Cloning and expression study of anIRF4agene and its two transcript variants in turbot,Scophthalmus maximus

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Abstract

Interferon regulatory factor 4 (IRF4) is known to be involved in antiviral response as well as regulation of functional and developmental processes in lymphomyeloid cell lineages in mammals. In this study, the gene of IRF4a and its two transcript variants (named IRF4a1 and -2) were cloned from turbot, Scophthalmus maximus, the tissue distributions and in vivo immune responsive expression patterns of the two transcripts were subsequently examined. The Scophthalmus maximus (Sm)IRF4a gene is 8367 nucleotide (nt) in length, consisting of eight exons and seven introns. The SmIRF4a1 transcript is 3185 nt long, containing an open reading frame (ORF) of 1401 nt that encodes a polypeptide of 466 amino acids (aa). The SmIRF4a2 transcript is 2265 nt long and identical with the SmIRF4a1 from position 1 to 1171, containing an ORF of 1164 nt that encodes a truncated protein of 387 aa as a result of a frame shift in exon 6 which introduces a premature stop codon. The deduced aa sequence of SmIRF4a1 posses a DNA-binding domain (DBD), a nuclear localization signal (NLS), a serine-rich domain (SRD) and an IRF association domain (IAD), while SmIRF4a2 lacks the C-terminal 52 residues of the IAD and the downstream C-terminal extension, instead, they are replaced by a 8-aa segment although the three upstream domains are intact. Quantitative real-time PCR analysis revealed a broad tissue expression for both SmIRF4a1 and -2 with the former showing a significantly higher expression in all examined tissues except skin. Expressions of two transcript variants after stimulation with polyinosinic:polycytidylic acid [poly(I:C)] and turbot reddish body iridovirus (TRBIV) were tested in gills, spleen, head kidney and muscle. A two-wave of induced expression pattern was observed for both transcripts with either stimulus treatment during a 7-day time course. SmIRF4a2 responded more promptly to the stimuli and showed a higher level of inducibility in the early phase while SmIRF4a1 was strongly detected in the later phase. These data suggest an important role of SmIRF4a2 in the fast immune response under a background of SmIRF4a1-dominant antiviral response in the IRF4a system of turbot.

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