Cross-Species Genome-Wide Analysis Reveals Molecular and Functional Diversity of the Unconventional Interferon-? Subtype

Shields, Lauren E.; Jennings, Jordan; Liu, Qinfang; Lee, Jinhwa; Ma, Wenjun; Blecha, Frank; Miller, Laura C.; Sang, Yongming

Compared with IFN-γ, two major physiochemical property of type I IFN peptides are their tolerance in acidic solutions (pH < 4.5), but less thermal stability in higher temperature (1, 20, 25) . For comparative activity assays, we synthesized several porcine IFN peptides using both a mammalian expression system (HEK293F, Invitrogen) for authentic verification at small scale (9, 13) and a yeast-expression system (Kingfisher Biotech) for bulk production (24) . Pertaining to antiviral activity compared in several virus-cell systems (conducted both previously and in this study), IFN peptides produced in both mammalian and yeast system were very comparable and exerted very similar antiviral activity (9, 24). We determined the acidic and thermal stability of IFN-ω1 and IFN-ω5 peptides in comparison to IFN-α1 peptide. As shown in Table 1 , both IFN-α and IFN-ω peptides had similar acidic stability after incubation with an acetic acid and identified genomic polymorphisms (Ws). SsIFNW1-7 represent seven original identifications of IFN-ω genes, of which they share <95% sequence identity among each other. SsIFNW4L, 5L, 6L, and 7L are nearly identical duplicates of SsIFNW4-7, respectively; they are further annotated from current swine genome assembly (Sscorfa10.2). All genomic polymorphisms generally have only one to several nucleotide mutations and are clustered with their parental SsIFNW genes in the same branches. Note, more SsIFNW5 polymorphisms have been identified, and they exert diverse antiviral activity as partly shown in Figure 9 . The associated taxa clustered together in the bootstrap test (1,000 replicates) are shown next to the branches. Evolutionary analyses were conducted with MEGA6 (http://www.megasoftware.net/). *Primary IFN-W5 sequence used for activity assay.