scholarly journals “Hit-and-Run” Transformation by Adenovirus Oncogenes

2001 ◽  
Vol 75 (7) ◽  
pp. 3089-3094 ◽  
Author(s):  
Michael Nevels ◽  
Birgitt Täuber ◽  
Thilo Spruss ◽  
Hans Wolf ◽  
Thomas Dobner
Keyword(s):  
Dna Sequences ◽  
Cellular Transformation ◽  
Adenoviral Vectors ◽  
Gene Products ◽  
Mutagenic Potential ◽  
Viral Origin ◽  
Adenovirus E1a ◽  
Viral Oncogenes ◽  
E4 Region ◽  
Hit And Run

ABSTRACT 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.

scholarly journals Molecular cloning and chromosomal localization of DNA sequences associated with a human DNA repair gene.

1985 ◽  
Vol 5 (2) ◽  
pp. 398-405 ◽  
Author(s):  
J S Rubin ◽  
V R Prideaux ◽  
H F Willard ◽  
A M Dulhanty ◽  
G F Whitmore ◽  
...  
Keyword(s):  
Dna Repair ◽  
Dna Sequences ◽  
Chromosomal Localization ◽  
Excision Repair ◽  
Repair Process ◽  
Human Cells ◽  
Gene Products ◽  
Repair Gene ◽  
Human Dna ◽  
Dna Repair Gene

The genes and gene products involved in the mammalian DNA repair processes have yet to be identified. Toward this end we made use of a number of DNA repair-proficient transformants that were generated after transfection of DNA from repair-proficient human cells into a mutant hamster line that is defective in the initial incision step of the excision repair process. In this report, biochemical evidence is presented that demonstrates that these transformants are repair proficient. In addition, we describe the molecular identification and cloning of unique DNA sequences closely associated with the transfected human DNA repair gene and demonstrate the presence of homologous DNA sequences in human cells and in the repair-proficient DNA transformants. The chromosomal location of these sequences was determined by using a panel of rodent-human somatic cell hybrids. Both unique DNA sequences were found to be on human chromosome 19.

scholarly journals Isolation of monoclonal antibodies specific for products of avian oncogene myb.

1984 ◽  
Vol 4 (12) ◽  
pp. 2843-2850 ◽  
Author(s):  
G I Evan ◽  
G K Lewis ◽  
J M Bishop
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Nuclear Protein ◽  
Gene Products ◽  
Viral Oncogenes ◽  
Myb Gene

We isolated a series of monoclonal antibodies which were raised against a bacterially expressed protein, bp37v-myb, and coded for by part of the avian v-myb gene. These monoclonal antibodies recognized a range of antigenic specificities on bp37v-myb, and this was reflected in their differing specificities for the gene products of the v-myb, c-myb, and E26 viral oncogenes. One monoclonal antibody recognized, in addition to the v-myb and c-myb gene products, a conserved nuclear protein found in all tested cells. We describe the characterization of these monoclonal antibodies.

ACR1, a yeast ATF/CREB repressor

1992 ◽  
Vol 12 (12) ◽  
pp. 5394-5405
Author(s):  
A C Vincent ◽  
K Struhl
Keyword(s):  
Dna Binding ◽  
Cyclic Amp ◽  
Dna Sequences ◽  
Transcriptional Repressors ◽  
Yeast Gene ◽  
Yeast Saccharomyces Cerevisiae ◽  
Viral Oncogenes ◽  
Activation Of Transcription ◽  
Bzip Domain ◽  
New Yeast

