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- Title
Trichinella spiralis: Knockdown of gamma interferon inducible lysosomal thiol reductase (GILT) results in the reduction of worm burden.
- Authors
Fei, Hong; Naqvi, Muhammad Ali-ul-Husnain; Naqvi, Sana Zahra; Xu, Lixin; Song, Xiaokai; Li, Xiangrui; Yan, Ruofeng
- Abstract
Trichinella spiralis is mammalian skeletal muscles parasite which may cause trichinellosis in animals and humans. Gamma interferon inducible lysosomal thiol reductase (GILT) is a widespread superfamily which plays key role in processing and presentation of MHC class II restricted antigen by catalyzing disulfide bond reduction. There are no reports about GILT in T. spiralis. In present study, GILT from T. spiralis (Tsp-GILT) was cloned, analyzed by multiple-sequence alignment, and predicted by 3D structure model. Recombinant Tsp-GILT (about 46 kDa) was efficiently expressed in Escherichia coli and thiol reductase activity suggested that in acidic environment the addition of a reducing agent is needed. Soaking method was used to knockdown expression of Tsp-GILT using small interference RNA (siRNA). Immunofluorescence assay confirmed the transformation of siRNA into muscle larva (ML) and new born larva (NBL). Quantitative real time-PCR (QRT-PCR) analysis revealed that transcription level of Tsp-GILT mRNA can be up-regulated by stimulation of mouse IFN-γ and down-regulated by siRNA2 in vitro. NBLs soaked with siRNA2 showed 32.3% reduction in the generation of MLs. MLs soaked with siRNA2 showed 26.2% reduction in the next generation of MLs, but no significant effect was observed on adult worms or NBLs. These findings concluded that GILT may play important roles in the development of T. spiralis parasite. Author summary: Trichinella spiralis is the most important pathogen causing trichinellosis of animals and humans. Reports in mammal animals showed that gamma interferon inducible lysosomal thiol reductase (GILT) play key roles in processing and presentation of MHC class II restricted antigens. However, there were no reports about GILT in nematode T. spiralis. In this study, GILT gene from T. spiralis (Tsp-GILT) was cloned and analyzed. The predicted 3D structure of Ts-GILT protein was found similar with that of Homo sapiens and Sus scrofa. To further evaluate the roles of Tsp-GILT in T. spiralis development, a soaking method was used to knockdown the expression of Tsp-GILT gene by siRNA. It was found that NBLs soaked with siRNA showed 32.3% reduction in the generation of MLs, and MLs soaked with siRNA showed 26.2% reduction in the next generation of MLs. Current study concluded that siRNA knockdown of gamma interferon inducible lysosomal thiol reductase from Trichinella spiralis significantly reduce the larval infectivity, development and survival.
- Subjects
TRICHINELLA spiralis; INTERFERON gamma; WILD boar; NON-coding RNA; PROTEIN structure; NEMATODE infections; GLYCOGEN storage disease type II; TRYPANOSOMA cruzi
- Publication
PLoS Neglected Tropical Diseases, 2021, Vol 15, Issue 11, p1
- ISSN
1935-2727
- Publication type
Article
- DOI
10.1371/journal.pntd.0009958