No abstract

Optimum growth conditions for human cytomegalovirus (HCMV) include the use of subconfluent, actively growing cultures of human embryonic fibroblasts. Many clinical virology laboratories, however, use tissue culture cells from commercial sources. These cells are usually confluent, static cultures that tend to be less sensitive to viral infection. To determine whether dimethyl sulfoxide (DMSO) or dexamethasone (DEX), which are known enhancers of HCMV, facilitates the detection of the virus in confluent cells, we tested both HCMV AD169 and a number of clinical specimens suspected to contain HCMV on MRC-5 cells in both shell vials and conventional tube cultures. We found that, in the shell vial test, treatment of the cultures with either DMSO or DEX before and after inoculation increased the number of cells staining positive by threeto sixfold compared with untreated controls. Best results were obtained by pretreating the cultures with DEX alone and by treating the cultures with a combination of DEX and 1% DMSO postinfection. In the conventional MRC-5 culture tubes, treatment with the reagents resulted in the more rapid appearance of cytopathic effect and a more extensive infection of the cell sheet. The experimental findings indicate that the enhancing effect of DEX is attributable mainly to the increased production of a cellular mRNA during the period preceding viral infection.

No abstract

The synthesis of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCase) in Rhodospirillum rubrum was regulated by the CO2 concentration in the culture medium. The specific activity of RuBPCase in cells grown photolithotrophically in low concentrations of CO2 (1.5%) was five to ten times higher than that in cultures grown at high concentrations of CO2 (10%). Increased enzyme activity was reflected by an increase in both RuBPCase mRNA and RuBPCase protein. RuBPCase expression was also studied in vitro with a plasmid-borne genomic clone (pRR117) as the template in a partially defined Escherichia coli system containing either E. coli or R. rubrum RNA polymerase. With both enzymes there was excellent synthesis of RuBPCase mRNA, but no signfficant synthesis of RuBPCase was detected. The promoter region of the RuBPCase gene was sequenced, and mRNA start sites were mapped. A single major in vivo transcriptional start site was detected in RuBPCase mRNA extracted from R. rubrum. However, transcripts synthesized from pRR117 in vitro or from E. coli transformed with pRR117 started at upstream sites that were different from the in vivo transcription site. Two major features of the RuBPCase promoter region are three 6-base-pair direct repeats and a 31-base-pair region of dyad symmetry.