AUTHOR=Hrithik Md Tafim Hossain , Hong Jooan , Kim Yonggyun 

TITLE=Identification of four secretory phospholipase A2s in a lepidopteran insect, Acrolepiopsis sapporensis, and their functional association with cellular immune responses

JOURNAL=Frontiers in Endocrinology

VOLUME=14

YEAR=2023

URL=https://www.frontiersin.org/journals/endocrinology/articles/10.3389/fendo.2023.1190834

DOI=10.3389/fendo.2023.1190834

ISSN=1664-2392

ABSTRACT=<sec><title>Background</title><p>Eicosanoids are a group of the oxygenated C20 polyunsaturated fatty acids and play crucial roles in mediating various insect physiological processes. Catalytic activity of phospholipase A<sub>2</sub> (PLA<sub>2</sub>) provides an initial substrate, arachidonic acid (AA), for subsequent eicosanoid biosynthesis.</p></sec><sec><title>Results</title><p>This study identified four different secretory PLA<sub>2</sub> (<italic>As-PLA<sub>2</sub>A</italic>–<italic>As-PLA<sub>2</sub>D</italic>) genes encoded in the Asian onion moth, <italic>Acrolepiopsis sapporensis</italic>. A phylogenetic analysis indicated that <italic>As-PLA<sub>2</sub>A</italic> and <italic>As-PLA<sub>2</sub>D</italic> are clustered with Group III PLA<sub>2</sub>s while <italic>As-PLA<sub>2</sub>B</italic> and <italic>As-PLA<sub>2</sub>C</italic> are clustered with Group XII and Group X PLA<sub>2</sub>s, respectively. Expression levels of these PLA<sub>2</sub> genes increased along with larval development, especially in the fat body. A bacterial immune challenge upregulated the basal expression levels of the four PLA<sub>2</sub> genes, which resulted in significant increases of the PLA<sub>2</sub> enzyme activity. The enzyme activity was susceptible to a calcium chelator or reducing agent, suggesting Ca<sup>2+</sup> dependency and disulfide linkage required for the catalytic activities of the secretory type of PLA<sub>2</sub>s. In addition, the PLA<sub>2</sub> activity was also susceptible to bromophenacyl bromide (BPB), a specific inhibitor to sPLA<sub>2</sub>, but not to intracellular PLA<sub>2</sub> inhibitors. An addition of BPB to the immune challenge significantly prevented hemocyte-spreading behavior of <italic>A. sapporensis</italic>. BPB treatment also suppressed a cellular immune response measured by hemocyte nodule formation. However, the immunosuppression was significantly rescued by the AA addition. To determine the PLA<sub>2</sub>(s) responsible for the immunity, individual RNA interference (RNAi) treatments specific to each of the four PLA<sub>2</sub>s were performed. Injection of gene-specific double-stranded RNAs caused significant reductions in the transcript level in all four PLA<sub>2</sub>s. In all four PLA<sub>2</sub>s, the RNAi treatments prevented the cellular immune response even after the immune challenge.</p></sec><sec><title>Conclusion</title><p>This study reports four secretory PLA<sub>2</sub>s encoded in <italic>A. sapporensis</italic> and their function in mediating cellular immunity.</p></sec>