Characterization and differential expression analysis of Toxocara canis aquaporin-1 gene

Abstract

Toxocara canis is an intestinal nematode of canids with a worldwide distribution, causing an important but neglected parasitic zoonosis in humans. Aquaporins (AQP) are a family of water channel proteins, which function as membrane channels to regulate water homeostasis. In this study, the coding sequence of aquaporin-1 gene of T. canis (Tc-aqp-1) was cloned and characterized. The obtained Tc-aqp-1 coding sequence was 933 bp in length, which predicted to encode 311 amino acids. Two conserved asparagine-proline-alanine (NPA) motifs were identified in the multiple sequence alignments. Phylogenetic analysis revealed the closest relationship between T. canis and Opisthorchis viverrini based on aquaporin-1 amino acid sequence. A structure was predicted with ligand binding sites predicted at H93, N95, N226, L94, I79, and I210 and with active sites predicted at I256 and G207. Gene Ontology (GO) annotations predicted its cellular component term of integral component of plasma membrane (GO: 0005887), molecular function term of channel activity (GO: 0015250), and biological process term of water transport (GO: 0006833). Tissue expression analysis revealed that the Tc-aqp-1 was highly expressed in the intestine of adult male. The findings of the present study provide the basis for further functional studies of T. canis aquaporin-1.

Aquaporin 1 is located on the intestinal basolateral membrane in Toxocara canis and might play a role in drug uptake

“…Although nematodes do not have tegument, which have evolved to possess the specialised coat (cuticle), the functional roles of intestinal AQPs in nematodes should be similar to the tegument AQPs in trematodes as both of them are important organs known for nutrient absorption. This proposal can be somewhat supported by the predominant gene transcription of Tc-aqp-1 and protein expression of Tc AQP1 in the intestine of adult T. canis [30]. Interestingly, Tc AQP1 was localised specifically on the intestinal basolateral membrane, which is consistent with the tissue expression of AQP1 in C. elegans [4].…”

“…Recent genomic and transcriptomic studies on this parasite have indicated that Tc-aqp-1 might play important roles in the host-parasite interactions [44–46]. In our recent work [30], we have revealed the transcription profile of Tc-aqp-1 in the different tissues of adult T. canis . As a logical extension, in this study, we explore the tissue expression and function of Tc AQP1 protein in the adult (female and male) worms of T. canis , using immunohistochemistry, RNA interference and nematocidal assays.…”

“…Taken together, Tc AQP1 might represent a novel anthelmintic target. First, the high transcriptional level of Tc-aqp-1 in the infective larvae of T. canis suggests its roles in host-parasite interaction [30, 44, 45]. Second, the predominant expression of Tc AQP1 in the intestine of adult T. canis indicates the crucial roles of this protein in parasite survival, which could be easily targeted.…”

“…After soaking for 24 h, the efficacy of gene knockdown was determined by comparing the relative mRNA levels of Tc-aqp- 1 between the worms treated with silencing and non-silencing RNAs. qRT-PCR was performed as described previously [30] to confirm the efficiency of gene knockdown. In brief, total RNA was extracted from an individual worm ( n = 3) and reversely transcribed.…”

“…These findings have been raising interest in the proposition that AQPs might represent novel therapeutic targets for parasitic diseases [24–27]. However, little is known about this area in parasitic nematodes, though selected genes coding for some AQPs have been cloned and analysed [28–30]. A better understanding of this area in parasitic nematodes would contribute to the discovery of novel anthelmintic targets.…”