Comparative Subcellular Proteomics Analysis of Susceptible and Near-isogenic Resistant Bombyx mori (Lepidoptera) Larval Midgut Response to BmNPV infection

Abstract

The molecular mechanism of silkworm resistance to Bombyx mori nucleopolyhedrovirus (BmNPV) infection remains largely unclear. Accumulating evidence suggests that subcellular fractionation combined with proteomics is an ideal technique to analyse host antiviral mechanisms. To clarify the anti-BmNPV mechanism of the silkworm, the near-isogenic line BC9 (resistant strain) and the recurrent parent P50 (susceptible strain) were used in a comparative subcellular proteomics study. Two-dimensional gel electrophoresis (2-DE) combined with mass spectrometry (MS) was conducted on proteins extracted from the cytosol, mitochondria, and microsomes of BmNPV-infected and control larval midguts. A total of 87 proteins were successfully identified from the three subcellular fractions. These proteins were primarily involved in energy metabolism, protein metabolism, signalling pathways, disease, and transport. In particular, disease-relevant proteins were especially changed in microsomes. After infection with BmNPV, differentially expressed proteins (DEPs) primarily appeared in the cytosolic and microsomal fractions, which indicated that these two fractions might play a more important role in the response to BmNPV infection. After removing genetic background and individual immune stress response proteins, 16 proteins were identified as potentially involved in repressing BmNPV infection. Of these proteins, the differential expression patterns of 8 proteins according to reverse transcription quantitative PCR (RT-qPCR) analyses were consistent with the 2-DE results.

Specific genes related to nucleopolyhedrovirus in Bombyx mori susceptible and near‐isogenic resistant strains through comparative transcriptome analysis

“…With the use of recently emerging data, researchers have begun to uncover the antiviral mechanisms in invertebrate immunity, particularly by screening for antiviral genes (Xia et al ., ; Bao et al ., ; Zhou et al ., ; Lekha et al ., ; Wang et al ., ; ; ; Xu et al ., ; Li et al ., ; Yu et al ., ). Whether these genes are resistance‐related genes remains unclear, however, and the exact antiviral mechanisms remain unknown.…”

“…A comparative subcellular proteomics study on the susceptible strain, P50 and the derived resistant near‐isogenic strain BC9 showed that 16 proteins were potentially involved in repressing BmNPV infection (Wang et al ., ). Comparative genomics, transcriptomics and proteomics analyses also revealed that numerous genes and proteins are possibly involved in resistance against BmNPV (Bao et al ., , , ; Zhou et al ., ; Wang et al ., , ; Li et al ., ; Yu et al ., ). Among these, V‐ATPase and Bm‐SP142 are upregulated in resistant strains (Lü et al ., ; Li et al ., ).…”

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

“…Two-dimensional electrophoresis and mass spectrometry (MS) techniques, isobaric tags for relative and absolute quantification (iTRAQ), Tandem Mass Tags (TMT) and label-free methods are attractive proteomic analysis technologies for researching the interaction between pathogens and B. mori. Wang et al [12] performed a comparative subcellular proteomics analysis of the silkworm larval midgut response to BmNPV infection by 2-DE combined with MS. The iTRAQ method was also used to explore the molecular mechanisms of B. mori response to B. mori cypovirus (BmCPV) [13] and BmNPV [14] challenge.…”

“…Furthermore, a GP64binding protein E3 ubiquitin-protein ligase SINA-like 10 (SINAL10) [20] and chaperonin containing t-complex polypeptide 1 (TCP-1) [21] could stimulate BmNPV proliferation in BmN cells. 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.…”

Immune function of a Rab‐related protein by modulating the JAK‐STAT signaling pathway in the silkworm, Bombyx mori

“…It is also a lepidopteran model organism for genetic and immunology research (Liu, Ling, & Wu, 2009;Rao et al, 2016;Wang et al, 2017). In silkworms, the Rab protein mainly functions in the development of neurons (Uno et al, 2010(Uno et al, , 2013(Uno et al, , 2014(Uno et al, , 2016(Uno et al, , 2017.…”