A secreted phospholipase A₂ (BmsPLA₂) regulates melanization of immunity through BmDDC in the silkworm Bombyx mori

Abstract Insect immune‐associated phospholipase A 2 (PLA 2 ) is an important target of pathogen invasion. Melanization, an effective defense response, has significant correlations with other immune responses to coordinate immune attack against invaders. However, the effect of PLA 2 on melanization has not yet been reported in insects or other arthropods. In this work, we cloned a PLA 2 gene ( BmsPLA 2 ), and its protein had characteristic features of secreted PLA 2 (sPLA 2 ). After injection of bacteria, BmsPLA 2 expression and sPLA 2 activity in hemolymph significantly increased. BmsPLA 2 fluorescence was transferred from the cytoplasm to the cell membranes of circulating hemocytes. These results indicated that BmsPLA 2 was related to hemolymph immunity in silkworms. Interestingly, reducing BmsPLA 2 by RNA interference decreased melanosis (melanistic hemocytes) levels in vivo and in vitro , while BmsPLA 2 overexpression had the opposite effect. The larval survival and melanization rate in the hemocoel both slowed depending on the PLA 2 inhibitor dosage. These results demonstrated that BmsPLA 2 plays a role in melanization during the immune process of silkworms. Surprisingly, the level of BmDDC matched the degree of melanization in various observations. BmDDC expression showed a significant increase, with the peak occurring later than that of BmsPLA 2 after injection of bacteria, implying that BmsPLA 2 was activated prior to BmDDC . Moreover, the alteration of BmsPLA 2 by RNA interference or overexpression led to altered BmDDC levels. These results suggested that BmsPLA 2 regulates the melanization response in silkworms through BmDDC . Our study proposes a new regulatory mechanism of the melanization response and new directions for understanding the complex immune networks of insects.