Genome-wide transcriptome profiling and spatial expression analyses identify signals and switches of development in tapeworms

Abstract

BackgroundTapeworms are agents of neglected tropical diseases responsible for significant health problems and economic loss. They also exhibit adaptations to a parasitic lifestyle that confound comparisons of their development with other animals. Identifying the genetic factors regulating their complex ontogeny is essential to understanding unique aspects of their biology and for advancing novel therapeutics. Here we use RNA sequencing to identify up-regulated signalling components, transcription factors and post-transcriptional/translational regulators (genes of interest, GOI) in the transcriptomes of Larvae and different regions of segmented worms in the tapeworm Hymenolepis microstoma and combine this with spatial gene expression analyses of a selection of genes.ResultsRNA-seq reads collectively mapped to 90% of the > 12,000 gene models in the H. microstoma v.2 genome assembly, demonstrating that the transcriptome profiles captured a high percentage of predicted genes. Contrasts made between the transcriptomes of Larvae and whole, adult worms, and between the Scolex-Neck, mature strobila and gravid strobila, resulted in 4.5–30% of the genes determined to be differentially expressed. Among these, we identified 190 unique GOI up-regulated in one or more contrasts, including a large range of zinc finger, homeobox and other transcription factors, components of Wnt, Notch, Hedgehog and TGF-β/BMP signalling, and post-transcriptional regulators (e.g. Boule, Pumilio). Heatmap clusterings based on overall expression and on select groups of genes representing ‘signals’ and ‘switches’ showed that expression in the Scolex-Neck region is more similar to that of Larvae than to the mature or gravid regions of the adult worm, which was further reflected in large overlap of up-regulated GOI.ConclusionsSpatial expression analyses in Larvae and adult worms corroborated inferences made from quantitative RNA-seq data and in most cases indicated consistency with canonical roles of the genes in other animals, including free-living flatworms. Recapitulation of developmental factors up-regulated during larval metamorphosis suggests that strobilar growth involves many of the same underlying gene regulatory networks despite the significant disparity in developmental outcomes. The majority of genes identified were investigated in tapeworms for the first time, setting the stage for advancing our understanding of developmental genetics in an important group of flatworm parasites.Electronic supplementary materialThe online version of this article (10.1186/s13227-018-0110-5) contains supplementary material, which is available to authorized users.

The tapeworm interactome: inferring confidence scored protein-protein interactions from the proteome of Hymenolepis microstoma

“…Differentially-expressed genes with a log2 fold-change ≥ 2 between whole, gravid adults and 5-day old larval worms [Olson et al, 2018].…”

“…Differential expression was calculated by re-analysing previously available RNA-seq data [Olson et al, 2018] in order to take advantage of the v.3 genome assembly and gene models. Briefly, raw reads were aligned to the genome using STAR [Dobin et al, 2013] v2.4.2a with the alignIntronMin 10 option, count files were produced using featureCounts v1.6.3 [Liao et al, 2014], and differential expression assessed using DESeq2 [Love et al, 2014] v1.20.0 with a p -value threshold of 0.00001.…”

“…Hymenolepis microstoma , the mouse bile-duct tapeworm, is one of three species of rodent/beetle-hosted hymenolepid tapeworms that have been used widely as laboratory models, as their entire life cycles can be passaged using hosts that are themselves model organisms [Cunningham and Olson, 2010]. A draft genome was published in 2013 [Tsai et al, 2013] and was followed in 2015 by the public release of an up-dated assembly (v.2) based on additional Illumina data, as described by Olson and colleagues [Olson et al, 2018]. This assembly was used to investigate differentially-expressed genes among different life cycle stages and regions of the adult, strobilar worm [Olson et al, 2018], and for characterisation of the microRNA complement [Macchiaroli et al, 2019].…”

“…A draft genome was published in 2013 [Tsai et al, 2013] and was followed in 2015 by the public release of an up-dated assembly (v.2) based on additional Illumina data, as described by Olson and colleagues [Olson et al, 2018]. This assembly was used to investigate differentially-expressed genes among different life cycle stages and regions of the adult, strobilar worm [Olson et al, 2018], and for characterisation of the microRNA complement [Macchiaroli et al, 2019]. In 2018 long-read sequence and optical mapping data were added and all available genome data re-assembled, resulting in a complete assembly consisting of six scaffolds that correspond to their six haploid chromosomes [Hossain and Jones, 1963]; any missing data that remain are likely to represent collapsed repeats rather than unique, non-repetitive sequence (Olson, in preparation).…”

Region-specific regulation of stem cell-driven regeneration in tapeworms

“…WISH and FISH protocols were modified from previously published methods for planarians 52 and the mouse bile-duct tapeworm Hymenolepis microstoma 54 . Tapeworms were heat killed and fixed in 4% formaldehyde/10% DMSO/1% NP40/PBSTx for 30 min at room temperature before washing and dehydration into methanol.…”