Studies on the replication of herpes simplex virus in organized cultures of rat central nervous system (CNS) and peripheral nervous system (PNS) tissue demonstrated synthesis of intra- and extracellular virus, as determined by plaque assay on HEp-2 cells. Newly synthesized intracellular virus appeared 12 to 14 hr after inoculation of CNS, followed 10 hr later by the appearance of extracellular virus. In PNS cultures, where higher inputs of virus were introduced, intracellular virus appeared 6 to 8 hr after inoculation, followed by extracellular virus 12 hr later. Polykaryocyte formation was observed in CNS and PNS tissue involving neuroglial, meningeal, or Schwann cells. Neuron somas did not participate in polykaryocyte formation, but they underwent progressive morphological changes starting with increased cytoplasmic granularity followed by nucleolar distortions and disintegration, margination of nuclear chromatin, and the appearance of intranuclear inclusions. Finally, all recognizable cellular detail was lost. Immune serum globulin failed to inhibit both the progressive nature of the cytopathic effect and the synthesis of intracellular virus. These findings are discussed in relation to other in vitro systems, as well as to disease processes in man and animals.
The morphogenesis of the Edmonston strain of measles is described in cultures of hamster dorsal-root ganglion maintained for as long as 63 days postinoculation. The patterns observed confirmed those previously reported in both neural and non-neural tissue. However, in the present tissue, the development of viral material could be followed chronologically within different cell types such as neurons and Schwann cells. Active replication was visualized up to 63 days postinoculation. The appearance of cytoplasmic nucleocapsid preceded that of intranuclear nucleocapsid, the latter occurring after 14 days. These intranuclear inclusions were formed after the transformation of the nucleoli into bizarre pleomorphic bodies which eventually segregated into clumps of nucleocapsid. These intranuclear inclusions mimic those seen in subacute sclerosing panencephalitis, now known to be etiologically related to a measles-like virus.
Data presented elsewhere in this volume amply demonstrate the dependence of the transformed phenotype on specific functions of the papovavirus genome (see, e.g. Crawford, this volume). Similarly, these and other studies have demonstrated that the viral genes are not sufficient in themselves. An understanding of the mechanism of transformation, therefore, will require the exploitation of methods permitting dissection of the cellular processes as well. Given the complexity of the mammalian cell genome, it is not surprising that such insights lag far behind those involving the viral genome. Nonetheless, it has been possible to identify, at least genetically, the role of cellular functions on expression of an integrated wild-type (SV40) genome. Basilico and coworkers (Renger and Basilico 1972; Basilico and Zouzias 1976) have defined a class of cell mutants in SV40-transformed 3T3 cells (SV3T3) that display a temperature-dependent transformed phenotype in regard to density-dependent inhibition of growth (DDIG) and other parameters;...
The effect of DNA antagonists and various antibiotics on steps in the synthesis of SV40 virus in green monkey kidney cells was investigated. Both the early forming tumor (T) antigen, as well as the later synthesized virus (V) antigen, were synthesized in the presence of fluorouracil and iododeoxyuridine. Cytosine arabinoside (and fluorodeoxyuridine in starved cells) prevented synthesis of V antigen but not T antigen. The synthesis of T antigen therefore does not require synthesis of virus DNA. Virus particles formed only in the presence of the iododeoxyuridine and they were non-infectious. Actinomycin D inhibited synthesis of both tumor and virus antigens, suggesting that the synthesis of these antigens involves DNA-dependent RNA. Puromycin allowed synthesis of the T antigen which remained localized at the nucleolar membrane. This finding with puromycin suggests that the T antigen is a protein of low molecular weight. Virus antigen forming in the presence of mitomycin C, p-fluorophenylalanine, iododeoxyuridine, or fluorouracil was distributed atypically. These inhibitors caused the V antigen to be diffusely spread throughout the nucleus, or to be concentrated at the nuclear membrane.
Rapp, Fred (Baylor University College of Medicine, Houston, Tex.), Lawrence A. Feldman, and Manley Mandel . Synthesis of virus deoxyribonucleic acid during abortive infection of simian cells by human adenoviruses. J. Bacteriol. 92: 931–936. 1966.—Inoculation of green monkey kidney cells (GMK) with adenovirus types 2 or 12, under conditions where neither infectious virus was synthesized, resulted in an increase in the uptake of H 3 -thymidine into deoxyribonucleic acid (DNA). Extraction of the DNA from infected cells, followed by identification by isopycnic analysis in CsCl gradients, revealed the presence of virus DNA. Cells infected with adenovirus type 2 yielded DNA giving bands with peak densities of 1.699 g/ml [GMK DNA with 40 moles% guanine + cytosine (GC)] and 1.714 g/ml (adenovirus type 2 DNA with 55 moles% GC). Cells infected with adenovirus type 12 also yielded the GMK DNA and a band at 1.706 g/ml (adenovirus type 12 DNA with 47 moles% GC). The rate of synthesis of adenovirus type 2 DNA in KB cells (productive cycle) and in GMK cells infected only with adenovirus (nonproductive cycle) or with adenovirus and simian virus 40 (adeno-productive cycle) was not significantly different.
Feldman, Lawrence A. (Baylor University College of Medicine, Houston, Tex.), Janet S. Butel, and Fred Rapp . Interaction of a simian papovavirus and adenoviruses. I. Induction of adenovirus tumor antigen during abortive infection of simian cells. J. Bacteriol. 91: 813–818. 1966.—Adenovirus types 2, 7, and 12 undergo an abortive growth cycle in green monkey kidney cells; they induce the formation of adenovirus tumor antigen, but synthesis of adeno capsid antigen and infectious adenovirus was observed only when cultures were concomitantly infected with a simian papovavirus (SV40). Several other viruses, including herpes simplex and measles which replicate in monkey cells, and rabbit papilloma and human wart papovaviruses which do not, failed to stimulate adenovirus replication in the monkey cells. Adenovirus tumor antigen was detected 8 to 10 hr postinfection by immunofluorescent techniques. The antigen induced by adenovirus types 2 and 7 appeared as intranuclear masses; adenovirus type 12 tumor antigen also appeared as cytoplasmic and nuclear flecks. Sera from hamsters bearing tumors induced by adenovirus type 12 cross-reacted with tumor antigens induced by types 2 and 7 but not with antigens induced by SV40.
