Molecular cloning of a second prophenoloxidase cDNA from the mosquito Armigeres subalbatus: prophenoloxidase expression in blood-fed and microfilariae-inoculated mosquitoes.
Melanization constitutes an important component in various aspects of insect life, including cuticular sclerotization, egg-shell tanning, melanization of parasites and wound healing. Recently, a cDNA encoding prophenoloxidase (pro-PO), a key enzyme in the biosynthesis of melanotic material in insects, was cloned from microfilariae (mf)-inoculated mosquitoes, Armigeres subalbatus. However, results of Northern blot analyses indicated that two pro-POs might be present in Ar. subalbatus and these pro-POs might be responsible for two distinct physiological functions, egg-shell tanning and melanization of parasites. Subsequently, the second pro-PO cDNA (As-pro-PO II) was cloned from blood-fed Ar. subalbatus by rapid amplification of cDNA ends polymerase chain reaction. The 2210 bp As-pro-PO II cDNA contains a 41 bp 5'-non-coding region, a 2064 bp open reading frame and a 105 bp 3'-non-coding region. A hydrophobic signal peptide for endoplasmic reticulum targeting is not found in the N-terminal region. The deduced amino acid sequence of As-pro-PO II shares a high degree of identity (81.5%) with that of the As-pro-PO I obtained from mf-inoculated Ar. subalbatus. Both Northern blot and reverse transcription polymerase chain reaction analysis demonstrated that these two mosquito pro-POs are persistently expressed in mosquito haemocytes and not in fat body, midgut, or ovaries. The expression of As-pro-PO I and As-pro-PO II in mosquitoes is associated with melanization of mf and blood feeding, respectively.
Department of Parasitology, National Yang-Ming University, Shihpai, Taipei 112, Taiwan.
L H
LH
Huang
B M
BM
Christensen
C C
CC
Chen
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern / Q000379:methods; D004156:Catechol Oxidase / Q000235:genetics*; D003001:Cloning, Molecular; D009033:Culicidae / Q000201:enzymology* / Q000235:genetics / Q000469:parasitology; D018076:DNA, Complementary; D004792:Enzyme Precursors / Q000235:genetics*; D005247:Feeding Behavior; D005260:Female; D015870:Gene Expression; D008842:Microfilariae / Q000502:physiology*; D008969:Molecular Sequence Data; D012333:RNA, Messenger; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
D018076:DNA, Complementary; D004792:Enzyme Precursors; D012333:RNA, Messenger; C038451:pro-phenoloxidase; D004156:Catechol Oxidase