Allograft inflammatory factor 1 functions as a pro-inflammatory cytokine in the oyster, Crassostrea ariakensis.

R Lee Mosley,Ting Xu,Xinzhong Wu,Jiasong Xie,Baojian Zhu,Xiao Liu

doi:10.1371/journal.pone.0095859

Abstract

The oyster Crassostrea ariakensis is an economically important bivalve species in China, unfortunately it has suffered severe mortalities in recent years caused by rickettsia-like organism (RLO) infection. Prevention and control of this disease is a priority for the development of oyster aquaculture. Allograft inflammatory factor-1 (AIF-1) was identified as a modulator of the immune response during macrophage activation and a key gene in host immune defense reaction and inflammatory response. Therefore we investigated the functions of C. ariakensis AIF-1 (Ca-AIF1) and its antibody (anti-CaAIF1) in oyster RLO/LPS-induced disease and inflammation. Ca-AIF1 encodes a 149 amino acid protein containing two typical Ca2+ binding EF-hand motifs and shares a 48–95% amino acid sequence identity with other animal AIF-1s. Tissue-specific expression analysis indicates that Ca-AIF1 is highly expressed in hemocytes. Significant and continuous up-regulation of Ca-AIF1 is detected when hemocytes are stimulated with RLO/LPS (RLO or LPS). Treatment with recombinant Ca-AIF1 protein significantly up-regulates the expression levels of LITAF, MyD88 and TGFβ. When anti-CaAIF1 antibody is added to RLO/LPS-challenged hemocyte monolayers, a significant reduction of RLO/LPS-induced LITAF is observed at 1.5–12 h after treatment, suggesting that interference with Ca-AIF1 can suppress the inflammatory response. Furthermore, flow cytometric analysis indicated that anti-CaAIF1 administration reduces RLO/LPS-induced apoptosis and necrosis rates of hemocytes. Collectively these findings suggest that Ca-AIF1 functions as a pro-inflammatory cytokine in the oyster immune response and is a potential target for controlling RLO infection and LPS-induced inflammation.

Highlights

Invertebrates lack adaptive immunity, but their immune systems are capable of effective immune responses and are complex and diverse [1,2]
Blastp analysis of the deduced amino acid sequence revealed that it is a homologue of Allograft inflammatory factor-1 (AIF-1) protein similar to that identified in other species
The results show that the rickettsia-like organism (RLO)/ LPS challenge can up-regulate the expression level of LITAF in hemocytes, while administration of anti-CaAIF1 reduces this effect

Summary

Introduction

Invertebrates lack adaptive immunity, but their immune systems are capable of effective immune responses and are complex and diverse [1,2]. C. ariakensis is one of the most economically important oysters cultivated in southeastern China, especially in Guangxi, Guangdong and Fujian provinces Aquaculture of this species has suffered from massive infection and severe mortalities caused by the pathogen rickettsia-like organism (RLO) [7,8]. We constructed a RLO-challenged oyster hemocyte cDNA library [22], from which a high mobility group box (HMGB) homolog Ca-HMGB was identified and found to function as a proinflammatory cytokine in oyster immune reaction [23]. We reported another protein identified from the hemocyte cDNA library, a novel allograft inflammatory factor 1 (AIF-1), which we named Ca-AIF1. The ability of the anti-CaAIF1 antibody treatment to reduce the oyster inflammatory reaction and to reduce hemocyte apoptosis and necrosis upon exposure to RLO/LPS challenge were analyzed

Materials and Methods

Results

Discussion