New Associations of Human Papillomavirus, Simian Virus 40, and Epstein-Barr Virus with Human Cancer

ABSTRACT The Epstein-Barr virus (EBV)-encoded viral protein kinase, EBV-PK (the BGLF4 gene product), is required for efficient nuclear viral egress in 293 cells. However, since EBV-PK phosphorylates a number of different viral and cellular proteins (including lamin A/C), the relative importance of each target during lytic viral replication remains unclear. We show here that an EBV PK mutant (PKmut; containing stop codons at residues 1 and 5 in EBV-PK) is highly defective for release of infectious virus from 293 cells but not 293T cells. Furthermore, the phenotype of the PKmut in 293 cells is substantially reversed by expression of the simian virus 40 (SV40) large (T) and small (t) T antigens. Efficient rescue requires the presence of both SV40 T/t proteins. We show that 293T cells have a much higher level of constitutive lamin A/C phosphorylation than do 293 cells over residues (S22 and S392) that promote phosphorylation-dependent nuclear disassembly and that both large T and small t contribute to enhanced lamin A/C phosphorylation. Finally, we demonstrate that knockdown of lamin A/C expression using small interfering RNA also rescues the PKmut phenotype in 293 cells. These results suggest that essential roles of EBV-PK during lytic viral replication include the phosphorylation and dispersion of lamin A/C.

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Analysis of mutation in human cells by using an Epstein-Barr virus shuttle system.

Molecular and Cellular Biology ◽

10.1128/mcb.7.1.379 ◽

1987 ◽

Vol 7 (1) ◽

pp. 379-387 ◽

Cited By ~ 784

Author(s):

R B DuBridge ◽

P Tang ◽

H C Hsia ◽

P M Leong ◽

J H Miller ◽

...

Keyword(s):

Cell Lines ◽

Epstein Barr Virus ◽

Shuttle Vector ◽

Simian Virus 40 ◽

Simian Virus ◽

Human Cells ◽

Missense Mutations ◽

Barr Virus ◽

Vector Systems ◽

Epstein Barr

We developed highly sensitive shuttle vector systems for detection of mutations formed in human cells using autonomously replicating derivatives of Epstein-Barr virus (EBV). EBV vectors carrying the bacterial lacI gene as the target for mutation were established in human cells and later returned to Escherichia coli for rapid detection and analysis of lacI mutations. The majority of the clonal cell lines created by establishment of the lacI-EBV vector show spontaneous LacI- frequencies of less than 10(-5) and are suitable for studies of induced mutation. The ability to isolate clonal lines represents a major advantage of the EBV vectors over transiently replicating shuttle vectors (such as those derived from simian virus 40) for the study of mutation. The DNA sequence changes were determined for 61 lacI mutations induced by exposure of one of the cell lines to N-nitroso-N-methylurea. A total of 33 of 34 lacI nonsense mutations and 26 of 27 missense mutations involve G X C to A X T transitions. These data provide support for the mutational theory of cancer.

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A putative origin of replication of plasmids derived from Epstein-Barr virus is composed of two cis-acting components.

Molecular and Cellular Biology ◽

10.1128/mcb.5.8.1822 ◽

1985 ◽

Vol 5 (8) ◽

pp. 1822-1832 ◽

Cited By ~ 256

Author(s):

D Reisman ◽

J Yates ◽

B Sugden

Keyword(s):

Dna Sequences ◽

Epstein Barr Virus ◽

Simian Virus 40 ◽

Repeated Sequence ◽

Simian Virus ◽

Nuclear Antigen ◽

Barr Virus ◽

Early Promoter ◽

The Family ◽

Epstein Barr

A genetic element of Epstein-Barr virus, oriP, when present on recombinant plasmids allows those plasmids to replicate and to be maintained in cells that express the Epstein-Barr virus-encoded nuclear antigen EBNA-1. Here we define the DNA sequences required for oriP activity. Two noncontiguous regions of oriP are required in cis for activity. One consists of approximately 20 tandem, imperfect copies of a 30-base-pair (bp) sequence. The other required region, approximately 1,000 bp away, is at most 114 bp in length and contains a 65-bp region of dyad symmetry. When present together on a plasmid, these two components supported plasmid replication even when the distance between them was varied or their relative orientation was altered, or both. When present alone on a plasmid that expresses a selectable marker, the family of 30-bp repeats efficiently conferred a transient drug-resistant phenotype in human 143 cells that is dependent on the presence of EBNA-1. This result leads us to suggest that EBNA-1 interacts with the 30-bp repeated sequence to activate oriP. To test whether the 30-bp repeats might cause the increased transient expression of drug resistance by enhancing transcription, the family of 30-bp repeats was tested for the ability to activate the simian virus 40 early promoter present in plasmid pA10CAT2 (Laimins, et al., Proc. Natl. Acad. Sci. U.S.A. 79:6453-6457). In this assay, the 30-bp repeats could activate the simian virus 40 early promoter in Raji cells, an EBNA-positive Burkitt's lymphoma cell line, but not detectably an EBNA-positive 143 cells in which oriP also functions.

