Identification and characterization of alternative splicing in parasitic nematode transcriptomes

Abstract

BackgroundAlternative splicing (AS) of mRNA is a vital mechanism for enhancing genomic complexity in eukaryotes. Spliced isoforms of the same gene can have diverse molecular and biological functions and are often differentially expressed across various tissues, times, and conditions. Thus, AS has important implications in the study of parasitic nematodes with complex life cycles. Transcriptomic datasets are available from many species, but data must be revisited with splice-aware assembly protocols to facilitate the study of AS in helminthes.MethodsWe sequenced cDNA from the model worm Caenorhabditis elegans using 454/Roche technology for use as an experimental dataset. Reads were assembled with Newbler software, invoking the cDNA option. Several combinations of parameters were tested and assembled transcripts were verified by comparison with previously reported C. elegans genes and transcript isoforms and with Illumina RNAseq data.ResultsThoughtful adjustment of program parameters increased the percentage of assembled transcripts that matched known C. elegans sequences, decreased mis-assembly rates (i.e., cis- and trans-chimeras), and improved the coverage of the geneset. The optimized protocol was used to update de novo transcriptome assemblies from nine parasitic nematode species, including important pathogens of humans and domestic animals. Our assemblies indicated AS rates in the range of 20-30%, typically with 2-3 transcripts per AS locus, depending on the species. Transcript isoforms from the nine species were translated and searched for similarity to known proteins and functional domains. Some 21 InterPro domains, including several involved in nucleotide and chromatin binding, were statistically correlated with AS genetic loci. In most cases, the Roche/454 data explored in this study are the only sequences available from the species in question; however, the recently published genome of the human hookworm Necator americanus provided an additional opportunity to validate our results.ConclusionsOur optimized assembly parameters facilitated the first survey of AS among parasitic nematodes. The nine transcriptome assemblies, their protein translations, and basic annotations are available from Nematode.net as a resource for the research community. These should be useful for studies of specific genes and gene families of interest as well as for curating draft genome assemblies as they become available.

Dynamic Changes in the Global Transcriptome and MicroRNAome Reveal Complex miRNA-mRNA Regulation in Early Stages of the Bi-Directional Development of Echinococcus granulosus Protoscoleces

“…This ratio is very close to the proportions recorded for Dictyocaulus viviparus (34.6%) (Cantacessi et al, 2010) and Cooperia oncophora (33.1%) (Heizer et al, 2013), but is significantly higher than that reported for C. elegans (17%) in WormBase (Yook et al, 2012). This result is consistent with observations of parasitic nematode transcriptomes (Abubucker et al, 2014), which may be due to a reduction in the number of functional genes in parasite genomes (Zheng H. et al, 2013) or the increased genomic complexity that may be required to interact with multiple hosts/vectors (Abubucker et al, 2014).…”

The genome of the soybean cyst nematode (Heterodera glycines) reveals complex patterns of duplications involved in the evolution of parasitism genes

“…By merging genomics and transcriptomics data, we can provide important insights into the molecular mechanisms regulated through alternative splicing. Alternative splicing is highly prevalent within H. glycines, echoing similar observations in other nematode species [48].…”

RNA-Sequencing of Heterorhabditis nematodes to identify factors involved in symbiosis with Photorhabdus bacteria

“…In addition, unique transcripts in symbiotic nematodes might also result from alternative splicing (AS) under specific physiological condition (here presence of bacterial symbionts). Nematodes are known to have AS in the range of 20–30%, typically with 2–3 transcripts per AS locus [ 31 ]. However, a deeper analysis of the AS in our experimental conditions is beyond the scope of this study and will be taken up in future.…”