The Effect of the DNA‐Suppressing Factor (DSF) on Host DNA Synthesis in Synchronized Cell Cultures

Abstract

Purified host DNA-suppressing factor (DSF) produced into culture fluid of HeLa C-9 cells infected with measles virus inhibited cellular DNA synthesis in HeLa cells. When purified DSF was added into cultures of synchronous HeLa cells at the early G1-phase, cellular DNA synthesis was irreversibly inhibited. However, DSF did not affect the stability of native double-stranded DNA nor the chain-elongation of single-stranded DNA in cells of the S-phase.Many RNA viruses are known to suppress host macromolecular synthesis (1,3,4,15,24). Among them, reovirus type 3 (2, 6, 16), measles virus (8, 18, 25) and pancreatic necrosis virus (7) inhibit only cellular DNA synthesis, while leaving unaffected the synthesis of protein and RNA. In our previous paper (9), it was reported that the host DNA-suppressing factor (DSF) which was produced into culture fluid after infection with measles virus inhibited cellular DNA synthesis. Recently we succeeded in purification of the DSF. The molecular weight of purified DSF was about 55,000 and the activity of DSF was inactivated by heating at 56 C for 30 min or by treatment with trypsin (22).In the suppression of DNA synthesis by reovirus and pancreatic necrosis virus, a factor similar to DSF has not yet been proved. However, Cox and Shaw (2) showed that the block of DNA synthesis by reovirus appeared to be due to an event(s) during the early G1-phase of the cell cycle. In addition, Lothrop and Nicholson (7) reported that DNA synthesis in the early synthetic phase was relatively insensitive to the inhibition by pancreatic necrosis virus. Thus it is of interest to determine how the DSF reacts at the specific stages of the cell cycle. The present study was undertaken to determine the steps of the suppression of cellular DNA synthesis in HeLa cells by the DSF. MATERIALS AND METHODSCell culture. HeLa C-9 cells which had been cloned from HeLa cells were cultivated in Eagle's minimum essential medium (MEM) supplemented with 5% calf serum. 639

Thymidine Metabolism in Cells Treated with DNA‐Suppressing Factor (DSF)

“…As reported in a previous paper (6), DSF affected cells in the G1-phase. We tried but failed to synchronize the growth of Meth-A cells.…”

“…As described in our previous paper (6), DNA-suppressing factor (DSF) inhibited cellular DNA synthesis only when added to cells in the early G1-phase, and did not affect the stability of double-stranded DNA or the chain-elongation of singlestranded DNA in HeLa cells. We discussed the mechanisms of the inhibition of DNA synthesis by DSF and suggested the following possibilities: 1) DSF might inhibit transport of exogenous thymidine across the cell membrane, 2) DSF might inhibit phosphorylation of thymidine, or 3) DSF might inhibit the initiation of DNA synthesis.…”

Suppression of Interferon-induced Oligo-2',5'-adenylate Synthetase Induction in Persistent Infection

“…Although the mechanism of this suppression is still unknown, it has been reported that 2-5AS mRNA was detectable in K562 cells persistently infected with AIK-C vaccine strain in spite of poor induction of this enzyme (Fujii et al, 1988a). Poor translation of virus matrix M protein mRNA and the inhibition of macromolecular synthesis have been reported in SSPE virusand measles virus-infected cells, respectively (Minagawa et al, 1977;Fujii et al, 1978;Haase et al, 1985). Therefore, it might be considered that translation of 2-5AS mRNA is inhibited by some mechanism in combinations of virus and cell.…”