Calmodulin disruption impacts growth and motility in juvenile liver fluke. Issue 1 (December 2016)

• Record Type: Journal Article
• Authors: McCammick, Erin; McVeigh, Paul; McCusker, Paul; Timson, David; Morphew, Russell; Brophy, Peter; Marks, Nikki; Mousley, Angela; Maule, Aaron
• Is Part Of: Parasites & vectors. Volume 9:Issue 1(2016)
• Page Start: 1
• Page End: 13
• Publication Date: 2016-12
• Subjects: Fasciola hepatica -- Calmodulin -- Flukicide target -- RNAi -- Growth phenotype -- Motility phenotype; Parasitism -- Periodicals; Parasites -- Periodicals; Vector-pathogen relationships -- Periodicals; Animals as carriers of disease -- Periodicals; Insects as carriers of disease -- Periodicals; 616.96
• DOI: 10.1186/s13071-016-1324-9
• Languages: English
• ISSNs: 1756-3305

Abstract

Abstract Background Deficiencies in effective flukicide options and growing issues with drug resistance make current strategies for liver fluke control unsustainable, thereby promoting the need to identify and validate new control targets inFasciola spp. parasites. Calmodulins (CaMs) are small calcium-sensing proteins with ubiquitous expression in all eukaryotic organisms and generally use fluctuations in intracellular calcium levels to modulate cell signalling events. CaMs are essential for fundamental processes including the phosphorylation of protein kinases, gene transcription, calcium transport and smooth muscle contraction. In the blood flukeSchistosoma mansoni, calmodulins have been implicated in egg hatching, miracidial transformation and larval development. Previously, CaMs have been identified amongst liver fluke excretory-secretory products and three CaM-like proteins have been characterised biochemically from adultFasciola hepatica, although their functions remain unknown. Methods In this study, we set out to investigate the biological function and control target potential ofF. hepatica CaMs (Fh CaMs) using RNAi methodology alongside novel in vitro bioassays. Results Our results reveal that: (i)Fh CaMs are widely expressed in parenchymal cells throughout the forebody region of juvenile fluke; (ii) significant transcriptional knockdown ofFhCaM1-3 was inducible by exposure to either long (~200 nt) double stranded (ds) RNAs or 27 nt short interfering (si) RNAs, although siRNAs were less effective than long dsRNAs; (iii) transient long dsRNA exposure-induced RNA interference (RNAi) ofFhCaM s triggered transcript knockdown that persisted for ≥ 21 days, and led to detectable suppression ofFh CaM proteins; (iv)FhCaM RNAi significantly reduced the growth of juvenile flukes maintained in vitro; (v)FhCaM RNAi juveniles also displayed hyperactivity encompassing significantly increased migration; (vi) both the reduced growth and increased motility phenotypes were recapitulated in juvenile fluke using the CaM inhibitor trifluoperazine hydrochloride, supporting phenotype specificity. Conclusions These data indicate that the Ca2+ -modulating functions ofFh CaMs are important for juvenile fluke growth and movement and provide the first functional genomics-based example of a growth-defect resulting from gene silencing in liver fluke. Whilst the phenotypic impacts ofFh CaM silencing on fluke behaviour do not strongly support their candidature as new flukicide targets, the growth impacts encourage further consideration, especially in light of the speed of juvenile fluke growth in vivo.