Proteomics analysis of digestive juice from silkworm during Bombyx mori nucleopolyhedrovirus infection

Abstract

Previous studies have analyzed the midgut transcriptome and proteome after challenge with Bombyx mori nucleopolyhedrovirus (BmNPV), however little information is available on the digestive juice proteome after BmNPV challenge. This study investigated BmNPV infection-induced protein changes in the digestive juice of silkworms using shotgun proteomics and MS sequencing. From the digestive juice of normal third-day, fifth-instar silkworm larvae, 75 proteins were identified, 44 of which were unknown; from larvae 6 h after inoculation with BmNPV, 106 proteins were identified, of which 39 were unknown. After BmNPV challenge, more secreted proteins appeared that had antiviral and digestive features. GO annotation analysis clustered most proteins in the lumen into catalytic, binding, and metabolic processes. Numerous proteins were reported to have BmNPV interactions. Hsp70 protein cognate, lipase-1, and chlorophyllide A-binding protein precursor were upregulated significantly after BmNPV challenge. Levels of trypsin-like serine protease, beta-1,3-glucanase, catalase, and serine protease transcripts decreased or were not significantly change after BmNPV challenge. Taken together, these findings provided insights into the interaction between host and BmNPV and revealed potential functions of digestive juice after per os BmNPV infection.

Quantitative label-free proteomic analysis reveals differentially expressed proteins in the digestive juice of resistant versus susceptible silkworm strains and their predicted impacts on BmNPV infection

“…Included in that total number are 72, 82, 82 and 87 proteins identified from P50-, P50+, A35-and A35+ strains, respectively. These results demonstrate that proteins derived from digestive juice mainly consist of enzymes, and are consistent with previous reports in B. mori and Helicoverpa armigera [5,6]. Compared to studies by Hu et al [6] which investigated protein expression in the digestive juice of silkworms, > 45 and 15 proteins were identified by Label-free quantitative approaches in P50-and P50+, respectively.…”

“…The BmNPV T3 strain was maintained in our laboratory and purified according to the protocol reported by Rahman et al [23]. The third day of fifth instar larvae were selected for oral feeding experiment according to Hu et al [6], and in each of the three independent experiments were administered 5 μL of BmNPV suspended in water (1 × 10 7 OBs/mL) per os and the control was treated with 5 μL of sterile water.…”

“…Digestive juice proteins are synthesized by midgut cells and secreted into the midgut lumen. To analyze the level of transcription of the genes corresponding to proteins in digestive juice, the cDNA of midgut RNA was used as a template for qPCR reactions according to Hu et al [6]. To measure between protein and transcript abundance, RT-qPCR was conducted to analyze transcript levels of nine identified DEPs, selected from the 80 total DEPs identified as an accumulation of those identified in all four groups (Fig.…”

“…peptides, carbohydrates, and lipids that are absorbed by epithelial cells and enter the circulatory system [6]. Previous studies have indicated that the enzymes form digestive juice not only function to digest food, but also play an important role in weakening or killing pathogens [5].…”

“…Some studies regarding the role of the B. mori larval midgut in response to BmNPV infection have been conducted in previous studies utilizing RNA-Seq [22], 2-DE combined MS [12], label free [16] and iTRAQ methods [14]. Hu et al [6] investigated changes in the protein composition of the silkworm digestive juice between non-infected and infected samples by shotgun and MS techniques. Their research, however, only involved the P50 susceptible strain.…”