Proteomics analysis reveals the differential protein expression of female and male adult Toxocara canis using Orbitrap Astral analyzer

Abstract

Background: Toxocara canis, the most prevalent helminth in dogs and other canines, is one of the socioeconomically important zoonotic parasites, particularly affecting pediatric and adolescent populations in impoverished communities. However, limited information is available regarding the proteomes of female and male adult T. canis. To address this knowledge gap, we performed a comprehensive proteomic analysis to identify the proteins with differential abundance (PDAs) and gender-specifically expressed proteins between the two sexes adult T. canis.

Methods: The comparative proteomic analysis was carried out by the Orbitrap mass spectrometry (MS) with asymmetric track lossless (Astral) analyzer. The difference analysis was conducted using t-test and the proteins verification was achieved through parallel reaction monitoring (PRM). The potential biological functions of identified adult T. canis proteins and PDAs were predicted by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. The domain, transcription factor and subcellular localization of the identified proteins and PDAs were analyzed by InterPro, AnimalTFDB 4.0 and Cell-mPLOC 2.0 databases, respectively.

Results: A total of 8565 somatic proteins of adult T. canis were identified. Compared to male adult, 682 up-regulated PDAs and 844 down-regulated PDAs were identified in female adult with P-values < 0.05 and |log₂^FC| > 1, including 139 proteins exclusively expressed in female and 272 proteins exclusively expressed in male. The GO annotation analysis using all PDAs revealed that the main biological processes, cellular components and molecular functions corresponded to aminoglycan metabolic process, extracellular region and protein tyrosine phosphatase activity, respectively. The KEGG analysis using all PDAs showed that the pathways were mainly associated with adipocytokine signaling pathway, proximal tubule bicarbonate reclamation and PPAR signaling pathway.

Conclusions: This study reveals the differential protein expression between female and male adult T. canis, providing valuable resource for developing the novel intervention strategies against T. canis infection in humans and animals, especially from the perspective of sexual development and reproduction.

Keywords: Toxocara canis; Orbitrap Astral; Proteomics; Reproduction; Toxocariasis; Zoonosis.

Fig. 1

The quantification and validation of adult Toxocara canis proteins. a The number of T. canis proteins identified from each sample using narrow-window data-independent acquisition (nDIA). b The principal component analysis (PCA) of proteome data among the six samples. c The quantitation of five T. canis proteins in each sample using parallel reaction monitoring (PRM). d The verification of T. canis proteins identified through nDIA analysis using PRM

Fig. 2

The functional prediction of adult Toxocara canis proteins. a The Venn diagrams showing the common and exclusive proteins annotated by GO, KEGG and IPR database. The top 30 enriched GO terms (including BP, CC and MF categories) (b), all enriched KEGG terms of level 2 signaling pathways (c) and top 20 protein domain (d) of all identified adult T. canis proteins

Fig. 3

The top 20 transcription factors (TFs) families (a) and the subcellular localization (b) of all identified adult Toxocara canis proteins

Fig. 4

The volcano plots (a) and heatmaps (b) showing the proteins with differential abundance (PDAs) identified between female and male adult Toxocara canis

Fig. 5

The functional prediction of proteins with differential abundance (PDAs) identified between female and male adult Toxocara canis. Scatter plots of the top 20 significantly enriched (P-values < 0.05) GO terms (a), top 20 significantly enriched (P-values < 0.05) KEGG pathways (b) and top 20 protein domain (c) of PDAs. The X-axis label represents the rich factor; the Y-axis label shows the terms name. The rich factor reflects the proportion of PDAs in each term. The greater the rich factor, the greater the degree of term enrichment. The color of the dots represents the enrichment score [− log₁₀^(Pvalue)], where red color indicates high enrichment, while green color indicates low enrichment. Dot size represents the number of PDAs in the corresponding term (bigger dots indicate larger PDAs numbers). d The subcellular localization of PDAs