Molecular cloning of two prophenoloxidase genes from the mosquito Aedes aegypti

The biosynthesis of melanotic materials is an important process in the life of a mosquito. Melanin production is critical for many diverse processes s uch as egg chorion tanning, cuticular sclerotization, and melanotic encapsu lation of metazoan parasites. Prophenoloxidase plays a critical role in thi s biochemical cascade. Two cDNAs, one full length and one partial clone, en d two genomic clones encoding prophenoloxidase (pro-PO) were isolated from the yellow fever mosquito, Aedes aegypti. The full-length cDNA, pAaProPO1, is 2286 bp long with a 2055 bp open reading frame encoding a 685 amino acid protein that shares 89% identity with Armigeres subalbatus pro-PO. It cont ains two putative copper binding domains (amino acids 197-243 and 346-423) that are homologous to other insect pro-POs. AaProPO1 messenger RNA (mRNA) was detected by reverse transcription polymerase chain reaction (RT-PCR) on ly from third-stage larvae and not in adult mosquitoes after blood feeding, during the melanotic encapsulation of Dirofilaria immitis microfilariae or following exposure to bacteria. A 750 bp fragment of the second cDNA (pAaP roPO2) was cloned using RT-PCR from mRNA obtained from 14-day postovipostio nal eggs. AaProPO2 mRNA was not found in any other life stages, and may be in low abundance or transiently expressed. AaProPO2 and AaProPO1 each conta in three introns that are 60, 68 and 58 bp and 61, 69 and 59 bp long, respe ctively, and the intron sequences of these two genes are not similar.