Human herpesvirus 6 (HHV-6) ORF-1 transactivating gene exhibits malignant transforming activity and its protein binds to p53.
The 357 amino acid open reading frame 1 (ORF-1), also designated DR7, within the SalI-L fragment of human herpesvirus 6 (HHV-6) exhibited transactivation of the human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) promoter and increased HIV-1 replication (Kashanchi et al., Virology, 201, 95-106, 1994). In the current study, the SalI-L transforming region was localized to the SalI-L-SH subfragment. Several ORFs identified in SalI-L-SH by sequence analysis were cloned into a selectable mammalian expression vector, pBK-CMV. Only pBK/ORF1 transformed NIH3T3 cells. Furthermore, cells expressing ORF-1 protein produced fibrosarcomas when injected into nude mice, whereas control cells, expressing either no ORF-1 protein or C-terminal truncated (after residue 172) ORF-1 protein, were not tumorigenic. Western blot analysis of proteins extracted from the tumors revealed ORF-1 protein. Additional studies indicated that ORF-1 was expressed in HHV-6-infected human T-cells by 18 h. Co-immunoprecipitation experiments showed that ORF-1 protein bound to tumor suppressor protein p53, and the ORF-1 binding domain on p53 was located between residues 28 and 187 of p53, overlapping with the specific DNA binding domain. Functional studies showed that p53-activated transcription was inhibited in ORF-1, but not in truncated ORF-1, expressing cells. Importantly, the truncated ORF-1 mutant also failed to cause transformation. Analysis of several human tumors by PCR revealed ORF-1 DNA sequences in some angioimmunoblastic lymphadenopathies, Hodgkin's and non-Hodgkin's lymphomas and glioblastomas. The detection of ORF-1 sequences in human tumors, while not proof per se, is a prerequisite for establishing its role in tumor development. Taken together, the results demonstrate that ORF-1 is an HHV-6 oncogene that binds to and affects p53. The identification of both transforming and transactivating activities within ORF-1 is a characteristic of other viral oncogenes and is the first reported for HHV-6.
Kashanchi
F
F
Laboratory of Molecular Virology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Araujo
J
J
Doniger
J
J
Muralidhar
S
S
Hoch
R
R
Khleif
S
S
Mendelson
E
E
Thompson
J
J
Azumi
N
N
Brady
J N
JN
Luppi
M
M
Torelli
G
G
Rosenthal
L J
LJ
D016475:3T3 Cells; D000818:Animals; D005354:Fibrosarcoma / Q000235:genetics; D005809:Genes, Regulator / Q000502:physiology*; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009857:Oncogenes; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism*
C087150:ORF-1 protein, Human herpesvirus 6; D015534:Trans-Activators; D016159:Tumor Suppressor Protein p53; D014764:Viral Proteins