Properties of Natural and Hybrid Murine Alpha Interferons

Abstract

SUMMARYFour natural murine interferon-~ genes (MuIFN-cd, -~2, -~4 and -cc6) and four hybrid genes (crick4, ~2cM, ~4cd and cc4~2) were transiently expressed in monkey COS cells under the transcriptional control of the simian virus 40 early promoter. The proteins were labelled with [35S]methionine during a 16 h incubation and proteins secreted by the cells during this period were separated by polyacrylamide gel electrophoresis and subsequently visualized by fluorography. Under the conditions used, the IFNs represented 5 to 10~ of the total amount of secreted proteins. All genes were found to encode biologically active IFN subspecies, including ~4 which has a deletion of five amino acids. When the specific activities of the proteins were compared, it appeared that the specific antiviral activity of c~4 on mouse cells was three-to sixfold higher than the activities of the other natural IFN subspecies. The specific activities of the hybrid proteins were similar to those of the natural proteins, except for the ~2c~4 hybrid which had a higher specific activity than the original proteins. The ability of the natural and hybrid subspecies to protect hamster cells against viral infection was determined using MuIFN-~I as a standard. Large differences in activity were found, with ~6 as the most and ~4 as the least active subspecies.

Characterization of the Murine Alpha Interferon Gene Family

“…These include IFN-␤, IFN-␣1, IFN-␣2, IFN-␣4, IFN-␣5, and IFN-␣13 (8, 43, 44). In contrast, IFN-␣6T and IFN-␣14 were known to lack N-glycosylation (43,44). It was, therefore, of interest to analyze the glycosylation status of the other IFN subtypes.…”

The interferon‐stimulated gene 54 K promoter contains two adjacent functional interferon‐stimulated response elements of different strength, which act synergistically for maximal interferon‐α inducibility

“…The supernatant of constitutively MuIFN-a6-producing CHO-12 cells (CHO-12 clone 28, which produces MuIFN-a6, a mouse IFN-a species which is active on both mouse and hamster cells [24,251) was used as the interferon source. MuIFN-a6 was purified by affinity chromatography over an anti-(MuIFN-a) immunoglobulin column.…”

Isolation and characterization of a cDNA encoding the low molecular weight insulin-like growth factor binding protein (IBP-1).

“…We constructed the expression plasmid pSV19 by inserting cDNA p19 (Figure 1) into expression vector pSV328 (Van Heuvel et al, 1986 (Figure 5, lanes A and D). Since the cDNA clone p19 was isolated from a placental cDNA library it was surprising to find that RNA isolated from placental tissue did not show any expression ( Figure 5, lane B).…”

“…The cDNA clone p19 encoding the IBP-1 gene was inserted into the EcoRI site of pSV328, which expresses cloned inserts using the simian virus 40 (SV40) early promoter (Van Heuvel et al, 1986). COS-1 cells (Gluzman, 1981) were transfected with the expression vector by incubation with 100 Ag/ml DEAE-dextran for 2 h (McCutchan and Pagano, 1968) followed by treatment with 100 zM chloroquine in Dulbecco's MEM (DMEM) for 4 h. After this treatment the cells were fed with DMEM plus 5 % fetal calf serum for 24 h. Subsequently, the cells were washed extensively with DMEM and incubated for 72 h with DMEM without serum.…”