Váczi L

Stimulation of host DNA synthesis and induction of early antigens by ultraviolet light irradiated human cytomegalovirus

SummaryThe ultraviolet (UV-)sensitivity of the human cytomegalovirus (HCMV) genes coding for very early complement fixing and early antigens in human embryonic fibroblasts (HEF) and mouse embryonic fibroblasts (MEF) and the relation of these genes to the ability of the virus to stimulate host cell DNA synthesis were investigated.After 14 minutes of UV-irradiation of the virus inoculum only the very early complement fixing nuclear antigen (CMNA) developed in the HEF cells and only the early cytoplasmic antigen(s) was present in the MEF. In both HEF and MEF, host cell DNA synthesis was stimulated. We conclude that the ability of HCMV to stimulate host DNA synthesis is an early function of the viral genome and shows a high resistance to UV-irradiation.There is no direct correlation, however, between the ability of the virus to stimulate host cell DNA synthesis and the genes which code for the CMNA or for early cytoplasmic antigens.

Cytomegalovirus latency in cultured human cells.

Summary Infectious virus could not be recovered by disruption of human embryonic fibroblast cultures inoculated with cytomegalovirus and treated with cytosine-arabinoside (ara-C), either during treatment or 4 to 5 days after removing ara-C. However, when intact cells were used for infection, at least 1 in 70 cells was found harbouring virus. Examination by immunofluorescence revealed that 40 to 50% of the cells contained virus-specific antigens in the form of small granules diffusely distributed in the nuclei. The presence of infectious virus could be detected even in disrupted cells 4 to 5 days after removing ara-C; intracellular antigens of every kind were also found to develop. Experiments showed that the cultures continued to harbour latent infection indefinitely in the presence of ara-C. Cultures harbouring latent virus were susceptible to superinfection with cytomegalovirus.

Detection of human cytomegalovirus-induced DNA-binding proteins on Western blots

DNA-binding proteins have been isolated from human embryonic fibroblast cells infected with human cytomegalovirus strain AD169 by native and denatured DNA-cellulose affinity chromatography and Polymin P precipitation. The DNA-binding proteins separated by polyacrylamide gel electrophoresis and electrophoretically transferred to a nitrocellulose sheet were identified by a modified ELISA reaction. Six DNA-binding proteins were found in infected cells, their molecular weights range from 52K to 18K.

Early Nuclear Antigen as DNA-Binding Protein in Cytomegalovirus-Infected Cells

Early nuclear complement-fixing antigen in cytomegalovirus (CMV) (strain Ad 169)-infected human embryonic fibroblasts is described. The nuclear antigen was solubilized from CMV-infected cells by high salt treatment. DNA-binding properties were studied by DNA-cellulose chromatography. The purified antigen, eluted from double-stranded DNA-cellulose columns, was added to acid-fixed nuclear preparations from human embryonic fibroblasts and then exposed to human sera containing antibodies to CMV. Positive staining was obtained by anti-complement immunofluorescence. These data identify the CMV-determined nuclear antigen as a DNA-binding protein. In this respect, it is similar to the Epstein-Barr virus nuclear antigen.

Activation of herpes specific antigens and infectious virus in a hamster cell line transformed by HSV 2

An established cell line transformed by herpes simplex virus type 2 and derived from hamster tumors contained herpesvirus specific intraeellular and surface antigens in about 2–5 per cent of cells but did not produce infectious virus. Arginine deprivation resulted in induction of virus specific intracellular and surface antigens in 70–75 and about 40 per cent of cells, respectively, while no infectious virus was induced. After treatment of the cultures with IUDR 70–75 per cent of the cells showed virus specific intracellular fluorescence and 45–50 per cent exhibited cell membrane fluorescence and infectious virus could be detected. The induction by IUDR could be inhibited by simultaneous treatment of cultures with cytosine arabinoside (ara-C).

Antibodies to the second Epstein-Barr virus nuclear antigen in non-Hodgkin lymphomas

Human antibodies against the first and second Epstein-Barr virus encoded nuclear antigens (EBNA-1 and EBNA-2) were analysed by modified ELISA reaction on immunoblots: 5 out of 20 EBNA-1 positive, healthy donor sera reacted with the EBNA-2. Healthy EBV seronegatives contained no antibodies against EBNA-2. The antibodies against this EBNA-2 developed several months after acute EBV infections. Four out of 8 infectious mononucleosis sera failed to react EBNA-2 in the late stage. In the group of EBV seropositive patients with non-Hodgkin lymphoma, 14 out of 18 sera contained EBNA-2 antibodies.

Vesicular stomatitis virus pseudotypes produced by cells abortively infected or transformed by human cytomegalovirus.

SummaryPseudotypes of vesicular stomatitis virus (VSV) and human cytomegalovirus (HCMV) were produced by normal hamster cells abortively infected with HCMV and superinfected by VSV at a certain stage of abortive HCMV infection. Hamster cells transformedin vitro by HCMV (87-TRH-5 and CX-90-3B cells) also produce VSV (HCMV) pseudotypes after infection of the cells by VSV, but the same cells after passagein vivo (TSC-1, TSC-2 cells) do not.