Genome-Wide Tissue-Specific Gene Expression, Co-expression and Regulation of Co-expressed Genes in Adult Nematode Ascaris suum

Abstract

Background Caenorhabditis elegans has traditionally been used as a model for studying nematode biology, but its small size limits the ability for researchers to perform some experiments such as high-throughput tissue-specific gene expression studies. However, the dissection of individual tissues is possible in the parasitic nematode Ascaris suum due to its relatively large size. Here, we take advantage of the recent genome sequencing of Ascaris suum and the ability to physically dissect its separate tissues to produce a wide-scale tissue-specific nematode RNA-seq datasets, including data on three non-reproductive tissues (head, pharynx, and intestine) in both male and female worms, as well as four reproductive tissues (testis, seminal vesicle, ovary, and uterus). We obtained fundamental information about the biology of diverse cell types and potential interactions among tissues within this multicellular organism.Methodology/Principal FindingsOverexpression and functional enrichment analyses identified many putative biological functions enriched in each tissue studied, including functions which have not been previously studied in detail in nematodes. Putative tissue-specific transcriptional factors and corresponding binding motifs that regulate expression in each tissue were identified, including the intestine-enriched ELT-2 motif/transcription factor previously described in nematode intestines. Constitutively expressed and novel genes were also characterized, with the largest number of novel genes found to be overexpressed in the testis. Finally, a putative acetylcholine-mediated transcriptional network connecting biological activity in the head to the male reproductive system is described using co-expression networks, along with a similar ecdysone-mediated system in the female.Conclusions/SignificanceThe expression profiles, co-expression networks and co-expression regulation of the 10 tissues studied and the tissue-specific analysis presented here are a valuable resource for studying tissue-specific biological functions in nematodes.

Functional and Phylogenetic Characterization of Proteins Detected in Various Nematode Intestinal Compartments*

“…The genes encoding all of the detected proteins were previously determined to be expressed in the A. suum intestine by RNA-Seq analysis (15) except for three (GS_14239 and GS_21581 in the PF, and GS_14331 in the PBS perfusate; The expression of these three genes was detected in in the head, pharynx, and/or testes). Proteins were assigned to intestinal compartments according to the method of collection and computational annotation of results ( Fig.…”

“…Functional Annotation-Interproscan (23-25) was used to determine associations of genes in the current A. suum genome (12) to Gene Ontology (GO) terms (26) and Interpro domains (27), as previously described (15). In addition, predicted proteins were searched against the KEGG database (28) using KAAS (29).…”

“…Transcription Factor Binding Site Motif Enrichment-Transcription factor (TF) consensus binding site (motif) enrichment and annotation was performed similarly to our previous study on tissue-specific overexpression in A. suum (15), but with a much more expansive reference database containing rich annotation for C. elegans (33). A maximum length of 2000bp 5Ј upstream regions (URs) were extracted for each gene based on the A. suum genome annotation, with shorter lengths where URs reached the end of a scaffold or overlapped with other coding sequences on either strand (12).…”

“…In addition, known C. elegans motifs from JASPAR (36) and UniProbe (37) were included in the search, as well as vertebrate and mouse (respectively) sequences from the same databases. Vertebrate sequences were included because (as the modENCODE project demonstrated (33)) orthologous transcription factors from vertebrates to nematodes maintain conserved sequence preferences, so these annotations were used when no significant C. elegans transcription factor matches were annotated (as previously performed for A. suum tissue-specific gene expression TF annotation, before the modENCODE database was available (15)). Fig.…”

“…However, the parasitic roundworm Ascaris suum (size 15-40 cm; which also has a complete draft genome available (12,13)) is large enough that the intestine can be physically manipulated for this purpose. The large size of the A. suum intestine has also supported the broad molecular characterization of intestinal cell functions based on gene expression studies using expressed sequence tags (3), microarray (14) and RNA-seq approaches (15). These studies analyzed the transcript expression level in whole intestinal tissue relative to other tissues but not protein expression or compartmentalization of proteins within intestinal tissue, although a proteomic study of A. suum proteins in nonintestinal fluids including perienteric and uterine fluids was recently reported (16).…”