Characterization of a homologue of the Rel/NF-kappaB transcription factor from a beetle, Allomyrina dichotoma.
A cDNA encoding a Rel/NF-kappaB homologue was cloned from a beetle, Allomyrina dichotoma, by reverse transcriptase-polymerase chain reactions (RT-PCR) taking advantage of the conserved Rel homology domain (RHD) to synthesize primers. The Rel/NF-kappaB homologue was designated A. dichotoma (A.d.) Rel A. The amino acid sequence of the A.d. Rel A RHD was compared with those of insect RHDs. The result showed that it has 70% identity with Tribolium castaneum Dorsal, 66% with Drosophila melanogaster Dorsal, 61% with Anopheles gambiae Gambif1, and 55% with D. melanogaster Dif. A putative phosphorylation site in the RHD, RRPS, and two putative nuclear localization signals were conserved in A.d. Rel A. A recombinant fusion protein containing the A.d. Rel A RHD was confirmed to bind specifically to the NF-kappaB site of a gene encoding A.d. coleoptericin A, an antibacterial peptide from A. dichotoma. The activity of A.d. Rel A in modulating a gene construct of the A.d. coleoptericin A promoter-luciferase reporter by expressing the A.d. coleoptericin A cDNA in a Bombyx mori cell line was analyzed. The result showed that A.d. Rel A strongly activates the A.d. coleoptericin A gene construct, whereas A.d. Rel A failed to activate the gene construct containing the mutated NF-kappaB site, suggesting the importance of the interaction between the NF-kappaB site and A.d. Rel A in the signal transduction for gene expression of antibacterial peptides in A. dichotoma.
Sagisaka
Aki
A
Innate Immunity Laboratory, National Institute of Agrobiological Sciences, Owashi 1-2, Tsukuba, Ibaraki 305-8634, Japan.
Tanaka
Hiromitsu
H
Furukawa
Seiichi
S
Yamakawa
Minoru
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D012831:Bombyx; D002460:Cell Line; D003001:Cloning, Molecular; D001517:Coleoptera; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D004330:Drosophila; D005982:Glutathione Transferase / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014162:Transfection
D017931:DNA Primers; D018076:DNA, Complementary; D016328:NF-kappa B; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D008156:Luciferases; D005982:Glutathione Transferase