Structural organization and expression of the gaegurin 4 gene of Rana rugosa

Abstract

Gaegurin 4 (GGN4) is a member of the antimicrobial peptide subfamily isolated from the skin of Rana rugosa. We cloned gDNA encoding GGN4 to study its gene organization and regulation of expression. The GGN4 gene occurs in single copy in the R. rugosa genome and contains a single intron of about 3.4 kb. The transcription start site is located 68 bases upstream of the translation initiation codon. The GGN4 gene was expressed both in Xenopus kidney epithelial cells (A6) and in Xenopus oocytes using the chloramphenicol acetyltransferase reporter gene system. The 5′ flanking region of the GGN4 gene contains a dl binding site that is known to regulate acute phase immune response related gene expression in mammals and insects. The dl protein bound specifically to the GGN4 gene promoter region. Mutants that serially delete the 5′ flanking region show that removal of the dl binding site inhibited GGN4 gene expression in both A6 cells and Xenopus oocytes. From these results, we propose that expression of the GGN4 gene may be regulated by the region containing the dl element which plays a key role in the regulation of antimicrobial peptide genes in Drosophila and mammals.