N-terminal amino acids of bovine alpha interferons are relevant for the neutralization of their antiviral activity

The structure-function relationship of interferons (IFNs) has been studied by epitope mapping. Epitopes of bovine IFNs, however, are practically unknown, despite their importance in virus infections and in the maternal recognition of pregnancy. It has been shown that recombinant bovine (rBo)IFN-alphaC and rBoIFN-alpha1 differ only in 12 amino acids and that the F12 monoclonal antibody (mAb) binds to a linear sequence of residues 10 to 34. We show here that the antiviral activities of these two IFNs were neutralized by the F12 mAb to different extents using two tests. In residual activity tests the antiviral activity dropped by more than 99% with rBoIFN-alphaC and by 84% with rBoIFN-alpha1. In checkerboard antibody titrations, the F12 mAb titer was 12,000 with rBoIFN-alphaC and only 600 with rBoIFN-alpha1. Since these IFNs differ in their amino acid sequence at positions 11, 16 and 19 of the amino terminus, only these amino acids could account for the different neutralization titers, and they should participate in antibody binding. According to the three-dimensional structure described for human and murine IFNs, these amino acids are located in the alpha helix A; amino acids 16 and 19 of the bovine IFNs would be expected to be exposed and could bind to the antibody directly. The amino acid at position 11 forms a hydrogen bond in human IFNs-alpha and it is possible that, in bovine IFNs-alpha, the F12 mAb, binding near position 11, would disturb this hydrogen bond, resulting in the difference in the extent of neutralization observed

Biological activities of a human amniotic membrane interferon

In order to characterize further the human amniotic membrane interferon (IFN-AM), an interferon antigenically unrelated to human IFN-α, -β, and -γ or TNF, we analysed its biological activities. Here, we present direct evidence of its ability to affect cell growth and to induce the IFN-stimulated genes (ISGs) 6-16 and 2'-5' oligoadenylate synthetase (OAS), in addition to its crossed anti-viral activity. The cellular growth arrest effect of IFN-AM was dose-dependent and paralleled that of IFN-β. IFN-AM was also able to inhibit thymidine incorporation into DNA, similar to IFN-β. The mRNA induction of 6-16 gene with IFN-AM treatment reached its highest level at 500 IU/ml and remained constant up to 2000 IU/ml. Conversely, 2'-5' OAS mRNA induction was dose-dependent, with the maximum level detected at 2000 IU/ml of IFN-AM treatment. The time course of mRNA accumulation by ISGs with IFN-AM (500 IU/ml) stimulation was also investigated. Gene induction reached a maximum at 16 h after IFN treatment for 2'-5' OAS and at 48 h for the 6-16 gene. IFN-AM and human IFN-α induced similar levels of the OAS enzyme. IFN-AM also showed small but significant activity in bovine cells. In conclusion, the amniotic membrane IFN here studied showed both anti-cellular activity and the ability to stimulate ISG-transcriptional activation in a similar manner to IFN-β. In addition, IFN-AM was also as able to induce the expression of the enzyme 2'-5' OAS, as did IFN-α. Lastly, amniotic IFN showed a significant cross-species anti-viral activity, which was different from both human IFN-α and -β. Taken together, these data strongly suggest that IFN-AM is a novel sub-type I IFN.SCOPUS: ar.jinfo:eu-repo/semantics/publishe