Re-annotation of the Theileria parva genome refines 53% of the proteome and uncovers essential components of N-glycosylation, a conserved pathway in many organisms

Abstract

<jats:title>Abstract</jats:title><jats:sec><jats:title>Background</jats:title><jats:p>The apicomplexan parasite <jats:italic>Theileria parva</jats:italic> causes a livestock disease called East coast fever (ECF), with millions of animals are at risk in sub-Saharan East and Southern Africa, the geographic distribution of <jats:italic>T. parva</jats:italic>. Over a million bovines die each year of ECF, with a tremendous economic burden to pastoralists in endemic countries. Comprehensive, accurate parasite genome annotation can facilitate the discovery of novel chemotherapeutic targets for disease treatment, as well as elucidate the biology of the parasite. However, genome annotation remains a significant challenge because of limitations in the quality and quantity of the data being used to inform the location and function of protein-coding genes and, when RNA data are used, the underlying biological complexity of the processes involved in gene expression. Here, we apply our recently published RNAseq dataset derived from the schizont life-cycle stage of <jats:italic>T. parva</jats:italic> to update structural and functional gene annotations across the entire nuclear genome.</jats:p></jats:sec><jats:sec><jats:title>Results</jats:title><jats:p>The re-annotation effort lead to evidence-supported updates in over half of all protein-coding sequence (CDS) predictions, including exon changes, gene merges and gene splitting, an increase in average CDS length of approximately 50 base pairs, and the identification of 128 new genes. Among the new genes identified were those involved in N-glycosylation, a process previously thought not to exist in this organism and a potentially new chemotherapeutic target pathway for treating ECF. Alternatively-spliced genes were identified, and antisense and multi-gene family transcription were extensively characterized.</jats:p></jats:sec><jats:sec><jats:title>Conclusions</jats:title><jats:p>The process of re-annotation led to novel insights into the organization and expression profiles of protein-coding sequences in this parasite, and uncovered a minimal N-glycosylation pathway that changes our current understanding of the evolution of this post-translation modification in apicomplexan parasites.</jats:p></jats:sec>

Integral Use of Immunopeptidomics and Immunoinformatics for the Characterization of Antigen Presentation and Rational Identification of BoLA-DR–Presented Peptides and Epitopes

“…The sequence interpretations of mass spectrometry spectra were performed using a database containing all bovine UniProt entries combined with entry P01012 for chicken ovalbumin (total of 41610 entries) and 4084 entries for Theileria parva Muguga proteome (40). The spectral interpretation was performed using de novo -assisted database search with PEAKS 10 (Bioinformatics Solutions), in 'no enzyme' mode, with mass tolerances of 5 ppm for precursor ions and 0.03 Da for fragment ions.…”

Comparative Transcriptomics of the Bovine Apicomplexan Parasite Theileria parva Developmental Stages Reveals Massive Gene Expression Variation and Potential Vaccine Antigens

“…It is likely that the structure of those genes was not correctly identified in the original annotation and thus was incomplete or had gaps in the reference transcriptome used for the mapping, as we demonstrated previously (16). These have now been re-annotated accordingly (44). Most of the 32 genes identified by both schizont and piroplasm contigs of unmapped reads were hypothetical proteins.…”

“…We further checked the integrity of the RNA using 1.2% agarose RNA gel, as described previously (20). The poly(A) + RNA was purified from the total (44).…”

“…The schizont reads mapped to transcripts of 3,891 genes, whereas reads from the piroplasm mapped to transcripts of 3,887 genes. In total, 4,061 different protein coding genes were identified by the combined schizont and piroplasm reads out of a possible 4,084 protein coding genes predicted by the recent re-annotation of the T. parva genome (44).…”

Antigenic Diversity in Theileria parva Populations From Sympatric Cattle and African Buffalo Analyzed Using Long Read Sequencing

“…This study aimed to dissect the genetic relationship between T. parva identified in cattle and African buffalo sampled in and around the Serengeti National Park, Tanzania, an area endemic for T. parva. Long read (PacBio) sequencing was used to obtain sequences from full or near-full length amplicons of genes encoding three antigens recognized by parasite-specific T cells; Tp1, Tp4, and Tp16 (Graham et al, 2006;Tretina et al, 2020;Morrison et al, 2021, submitted), which were selected on the basis of amplifying from T. parva, but not from the closely related T. sp. buffalo.…”