Identification and characterization of a prophenoloxidase-1 (PPO1) cDNA in the cabbage butterfly Pieris rapae L.

摘要

Phenoloxidase (PO) is a melanin-synthesizing enzyme widely distributed in animals, plants and microorganisms. It plays an important role in insect immune response. Here we report the cloning of a prophenoloxidase (PPO) cDNA from Pieris rapae, prophenoloxidase-1 (PrPPO1). The full-length cDNA consisted of 3338bp, containing an open reading frame of 2049bp encoding 682 amino acid (aa) residues. Two putative copper-binding sites, a proteolytic activation site, three conserved hemocyanin domains and a thiolester motif, but no signal sequence, were found in the deduced aa sequence. A BLASTp search and neighbor-joining analysis showed the deduced aa sequence had high identity to the published sequence of PPO1 from other lepidopteran insects, ranging from 71.31 to 73.15%. The results of real-time polymerase chain reaction (PCR) showed that the highest expression of PrPPO1 was in the 4th instar larvae and the highest density of oenocytoids was observed in the 4th instar larvae also. PrPPO1 transcripts in the 4th instar larvae were significantly decreased at 6h and 12h with Beauveria bassiana infection; however, they were sharply increased at 72h. PrPPO1 transcriptional level in the 5th instar larvae showed no significant changes compared with the control at 4h with quercetin injection; however it was significantly increased at 12h. The expression level of PrPPO1 was mainly related to the treatment time of the quercetin, but not the dose.