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The cellular oncogene c-myb can interact synergistically with the Epstein-Barr virus BZLF1 transactivator in lymphoid cells

Molecular and Cellular Biology ◽

10.1128/mcb.12.1.136-146.1992 ◽

1992 ◽

Vol 12 (1) ◽

pp. 136-146

Author(s):

S C Kenney ◽

E Holley-Guthrie ◽

E B Quinlivan ◽

D Gutsch ◽

Q Zhang ◽

...

Keyword(s):

Gene Product ◽

Epstein Barr Virus ◽

Sequence Similarity ◽

Simian Virus 40 ◽

Simian Virus ◽

Lymphoid Cells ◽

Jurkat Cells ◽

Barr Virus ◽

Early Promoter ◽

Epstein Barr

Regulation of replicative functions in the Epstein-Barr virus (EBV) genome is mediated through activation of a virally encoded transcription factor, Z (BZLF1). We have shown that the Z gene product, which binds to AP-1 sites as a homodimer and has sequence similarity to c-Fos, can efficiently activate the EBV early promoter, BMRF1, in certain cell types (i.e., HeLa cells) but not others (i.e., Jurkat cells). Here we demonstrate that the c-myb proto-oncogene product, which is itself a DNA-binding protein and transcriptional transactivator, can interact synergistically with Z in activating the BMRF1 promoter in Jurkat cells (a T-cell line) or Raji cells (an EBV-positive B-cell), whereas the c-myb gene product by itself has little effect. The simian virus 40 early promoter is also synergistically activated by the Z/c-myb combination. Synergistic transactivation of the BMRF1 promoter by the Z/c-myb combination appears to involve direct binding by the Z protein but not the c-myb protein. A 30-bp sequence in the BMRF1 promoter which contains a Z binding site (a consensus AP-1 site) is sufficient to transfer high-level lymphoid-specific responsiveness to the Z/c-myb combination to a heterologous promoter. That the c-myb oncogene product can interact synergistically with an EBV-encoded member of the leucine zipper protein family suggests c-myb is likely to engage in similar interactions with cellularly encoded transcription factors.

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Deletion mutants that affect expression of Epstein-Barr virus nuclear antigen in COS-1 cells after gene transfer with simian virus 40 vectors containing portions of the BamHI K fragment.

Journal of Virology ◽

10.1128/jvi.58.2.324-330.1986 ◽

1986 ◽

Vol 58 (2) ◽

pp. 324-330 ◽

Cited By ~ 2

Author(s):

M Polvino-Bodnar ◽

D Shedd ◽

G Miller

Keyword(s):

Gene Transfer ◽

Epstein Barr Virus ◽

Simian Virus 40 ◽

Simian Virus ◽

Nuclear Antigen ◽

Deletion Mutants ◽

Barr Virus ◽

Affect Expression ◽

Epstein Barr

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A putative origin of replication of plasmids derived from Epstein-Barr virus is composed of two cis-acting components

Molecular and Cellular Biology ◽

10.1128/mcb.5.8.1822-1832.1985 ◽

1985 ◽

Vol 5 (8) ◽

pp. 1822-1832

Author(s):

D Reisman ◽

J Yates ◽

B Sugden

Keyword(s):

Dna Sequences ◽

Epstein Barr Virus ◽

Simian Virus 40 ◽

Repeated Sequence ◽

Simian Virus ◽

Nuclear Antigen ◽

Barr Virus ◽

Early Promoter ◽

The Family ◽

Epstein Barr

A genetic element of Epstein-Barr virus, oriP, when present on recombinant plasmids allows those plasmids to replicate and to be maintained in cells that express the Epstein-Barr virus-encoded nuclear antigen EBNA-1. Here we define the DNA sequences required for oriP activity. Two noncontiguous regions of oriP are required in cis for activity. One consists of approximately 20 tandem, imperfect copies of a 30-base-pair (bp) sequence. The other required region, approximately 1,000 bp away, is at most 114 bp in length and contains a 65-bp region of dyad symmetry. When present together on a plasmid, these two components supported plasmid replication even when the distance between them was varied or their relative orientation was altered, or both. When present alone on a plasmid that expresses a selectable marker, the family of 30-bp repeats efficiently conferred a transient drug-resistant phenotype in human 143 cells that is dependent on the presence of EBNA-1. This result leads us to suggest that EBNA-1 interacts with the 30-bp repeated sequence to activate oriP. To test whether the 30-bp repeats might cause the increased transient expression of drug resistance by enhancing transcription, the family of 30-bp repeats was tested for the ability to activate the simian virus 40 early promoter present in plasmid pA10CAT2 (Laimins, et al., Proc. Natl. Acad. Sci. U.S.A. 79:6453-6457). In this assay, the 30-bp repeats could activate the simian virus 40 early promoter in Raji cells, an EBNA-positive Burkitt's lymphoma cell line, but not detectably an EBNA-positive 143 cells in which oriP also functions.

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