Butterfly wings, a new site of porphyrin synthesis and cleavage: Studies on the expression of the lipocalin bilin-binding protein in Pieris brassicae

The bilin-binding protein (BBP), a member of the lipocalin protein superfamily, is synthesized mainly in last instar larvae and in late pupae and newly emerged adults of Pieris brassicae, as previously reported. Here we present results from Northern blot analysis of the BBP gene transcript and from in vitro studies of holo-BBP biosynthesis with isolated wings using [14C]5-aminolevulinic acid as a precursor to the bilin ligand, [14C]-amino acids to label the apo-protein and inhibitors for both processes. Our combined data clearly demonstrate that BBP, which accumulates around pupa-adult transformation, is produced as holoprotein in the developing wings, while the BBP gene transcript is no longer detected in the rest of the body. Forewings and hind wings behave markedly different as the latter represent the major site of BBP synthesis, in agreement with the unequal distribution of BBP in the wings. The presence of an active pathway of porphyrin synthesis and cleavage in insect wings, shown here for the first time, and the role of the biliprotein during wing development remains an enigma so far. As part of this work sequences of fragments of the genes for actin and glyceraldehyde-3-phosphate dehydrogenase were obtained and examined as reference house-keeping genes in the expression studies.