Gene cloning, sequencing, expression and biological activity of giant panda ( Ailuropoda melanoleuca) interferon-α

• Published in: Molecular immunology Vol. 44; no. 11; pp. 3061 - 3069
• Main Authors: Tan, Xue-Mei, Tang, Yang, Yang, Yun-Fei, Song, Hong-Mei, Zhang, Yi-Zheng
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.04.2007
• Subjects: Ailuropoda melanoleuca; alpha -Interferon; Amino Acid Sequence; Animals; Antiproliferative activity; Antiviral activity; Antiviral Agents - pharmacology; Cell culture; Cell Line; Cell Proliferation - drug effects; Cloning, Molecular; Endangered species; Expression; Gene cloning; Giant panda ( Ailuropoda melanoleuca); HEK293 cell; Humans; Interferon; Interferon-alpha - analysis; Interferon-alpha - genetics; Interferon-alpha - pharmacology; Interferon-α; Molecular Sequence Data; pH effects; Phylogeny; Sequence Alignment; Ursidae - genetics; Western blotting; China, People's Rep; Interferon-α; HEK293 cell; Antiproliferative activity; Expression; Antiviral activity; Gene cloning; Giant panda ( Ailuropoda melanoleuca)
• ISSN: 0161-5890; 1872-9142
• DOI: 10.1016/j.molimm.2006.12.017

The giant panda ( Ailuropoda melanoleuca) is an endangered species and indigenous to China. In mammals, multiple subtypes of interferon-α (IFN-α) exist, most of which possess antiviral activity. Little is known about giant panda IFN-α genes and the role they may play in giant panda immunological responses to viruses. We have cloned genes encoding 12 giant panda IFN-α (AmIFN-α or AmIFNA) subtypes that share from 90 to 99% amino acid sequence identity. AmIFN-α12 has one additional amino acid at position 57, which is not present in other subtypes. Sequence identity of the AmIFN-α proteins encoded by the 12 genes compared to human IFN-α2 is approximately 58%. Unlike most of the human subtypes, each of the 12 giant panda IFN sequences has an N-glycosylation recognition site. Expression of all 12 AmIFN-α subtypes in 293 cells was confirmed by SDS-PAGE and Western blotting analysis. The antiviral activity and antiproliferative activity of each AmIFN-α subtype produced in transiently transfected 293 cell cultures were tested in vitro. All AmIFN-α subtypes were found to be stable at pH 2 or 65 °C and to exhibit antiviral activity. Some IFN subtypes (AmIFN-α8 and AmIFN-α4) showed higher biological activity levels than others, whereas AmIFN-α11 exhibited lower activity. AmIFN-α had various antiproliferative activities to different target cells. To B16 cells, AmIFN-α3, AmIFN-α4, AmIFN-α8 had the highest activities, while to K562 cells, AmIFN-α3, AmIFN-α7, AmIFN-α10 had the highest activities. The various IFN-α subtypes displayed a good correlation between their antiviral and antiproliferative potencies.