Control of Interferon Production in Rabbit Cells Treated with Poly I:C

I would like to report some of our recent work on the stimulation of rabbit interferon by the double-stranded complex of polyinosinic acid and polycytidylic acid (poly I:C). Most of the work I shall describe was done by Doctors Chris Tan, Yan-hsien Ke, Monto Ho, and myself. Our starting point was Vilcek's interesting work on the accentuation of interferon response in rabbit kidney cells by cycloheximide and his concepts of a mechanism which controls interferon synthesis (Vilcek, 1969). Our work supports this idea of such a mechanism and provides evidence that messenger RNA synthesis is required for a hypothetical control protein. In addition, we found that this control mechanism operates differently in different rabbit tissues. We found that if rabbit kidney cultures are treated with poly I:C (40 /ug/ml) for 1 hr, the expected interferon response is inhibited 90-100% by puromycin (100 tzg/ml) added either 1 or 2 hr after the initiation of poly I: C treatment. At 3 hr, 50% inhibition was observed, and by 4 hr no further inhibition was possible since, by this time, interferon production was already terminated. We concluded that protein synthesis is required for interferon production in this system as was previously shown in slices of tissues from rabbits injected with poly I:C (Ho and Ke, 1970). If cycloheximide (20 /,g/ml) was added to rabbit kidney cultures together with poly I: C and removed at various times thereafter, we found that there was accentuation of interferon production when the blockade of protein synthesis was lifted. Removal of cycloheximide at 2 hr caused a 10-fold accentuation of production when compared with controls receiving no cycloheximide. Removal of cycloheximide 3 hr after initiation of poly I: C treatment caused more than 30-fold accentuation. If removal was delayed for 4 hr, there was no further increase in accentuation. Similar results were obtained with puromycin; for example, a 2 hr blockade of protein synthesis with this compound resulted in an 8-fold accentuation when the blockade was lifted. We concluded from these experiments that the inhibition of protein synthesis blocks the control mechanism, which in the absence of inhibitors