Cloning, Eukaryotic Expression And Detection Of Biological Activity Of Human Interferon-λ3 And Human Interferon-ε

Objective: Human interferon lamda3 (hIFN-λ3) and epsilon (hIFN- ε ) have recently been identified as new types of interferons. Study for both interferons has not been two much so far. Thus, we want to obtain full-length cDNAs for both interferons by RT-PCR amplification, subclone the cDNAs into eukaryotic expression vector and transfect the resulting constructs into mammalian cells. Furthermore, biological activity of proteins expressed by the inserts of the interest was evaluated in mammalian cells. Methods: Total RNA was isolated from mammalian cells stimulated by cytokines. Full-length human hIFN- λ 3 or hIFN- ε cDNA was obtained by RT-PCR and subcloned into the vector, pcDNA3.1/myc-his(-)A. Both constructs was transfected into CHO or COS-7 cells transiently or stably and subsequently detected biological activities and functions of encoded proteins. Results: Both constructs, pcDNA3.1/myc-his(-)A-IFN- ε including 627bp nucleotides and pcDNA3.1/myc-his(-)A-hIFN-λ 3 with 603bp nucleotides, was successfully generated as verified by PCR amplification, restriction endonucleases digestion and following DNA sequencing. Alignment ofresulting cDNAs of the interest with corresponding sequences in Genbank database indicates that our target genes are completely correct in the present study. Both fused plamids can readily expressed their encoded proteins in CHO or COS-7 cells. Both the hlFN- e and hlFN- A 3 proteins show intrinsic cellular antiviral activity. Additionally, hlFN- e protein inhibited cellular proliferation, which is associated with induced expression of MxA in these cells. Altogether, our study provides some clues or probability for future gene engineering and related drugs developments of these interferons. Conclusions: In present study hlFN- k 3 and hlFN- e we successfully obtained and expressed in mammalian cells. Moreover, we demonstrated that both ectopic interferons possessed antiviral and/or antiproliferative activities. Meanwhile our study provide some suggestions for future study of these interferons about their antiviral and antiproliferative activities and possible mechanisms of immunological regulations.