Members of the mammalian ATF/CREB family of transcription factors, which are associated with regulation by cyclic AMP and viral oncogenes, bind common DNA sequences (consensus TGACGTCA) via a bZIP domain. In the yeast Saccharomyces cerevisiae, ATF/CREB-like sequences confer either repression or activation of transcription, depending on the promoter context. By isolating mutations that alleviate the repression mediated by ATF/CREB sites, we define a new yeast gene, ACR1, which encodes an ATF/CREB transcriptional repressor. ACR1 contains a bZIP domain that is necessary for homodimer formation and specific DNA binding to an ATF/CREB site. Within the bZIP domain, ACR1 most strongly resembles the mammalian cyclic AMP-responsive transcriptional regulators CREB and CREM; it is less similar to GCN4 and YAP1, two previously described yeast bZIP transcriptional activators that recognize the related AP-1 sequence (consensus TGACTCA). Interestingly, deletion of the ACR1 gene causes increased transcription through ATF/CREB sites that does not depend on GCN4 or YAP1. Moreover, extracts from acr1 deletion strains contain one or more ATF/CREB-like DNA-binding activities. These genetic and biochemical observations suggest that S. cerevisiae contains a family of ATF/CREB proteins that function as transcriptional repressors or activators.

scholarly journals Adenovirus E1A specifically blocks SWI/SNF-dependent transcriptional activation.

1996 ◽  
Vol 16 (10) ◽  
pp. 5737-5743 ◽  
Author(s):  
M E Miller ◽  
B R Cairns ◽  
R S Levinson ◽  
K R Yamamoto ◽  
D A Engel ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Cellular Transformation ◽  
Transcriptional Coactivator ◽  
Gene Encoding ◽  
Adenovirus E1a ◽  
Transcriptional Regulatory ◽  
Genes Encoding ◽  
Regulatory Complex ◽  
Activation Pathways ◽  
Coactivator P300

Expression of the adenovirus E1A243 oncoprotein in Saccharomyces cerevisiae produces a slow-growth phenotype with accumulation of cells in the G1 phase of the cell cycle. This effect is due to the N-terminal and CR1 domains of E1A243, which in rodent cells are involved in triggering cellular transformation and also in binding to the cellular transcriptional coactivator p300. A genetic screen was undertaken to identify genes required for the function of E1A243 in S. cerevisiae. This screen identified SNF12, a gene encoding the 73-kDa subunit of the SWI/SNF transcriptional regulatory complex. Mutation of genes encoding known members of the SWI/SNF complex also led to loss of E1A function, suggesting that the SWI/SNF complex is a target of E1A243. Moreover, expression of E1A in wild-type cells specifically blocked transcriptional activation of the INO1 and SUC2 genes, whose activation pathways are distinct but have a common requirement for the SWI/SNF complex. These data demonstrate a specific functional interaction between E1A and the SWI/SNF complex and suggest that a similar interaction takes place in rodent and human cells.

scholarly journals E1A 13S and 12S mRNA products made in Escherichia coli both function as nucleus-localized transcription activators but do not directly bind DNA.

1985 ◽  
Vol 5 (10) ◽  
pp. 2653-2661 ◽  
Author(s):  
B Ferguson ◽  
B Krippl ◽  
O Andrisani ◽  
N Jones ◽  
H Westphal ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Mammalian Cells ◽  
Biochemical Properties ◽  
Protein Product ◽  
Gene Products ◽  
E Coli ◽  
Adenovirus E1a ◽  
Myc Gene ◽  
E1a Gene ◽  
Transcription Activators

We previously purified and characterized functionally the Escherichia coli-expressed product of the human subgroup C adenovirus E1A 13S mRNA (B. Ferguson, N. Jones, J. Richter, and M. Rosenberg, Science 224:1343-1346, 1984; B. Krippl, B. Ferguson, M. Rosenberg, and H. Westphal, Proc. Natl. Acad. Sci. USA 81:6988-6992, 1984). We have now expressed in E. coli and purified the protein product encoded by the human subgroup C adenovirus E1A 12S mRNA and have compared the functional properties of this protein with those of the E1A 13S mRNA product. Using microinjection techniques to introduce these proteins into mammalian cells, we found that the E1A 12S mRNA product, like the 13S mRNA product, localized rapidly to the cell nucleus and induced adenovirus gene expression. Although both E1A gene products localized to the nucleus and stimulated adenovirus gene transcription, these proteins did not directly bind to DNA under conditions in which a known DNA-binding protein, the human c-myc gene product, bound DNA efficiently. Thus, the E1A and myc gene products, which have been related both structurally and functionally, exhibit distinctly different biochemical properties.

scholarly journals Isolation and characterization of human DNA fragments with nucleotide sequence homologies with the simian virus 40 regulatory region.

