The developmental phosphoproteome of Haemonchus contortus

Protein phosphorylation plays essential roles in the biology of cellular processes. Despite recent progress in the genomics, transcriptomics and proteomics of a range of socioeconomically important parasitic nematodes, there is scant phosphoproteomic data to underpin fundamental molecular biological discovery. Here, using the phosphopeptide enrichment-based LC-MS/MS and DIA quantitation, we characterised the first developmental phosphoproteome of the parasitic nematode Haemonchus contortus, one of the most pathogenic parasites of livestock animals. In total, 1804 phosphorylated proteins with 4406 phosphorylation sites from different developmental stages/sexes (i.e. egg, L3 and L4, female (Af) and male (Am) adults) were identified with confidence (false discovery rate: < 0.01; localisation probability: ≥ 75%). Bioinformatic analyses of quantified phosphopeptides (with a single phosphorylation site) exhibited distinctive stage and sex-specific pattern during H. contortus development. A subset of phosphoproteins is proposed to play crucial roles in fundamental biological processes, such as spindle positioning (e.g. LIN-5 and LFI proteins), signal transduction (e.g. IGF-1 signalling related proteins) and kinase activity (e.g., auto-phosphorylated kinases). In addition, a sequence-based comparison of the phosphoproteome of H. contortus with those of free-living nematodes (i.e. Caenorhabditis elegans and Pristionchus pacificus) revealed that protein phosphorylation is likely regulated in a species-specific manner. Our findings infer active roles for protein phosphorylation in the adaptation of a parasitic nematode to a constantly changing external environment. The phosphoproteome data set for H. contortus provides a foundation for a better understanding of phosphorylation and associated biological processes (e.g. regulation of signal transduction), and might enable the discovery of novel anthelmintic targets.