Molecular characterization and gene expression of apolipophorin III from the ghost moth, Thitarodes pui (Lepidoptera, Hepialidae)

Abstract

Apolipophorin III (apoLp-III) functions in lipid transport and immune activation in insects. We cloned a cDNA encoding putative apoLp-III from larvae of Thitarodes pui, a host species of Ophiocordyceps sinensis, with great economic importance in the Tibetan Plateau. Excluding a putative signal peptide of the first 20 amino acid residues, the 171-residue mature apoLp-III has a calculated molecular mass of 18,606 Da. T. pui apoLp-III shares little sequence homologies (<36%) with other apoLp-IIIs. Phylogenetic analysis reveals that T. pui apoLp-III belongs to a distinct, early diverging lineage of lepidopteran apoLp-IIIs. Homology modeling of T. pui apoLp-III shows a bundle of five amphipathic α-helices, including a short helix 3'. T. pui apoLp-III was constitutively expressed in larval fat body at lower levels than pupal and adult fat body. Significant induction of apoLp-III expression, associated with strongest nodulation response, was observed in both sixth and eighth instar larvae challenged with Beauveria bassiana conidia at 1 hr after inoculation, compared with saline-injected controls. The inoculation experiment as well as previous field studies revealed the relative susceptibility of the sixth instar to the entomopathogenic fungus. ApoLp-III transcripts in the infected sixth and eighth instars were found to be induced highest 2- and 14.7-fold, respectively, during the first 12 hr. In late-stage infection, the infected susceptible sixth instar showed decrease in apoLp-III expression followed by production of B. bassiana hyphal bodies, whereas the infected eighth instar showed longer lasting increase in the expression. These results suggest that apoLp-III might contribute to T. pui immune response against fungal pathogens.

De novo transcriptome characterization of the ghost moth, Thitarodes pui, and elevation-based differences in the gene expression of its larvae

“…Third, T. pui is a member of the primitive lepidopteran family Hepialidae (Hebert et al, 2003) and has a long evolution distance in relation to other lepidopteran insects. For example, phylogenetic analysis clustered the heat shock proteins, beta-1,3-glucan recognition proteins, and apolipophorin III proteins of T. pui into a new clade or formed a distinct diverging branch, respectively Zou et al, 2011;Sun et al, 2012). Thus, sequence divergences between T. pui and other lepidopteran species is relatively large, which may have led to the low percentage of matches in the species distribution results.…”

“…Oligonucleotide primers for qRT-PCR were designed using Primer3web version 4.0.0 (http://primer3.ut.ee/) and are detailed in Supplementary Sun et al, 2012). All quantitative PCR runs were repeated in 3 replications.…”

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

“…and/or other posttranslational processing of the mature apoLp-III. Since the fat body is the major site of apoLp-III synthesis in insects [17,19,21,23,39,41] and presence of different apoLp-III forms was demonstrated by us in G. mellonella fat body (this report), it can be suggested that additional apoLp-III forms detected in the hemolymph are generated during and/or after release of apoLp-III from the fat body cells into the hemolymph. Many proteins synthesized as inactive precursors are subjected to limited proteolysis by specific proteases to form their final active structures.…”

CHANGES IN THE HEMOLYMPH PROTEIN PROFILES IN Galleria mellonella INFECTED WITH Bacillus thuringiensis INVOLVE APOLIPOPHORIN III. THE EFFECT OF HEAT SHOCK

“…An increase in apoLp‐III transcripts was observed in T. molitor infected with intracellular pathogen Listeria monocytogenes after 24–72 h. In the same insect, immunization with E. coli caused an increase in apoLp‐III expression 24–72 h after the challenge but no increase was observed 12 h after challenge (Noh et al., ). Upregulation of apoLp‐III expression, followed by its decrease at later time‐points was observed in Thitarodes pui after infection with fungus Beauveria bassiana (Sun et al., ). On the other hand no change in apoLp‐III expression was observed in infected Anopheles gambiae (Mendes et al., ).…”