1982 ◽  
Vol 2 (8) ◽  
pp. 949-965 ◽  
Author(s):  
S E Conrad ◽  
M R Botchan
Keyword(s):  
Thymidine Kinase ◽  
Dna Sequences ◽  
Regulatory Region ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Dna Fragments ◽  
Viral Origin ◽  
Isolation And Characterization ◽  
Human Dna ◽  
Sv40 Dna

A recombinant library of human DNA sequences was screened with a segment of simian virus 40 (SV40) DNA that spans the viral origin of replication. One hundred and fifty phage were isolated that hybridized to this probe. Restriction enzyme and hybridization analyses indicated that these sequences were partially homologous to one another. Direct DNA sequencing of two such SV40-hybridizing segments indicated that this was not a highly conserved family of sequences, but rather a set of DNA fragments that contained repetitive regions of high guanine plus cytosine content. These sequences were not members of the previously described Alu family of repeats and hybridized to SV40 DNA more strongly than do Alu family members. Computer analyses showed that the human DNA segments contained multiple homologies with sequences throughout the SV40 origin region, although sequences on the late side of the viral origin contained the strongest cross-hybridizing sequences. Because of the number and complexity of the matches detected, we could not determine unambiguously which of the many possible heteroduplexes between these DNAs was thermodynamically most favored. No hybridization of these human DNA sequences to any other segment of the SV40 genome was detected. In contrast, the human DNA segments isolated cross-hybridized with many sequences within the human genome. We tested for the presence of several functional domains on two of these human DNA fragments. One SV40-hybridizing fragment, SVCR29, contained a sequence which enhanced the efficiency of thymidine kinase transformation in human cells by approximately 20-fold. This effect was seen in an orientation-independent manner when the sequence was present at the 3' end of the chicken thymidine kinase gene. We propose that this segment of DNA contains a sequence analogous to the 72-base-pair repeats of SV40. The existence of such an "activator" element in cellular DNA raises the possibility that families of these sequences may exist in the mammalian genome.

scholarly journals Protein phosphatases and DNA tumor viruses: transformation through the back door?

1991 ◽  
Vol 2 (8) ◽  
pp. 589-598 ◽  
Author(s):  
M C Mumby ◽  
G Walter
Keyword(s):  
Phosphatase Activity ◽  
Protein Kinases ◽  
Normal Growth ◽  
Cellular Transformation ◽  
Cellular Protein ◽  
Oncogenic Viruses ◽  
Biochemical Control ◽  
Viral Oncogenes ◽  
Tumor Viruses ◽  
Back Door

Cellular transformation by many oncogenic viruses is mediated by alterations in signal transduction pathways that control normal growth and proliferation. Common targets for many transforming viruses are pathways regulated by protein phosphorylation. The biochemical control of proteins in these pathways is a dynamic process that is regulated by the relative activities of protein kinases and phosphatases. Although there are numerous examples of viral oncogenes that encode protein kinases (Hunter, 1991), until recently there has been no evidence linking altered phosphatase activity to transformation. In this review we describe a novel mechanism, utilized by small DNA tumor viruses, in which viral oncogenes bind to and regulate a cellular protein serine/threonine phosphatase. The currently available evidence indicates that alteration of phosphatase activity and subsequent changes in phosphorylation levels is an important step in transformation by these viruses.

scholarly journals DNA-binding properties of simian virus 40 T-antigen mutants defective in viral DNA replication.

