Proteome-wide analysis of lysine 2-hydroxyisobutyrylation in Frankliniella occidentalis

Abstract

Background: Lysine 2-hydroxyisobutyrylation (Khib) is a novel and conserved post-translational modification (PTM). Frankliniella occidentalis are economically important agricultural pests globally and also notorious for vectoring destructive plant viruses. To better study the disease transmission mechanism of F. occidentalis, it is necessary to conduct in-depth analysis of it. So far, no Khib modification of insects has been reported.

Results: In this study, a proteome-wide analysis of Khib modifications in F. occidentalis was analyzed for the first time through the combination of high performance liquid chromatography fractionation technology and 2-hydroxyisobutyrylated peptide enrichment and other advanced technologies, 4093 Khib sites were identified on 1125 modified proteins. Bioinformatics and functional enrichment analyses showed that Khib-modified proteins were significantly enriched in many cell compartments and pathways, especially related to various cellular components and biological processes, and were more concentrated in ribosomes and proteasome subunits, involved in energy metabolism, protein synthesis and degradation, compared to the other nine species including Japonica rice, Homo sapiens, P. patens, Botrytis, Ustilaginoidea virens, Saccharomyces cerevisiae, T. gondii, C. albicans, and F. oxysporum. And Khib sites on virus-interacting insect proteins were discovered for the first time, such as cyclophilin and endoCP-GN.

Conclusions: After three repeated experiments, we found a total of 4093 Khib sites on 1125 proteins. These modified proteins are mainly concentrated in ribosomes and proteasome subunits, and are widely involved in a variety of critical biological activities and metabolic processes of F. occidentalis. In addition, for the first time, Khib modification sites are found on the proteome of F. occidentalis, and these sites could be acted as for the virus interaction, including cyclophilin and endoCP-GN. The global map of 2-hydroxyisobutyrylation in thrips is an invaluable resource to better understand the biological processes of thrips and provide new means for disease control and mitigation of pest damage to crops.

Keywords: Frankliniella occidentalis; Lysine 2-hydroxyisobutyrylation; Pathogenicity; Post-translational modifications; Proteome.

Fig. 1

Identification of 2-hydroxyisobutyrylated proteins in thrips. A Overview of experimental procedures used in this study. B Immunoblot analysis with pan anti-Khib antibody in F. occidentalis, each protein lane represents a biological replicate (Figure S1). C Peptide length distribution. D Mass error distribution of all identified peptides. E Number of modified sites in a protein

Fig. 2

Analysis of the Khib site motifs. A Motif analysis shows Khib peptide motifs and conservation of Khib sites. The intensity map shows enrichment of amino acids in particular positions around Khib lysine residues. B Heat map of the amino acid compositions around Khib sites. Red indicates enrichment and green indicates depletion

Fig. 3

Structural analysis of all 2-hydroxyisobutyrylated proteins and predicted surface accessibility of Khib sites

Fig. 4

Test of the evolutionary conservation degree of Khib in different species. A Proportion of conserved Khib proteins of 9 distant species to total modified proteins. B A pie chart of conservation of Khib sites in 9 organisms

Fig. 5

Pie charts showing the distribution of Khib proteins. A Khib proteins categorized according to the biological process. B Khib proteins categorized according to the cellular component. C Khib proteins categorized according to the molecular function

Fig. 6

Enrichment analysis of 2-hydroxyisobutyrylated proteins in thrips. A, B, C GO enrichment analysis was based on the biological process, cellular component, and molecular function. D KEGG pathway-based enrichment analysis (www.kegg.jp kegg / kegg1.html). E Protein domain enrichment analysis of all identified proteins

Fig. 7

The PPI network of Khib proteins in thrips. Two highly interconnected Khib protein clusters are related to the ribosome and proteasome. These clusters were represented by different colors and black dots

Fig. 8

Three-dimensional structure of two thrip proteins involved in pathogenesis with the identified Khib site. A Cyclophilin. B endoCP-GN. The structure was predicted by I-TASSER. The recognized sites are highlighted in black