigate the effect of E4orf3 expression on the transformation of pr... [more] Previous observations that the adenovirus type 5 (Ad5) E4orf6 and E4orf3 gene products have redundant effects in viral lytic infection together with the recent findings that E4orf6 possesses transforming potential prompted us to investigate the effect of E4orf3 expression on the transformation of primary rat cells in combination with adenovirus E1 oncogene products. Our results demonstrate for the first time that E4orf3 can cooperate with adenovirus E1A and E1A plus E1B proteins to transform primary baby rat kidney cells, acting synergistically with E4orf6 in the presence of E1B gene products. Transformed rat cells expressing E4orf3 exhibit morphological alterations, higher growth rates and saturation densities, and increased tumorigenicity compared with transformants expressing E1 proteins only. Consistent with previous results for adenovirus-infected cells, the E4orf3 protein is immunologically restricted to discrete nuclear structures known as PML oncogenic domains (PODs) in transformed rat cells. As opposed to E4orf6, the ability of E4orf3 to promote oncogenic cell growth is probably not linked to a modulation of p53 functions and stability. Instead, our results indicate that the transforming activities of E4orf3 are due to combinatorial effects that involve the binding to the adenovirus 55-kDa E1B protein and the colocalization with PODs independent from interactions with the PML gene product. These data fit well with a model in which the reorganization of PODs may trigger a cascade of processes that cause uncontrolled cell proliferation and neoplastic growth. In sum, our results provide strong evidence for the idea that interactions with PODs by viral proteins are linked to oncogenic transformation.

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• 7.14

  Impact points

  The adenovirus E4orf6 protein contributes to malignant transformation by antagonizing E1A-induced accumulation of the tumor suppressor protein p53.

  M Nevels, T Spruss, H Wolf, T Dobner

  Oncogene. 02/1999; 18(1):9-17.

  The adenovirus type 5 (Ad5) E4orf6 protein promotes focus formation of primary baby rat kidney (BRK) cells in cooperation with Ad5 E1 proteins. This activity is most likely related to the ability of the E4orf6 protein to bind to p53 and modulate its tumor suppressor functions. In this study we repor... [more] The adenovirus type 5 (Ad5) E4orf6 protein promotes focus formation of primary baby rat kidney (BRK) cells in cooperation with Ad5 E1 proteins. This activity is most likely related to the ability of the E4orf6 protein to bind to p53 and modulate its tumor suppressor functions. In this study we report that transformed BRK cells that stably express E4orf6 in addition to E1A and E1B (ABS cells) displayed multiple additional properties commonly associated with a high grade of oncogenic transformation compared to cells expressing only E1A and E1B (AB cells). These properties included morphological alterations, markedly enhanced growth rates and growth to much higher saturation densities. Following injection into nude mice ABS-derived tumors exhibited accelerated growth and, based on histopathological criteria, proofed to be much more malignant compared to tumors generated by AB cells. Interestingly, these highly transformed properties of ABS cells correlated with a dramatic reduction of p53 steady-state levels which inversely correlated with E4orf6 expression. From these results we conclude that expression of the Ad5 E4orf6 protein (i) confers additional transformed in vitro properties to primary rat cells expressing the Ad5 E1 proteins, and (ii) increases the tumorigenic and malignant potential of these cells in vivo. Our data suggest that the Ad5 E4orf6 protein enhances the intrinsic ability of E1-transformed rat cells to grow in a neoplastic state by completely inactivating p53 tumor suppressor function in combination with the E1A and E1B proteins.