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Parasit Vectors. 2018 Mar 6;11(1):152. doi: 10.1186/s13071-018-2745-4.

A recombinant Fasciola gigantica 14-3-3 epsilon protein (rFg14-3-3e) modulates various functions of goat peripheral blood mononuclear cells.

Author information

1
State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu Province, 730046, People's Republic of China.
2
College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.
3
European Molecular Biology Laboratory-European Bioinformatics Institute, Wellcome Genome Campus, Hinxton, CB10 1SD, UK.
4
Faculty of Medicine and Health Sciences, School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington Campus, Loughborough, LE12 5RD, UK.
5
Jiangsu Co-innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University College of Veterinary Medicine, Yangzhou, Jiangsu Province, 225009, People's Republic of China.
6
Faculty of Medicine and Health Sciences, School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington Campus, Loughborough, LE12 5RD, UK. hany.elsheikha@nottingham.ac.uk.
7
State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu Province, 730046, People's Republic of China. xingquanzhu1@hotmail.com.

Abstract

BACKGROUND:

The molecular structure of Fasciola gigantica 14-3-3 protein has been characterized. However, the involvement of this protein in parasite pathogenesis remains elusive and its effect on the functions of innate immune cells is unknown. We report on the cloning and expression of a recombinant F. gigantica 14-3-3 epsilon protein (rFg14-3-3e), and testing its effects on specific functions of goat peripheral blood mononuclear cells (PBMCs).

METHODS:

rFg14-3-3e protein was expressed in Pichia pastoris. Western blot and immunofluorescence assay (IFA) were used to examine the reactivity of rFg14-3-3e protein to anti-F. gigantica and anti-rFg14-3-3e antibodies, respectively. Various assays were used to investigate the stimulatory effects of the purified rFg14-3-3e protein on specific functions of goat PBMCs, including cytokine secretion, proliferation, migration, nitric oxide (NO) production, phagocytosis, and apoptotic capabilities. Potential protein interactors of rFg14-3-3e were identified by querying the databases Intact, String, BioPlex and BioGrid. A Total Energy analysis of each of the identified interaction was performed. Gene Ontology (GO) enrichment analysis was conducted using Funcassociate 3.0.

RESULTS:

Sequence analysis revealed that rFg14-3-3e protein had 100% identity to 14-3-3 protein from Fasciola hepatica. Western blot analysis showed that rFg14-3-3e protein is recognized by sera from goats experimentally infected with F. gigantica and immunofluorescence staining using rat anti-rFg14-3-3e antibodies demonstrated the specific binding of rFg14-3-3e protein to the surface of goat PBMCs. rFg14-3-3e protein stimulated goat PBMCs to produce interleukin-10 (IL-10) and transforming growth factor beta (TGF-β), corresponding with low levels of IL-4 and interferon gamma (IFN-γ). Also, this recombinant protein promoted the release of NO and cell apoptosis, and inhibited the proliferation and migration of goat PBMCs and suppressed monocyte phagocytosis. Homology modelling revealed 65% identity between rFg14-3-3e and human 14-3-3 protein YWHAE. GO enrichment analysis of the interacting proteins identified terms related to apoptosis, protein binding, locomotion, hippo signalling and leukocyte and lymphocyte differentiation, supporting the experimental findings.

CONCLUSIONS:

Our data suggest that rFg14-3-3e protein can influence various cellular and immunological functions of goat PBMCs in vitro and may be involved in mediating F. gigantica pathogenesis. Because of its involvement in F. gigantica recognition by innate immune cells, rFg14-3-3e protein may have applications for development of diagnostics and therapeutic interventions.

KEYWORDS:

14-3-3 epsilon protein isoform; Fasciola gigantica; Goat; Homology modelling; Peripheral blood mononuclear cells

PMID:
29510740
PMCID:
PMC5840819
DOI:
10.1186/s13071-018-2745-4
[Indexed for MEDLINE]
Free PMC Article

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