Different forms of apolipophorin III in Galleria mellonella larvae challenged with bacteria and fungi.

Apolipophorin III (apoLp-III), a lipid-binding protein and an insect homolog of human apolipoprotein E, plays an important role in lipid transport and immune response in insects. In the present study, we have demonstrated a correlation in time between changes in the apoLp-III abundance occurring in the hemolymph, hemocytes, and fat body after immunization of Galleria mellonella larvae with Gram-negative bacteria Escherichia coli, Gram-positive bacteria Micrococcus luteus, yeast Candida albicans, and a filamentous fungus Fusarium oxysporum. Using two-dimensional electrophoresis (IEF/SDS-PAGE) and immunoblotting with anti-apoLp-III antibodies, the profile of apoLp-III forms in G. mellonella larvae challenged with the bacteria and fungi has been analyzed. Besides the major apoLp-III protein (pI=6.5), one and three additional apoLp-III forms differing in the pI value have been detected, respectively, in the hemolymph, hemocytes, and fat body of non-immunized insects. Also, evidence has been provided that particular apoLp-III-derived polypeptides appear after the immune challenge and are present mainly in the hemolymph and hemocytes. The time of their appearance and persistence in the hemolymph was dependent on the pathogen used. At least two of the apoLp-III forms detected in hemolymph bound to the microbial cell surface. The increasing number of hemolymph apoLp-III polypeptides and differences in their profiles observed in time after the challenge with different immunogens confirmed the important role of apoLp-III in discriminating between pathogens by the insect defense system and in antibacterial as well as antifungal immune response.