Hemocyte siRNA uptake is increased by 5′ cholesterol-TEG addition in Biomphalaria glabrata , snail vector of schistosome

Type Article
Date 2021-02
Language English
Author(s) Portet Anaïs1, 2, Galinier Richard1, Lassalle Damien1, Faille Alexandre3, 4, 5, Gourbal Benjamin1, Duval David1
Affiliation(s) 1 : IHPE UMR 5244, CNRS, IFREMER, University of Montpellier, University of Perpignan, Perpignan, France
2 : Department of Medicine, Molecular Immunity Unit, University of Cambridge, Cambridge, United Kingdom
3 : Department of Haematology, University of Cambridge, Cambridge, United Kingdom
4 : Wellcome Trust-Medical Research Council Stem Cell Institute, University of Cambridge, Cambridge, United Kingdom
5 : Cambridge Institute for Medical Research, Cambridge, United Kingdom
Source Peerj (2167-8359) (PeerJ), 2021-02 , Vol. 9 , P. e10895 (19p.)
DOI 10.7717/peerj.10895
Keyword(s) Biomphalaria glabrata, Cholesteryl TEG, RNAi interference, BgTEP1, Hemocyte
Abstract

Biomphalaria glabrata is one of the snail intermediate hosts of Schistosoma mansoni, the causative agent of intestinal schistosomiasis disease. Numerous molecular studies using comparative approaches between susceptible and resistant snails to S. mansoni infection have helped identify numerous snail key candidates supporting such susceptible/resistant status. The functional approach using RNA interference (RNAi) remains crucial to validate the function of such candidates. CRISPR-Cas systems are still under development in many laboratories, and RNA interference remains the best tool to study B. glabrata snail genetics. Herein, we describe the use of modified small interfering RNA (siRNA) molecules to enhance cell delivery, especially into hemocytes, the snail immune cells. Modification of siRNA with 5′ Cholesteryl TriEthylene Glycol (Chol-TEG) promotes cellular uptake by hemocytes, nearly eightfold over that of unmodified siRNA. FACS analysis reveals that more than 50% of hemocytes have internalized Chol-TEG siRNA conjugated to Cy3 fluorophores, 2 hours only after in vivo injection into snails. Chol-TEG siRNA targeting BgTEP1 (ThioEster-containing Protein), a parasite binding protein, reduced BgTEP1 transcript expression by 70–80% compared to control. The level of BgTEP1 protein secreted in the hemolymph was also decreased. However, despite the BgTEP1 knock-down at both RNA and protein levels, snail compatibility with its sympatric parasite is not affected suggesting functional redundancy among the BgTEP genes family in snail-schistosoma interaction.

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Publisher's official version 19 3 MB Open access
Intracellular uptake of siRNA in hemocytes After 2 hours Chol-siRNA-cy3 exposure, hemocytes were analyzed by confocal fluorescence microscopy.... 1 MB Open access
Raw data Four tables with the corresponding raw data for Figures 1C, 2, 3 and 4A. 18 KB Open access
Uncropped Figure 4B 280 KB Open access
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How to cite 

Portet Anaïs, Galinier Richard, Lassalle Damien, Faille Alexandre, Gourbal Benjamin, Duval David (2021). Hemocyte siRNA uptake is increased by 5′ cholesterol-TEG addition in Biomphalaria glabrata , snail vector of schistosome. Peerj, 9, e10895 (19p.). Publisher's official version : https://doi.org/10.7717/peerj.10895 , Open Access version : https://archimer.ifremer.fr/doc/00682/79376/