1983 ◽  
Vol 3 (11) ◽  
pp. 1958-1966 ◽  
Author(s):  
C Prives ◽  
L Covey ◽  
A Scheller ◽  
Y Gluzman
Keyword(s):  
Amino Acid ◽  
Dna Sequences ◽  
Simian Virus 40 ◽  
Amino Acid Position ◽  
Simian Virus ◽  
T Antigen ◽  
Regulatory Sequences ◽  
Viral Dna ◽  
Viral Origin ◽  
Specific Affinity

Three simian virus 40 (SV40)-transformed monkey cell lines, C2, C6, and C11, producing T-antigen variants that are unable to initiate viral DNA replication, were analyzed with respect to their affinity for regulatory sequences at the viral origin of replication. C2 and C11 T antigens both bound specifically to sequences at sites 1 and 2 at the viral origin region, whereas C6 T antigen showed no specific affinity for any viral DNA sequences under all conditions tested. Viral DNA sequences encoding the C6 T antigen have recently been cloned out of C6 cells and used to transform an established rat cell line. T antigen from several cloned C6-SV40-transformed rat lines failed to bind specifically to the origin. C6 DNA contains three mutations: two located close to the amino terminus of T antigen at amino acid positions 30 and 51 and a third located internally at amino acid position 153. Two recombinant SV40 DNA mutants were prepared containing either the amino-terminal mutations at positions 30 and 51 (C6-1) or the internally located mutation at position 153 (C6-2) and used to transform Rat 2 cells. Whereas T antigen from C6-2-transformed cells lacked any specific affinity for these sequences. Therefore, the single mutation at amino acid position 153 (Asn leads to Thr) is sufficient to abolish the origin-binding property of T antigen. A T antigen-specific monoclonal antibody, PAb 100, which had been previously shown to immunoprecipitate an immunologically distinct origin-binding subclass of T antigen, recognized wild-type or C6-1 antigens, but failed to react with C6 or C6-2 T antigens. These results indicate that viral replication function comprises properties of T antigen that exist in addition to its ability to bind specifically to the SV40 regulatory sequences. Furthermore, it is concluded from these data that specific viral origin binding is not a necessary feature of the transforming function of T antigen.

scholarly journals Regulation of Adenovirus-Mediated Transgene Expression by the Viral E4 Gene Products: Requirement for E4 ORF3

1999 ◽  
Vol 73 (10) ◽  
pp. 8308-8319 ◽  
Author(s):  
M. Lusky ◽  
L. Grave ◽  
A. Dieterlé ◽  
D. Dreyer ◽  
M. Christ ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Transgene Expression ◽  
Cystic Fibrosis Gene ◽  
Gene Products ◽  
In Vivo Evaluation ◽  
Cmv Promoter ◽  
Reading Frame ◽  
E4 Region

ABSTRACT In a previous study we showed that multiple deletions of the adenoviral regulatory E1/E3/E4 or E1/E3/E2A genes did not influence the in vivo persistence of the viral genome or affect the antiviral host immune response (Lusky et al., J. Virol. 72:2022–2032, 1998). In this study, the influence of the adenoviral E4 region on the strength and persistence of transgene expression was evaluated by using as a model system the human cystic fibrosis transmembrane conductance regulator (CFTR) cDNA transcribed from the cytomegalovirus (CMV) promoter. We show that the viral E4 region is indispensable for persistent expression from the CMV promoter in vitro and in vivo, with, however, a tissue-specific modulation of E4 function(s). In the liver, E4 open reading frame 3 (ORF3) was necessary and sufficient to establish and maintain CFTR expression. In addition, the E4 ORF3-dependent activation of transgene expression was enhanced in the presence of either E4 ORF4 or E4 ORF6 and ORF6/7. In the lung, establishment of transgene expression was independent of the E4 gene products but maintenance of stable transgene expression required E4 ORF3 together with either E4 ORF4 or E4 ORF6 and ORF6/7. Nuclear run-on experiments showed that initiation of transcription from the CMV promoter was severely reduced in the absence of E4 functions but could be partially restored in the presence of either ORF3 and ORF4 or ORFs 1 through 4. These results imply a direct involvement of some of the E4-encoded proteins in the transcriptional regulation of heterologous transgenes. We also report that C57BL/6 mice are immunologically weakly responsive to the human CFTR protein. This observation implies that such mice may constitute attractive hosts for the in vivo evaluation of vectors for cystic fibrosis gene therapy.

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