Expression of "early" and "late" viral functions in a somatic cell hybrid between a mouse cell and a spontaneous yielder SV 40-transformed Chinese hamster cell.

A somatic cell hybrid (Cl. 6d) was originated from the fusion of mouse 3T3-4E) and spontaneous yielder SV 40-transformed Chinese hamster (CHK/SVLP AG) cells. During the early stages of its history, the C1. 6d hybrid underwent a rapid chromosome loss, preferentially loosing hamster chromosomes. This was not a constant tendency of the hybrid cells. As the parental CHK)SVLP AG cells, the hybrid cells were always found 100 per cent SV40 T-antigen positive. While CHK/SVLP AG cells infectious SV 40 DNA, V-antigen and virus were regularly detected, in the hybrid cells only infectious DNA was occasionally detected. This was not due either to the loww of an essential Chinese hamster gene(s) or to the presence of an inhibiting mouse cell component(s); it was apparently the consequence of inability of the cells to properly activate the resident SV 40 genome(s). After superinfection with SV 40 DNA, the hybrid cells-though capable of synthesizing SV 40 V-antigen--were unable to ensure virus assembly. Experimental evidence was obtained suggesting that SV 40 maturation is dependent of a cellular function(s).

Simian virus 40-Chinese hamster kidney cell interaction. III. Characteristics of chemical induction in a clone of virogenic transformed cells.

A number of chemical and physical agents were screened to determine their effectiveness in inducing simian virus (SV40) production in a virogenic clone of SV40-transformed Chinese hamster cells. Mitomycin C (MC) was the most effective inducing agent, and MC induction was further characterized. It was found that levels of infectious SV40 DNA were increased above control levels as early as 6 h after addition of MC to the culture medium and reached maximum levels by 48 h. Virus capsid (V) antigen and virions followed with a lag of about 24 h. V antigen production was sensitive to hydroxyurea, suggesting a dependence on virus DNA synthesis. The proportion of virus-producing cells (infectious centres) and the virus burst per cell were both stimulated by MC. Studies of 3H-thmidine incorporation demonstrated that the rate of SV40 DNA synthesis was maximal at 48 h post-induction, at which time cellular DNA synthesis was almost abolished. Caffeine, at doses not toxic to non-induced cells, strongly inhibited SV40production in both non-induced and induced cells, suggesting some role for DNA repair mechanisms.

Simian virus 40-Chinese hamster kidney cell interaction. IV. Enhanced virus replication in infected cells upon treatment with mitomycin C.

Chinese hamster kidney cells are semi-permissive to simian virus 40 (SV40). Exposure to mitomycin C (MC) of Chinese hamster kidney cells infected with SV40 DNA enhanced the yield of infectious virus 10- to 100-fold. This stimulation occurred whether the treatment was performed before or after infection. A simultaneous increase in the number of V antigen-synthesizing cells and virus-producing cells, as well as the virus burst size, was observed upon MC pretreatment, whereas the proportion of T antigen-synthesizing cells remained unchanged. MC pretreatment clearly stimulated virus DNA replication in SV40 virus-infected cells. Cells treated with MC exhibited an unbalanced growth pattern, with continuing protein synthesis in the absence of cell division and a markedly reduced ability to replicate the cellular DNA. These results suggest that MC enhances the permissiveness of Chinese hamster kidney cells by inducing the synthesis of a specific cellular factor(s) required for SV40 reproduction in Chinese hamster kidney cells.

Simian virus 40-chinese hamster kidney cell interaction. II. The semipermissivity of the cell system.

Throughout in vitro passages, Chinese hamster kidney (CHK) cells progressively lost susceptibility to SV 40 virus infection while remaining continuously susceptible to viral DNA infection. Upon infection with SV 40 virus or viral DNA, the CHK cell line supported viral DNA and virus replication at a low level. SV 40 transformed CHK cell lines spontaneously produced small amounts of viral DNA and virions. The percentage of virus-producing cells was low. Various clones derived from each of these lines behaved as the parental cell population, leading to the conclusion that each CHK cell, whether transformed or not with SV 40, is potentially permissive for this virus.

Simian virus 40-chinese hamster kidney cell interaction. I. Relationship of chromosome changes to transformation.

After approximately 20 in vitro passages, Chinese hamster kidney (CHK) cell cultures transformed upon exposure to different strains of SV 40 can show a diploid modal chromosome number of 22 with chromosome counts exclusively or essentially in the diploid range (20-25). In primary culture and at the 5th-7th subculture, tumors produced in nude mice by such cells can display a diploid modal value of 22 chromosomes. Numerical chromosome variations and karyotype abnormalities observed in CHK cells transformed upon infection with SV 40, are apparently indistinguishable from those that can be observed in uninfected CHK cells undergoing "spontaneous"-transformation. These results indicate that polyploidization is not a necessary step either in the process of transformation or in that of tumorigenic conversion with SV 40.