Characterization of a prophenoloxidase from hemocytes of the shrimp Litopenaeus vannamei that is down-regulated by white spot syndrome virus

Abstract

Previously, a prophenoloxidase (proPO) gene (named proPO-a here) from hemocytes of Litopenaeus vannamei was isolated. Here, a proPO-b gene was also identified and characterized from hemocytes of L. vannamei. The cDNA sequences of proPO-a and proPO-b were compared, and it was found that both proPOs had a microsatellite DNA site near the 3′ end of the open reading frame (ORF). However, the microsatellite DNA of proPO-b contained a compound imperfect simple sequence repeats (SSR) ((CT) 38(CA) 8(AA)(CA) 3(TA)(CA) 14), which was different from the perfect one ((CT) 20) of proPO-a, and the cDNA sequences of proPO-a and proPO-b prior to the microsatellite DNA were almost identical, but differed after the microsatellite DNA. ProPO-b (3232 bp) was longer than proPO-a (2471 bp). The 3′ UTR sequence after SSR of proPO-a was not detected in shrimp randomly collected from five different geographically separate populations by reverse-transcription polymerase chain reaction (RT–PCR). On the contrary, the 3′ UTR sequence of proPO-b was detected in all five groups of shrimps. Northern blot analysis showed that a transcript at approximately 3.2 kb, but not 2.5 kb, was detected mainly in hemocytes, and also present in midgut, gill, heart, stomach, posterior midgut cecum, and cuticular epidermis, but no signal was detected in hepatopancreas and musculature. RT–PCR and quantitative real-time RT–PCR analysis showed similar results of the proPO-b expression profile in these shrimp tissues. We also observed that proPO-b expression was down-regulated in shrimp challenged with white spot syndrome virus (WSSV). Our results suggest that proPO-b is a main transcript form of proPO gene in L. vannamei, and it may play a role in defence against WSSV virus.