Identification of a novel methyltransferase-type 12 protein from Haemonchus contortus and its effects on functions of goat PBMCs

Abstract

Background: Methyltransferases (MTFs) are broad range of enzymes, which are ubiquitously expressed in diverse organisms ranging from bacteria to animals. MTFs proteins have been associated with various biological/cellular processes including transcriptional regulation, subcellular protein and RNA localization, signal transduction and DNA-damage repair. However, the role of MTFs in immune mechanism during host-parasite interaction has not been addressed yet. Results: An open reading frame (764 bp) of methyltransferase-type 12 gene of H. contortus denoted as HcMTF-12, was successfully cloned using reverse transcriptase-polymerase chain reaction (RT-PCR) followed by prokaryotic expression in Escherichia coli BL21 (DE3 strain). The recombinant HcMTF-12 protein (rHcMTF-12) was about 47 kDa along with a fusion vector protein of 18 kDa. Immunoblot results identified the native protein MTF-12 with antibodies produced in rats against rHcMT-12, whereas rHcMTF-12 protein was recognized with sera of goat experimentally infected with H. contortus. Immunohistochemical analysis revealed that the native MTF-12 protein was mainly located in the periphery (cuticle) of parasite sections as well as within the pharynx and intestinal region. An immunofluorescence assay validated that rHcMTF-12 attached to the surface of goat PBMCs. Furthermore, the cytokines transcription of IL-2, IFN-γ and IL-4 transcripts of PBMCs incubated with rHcMTF-12 were enhanced in a dose-dependent manner. The secretion of TGF-β1 and IL-10 was significantly decreased. However, IL-6 production was not significantly different as compared to the control groups. Moreover, the migration activity and nitric oxide (NO) production by PBMCs were induced considerably, whereas the proliferation of PBMCs cells was negatively affected when incubated with the rHcMTF-12 protein. Conclusions: Our findings suggest that HcMTF-12 significantly mediated the functions of PBMCs, and it might be a potential candidate for therapeutic interventions against haemonchosis.

The GT1-TPS Structural Domain Protein From Haemonchus contortus Could Be Suppressive Antigen of Goat PBMCs

“…The binding of recombinant proteins to specific receptors of PBMCs is an important way for them to exert immuno-modulation ( 32 , 33 ). Previous studies in our laboratory have shown that His-tag could not affect the binding of recombinant proteins to PBMCs ( 31 , 34 , 35 ). The binding of rHc-TPS to goat PBMCs was confirmed by IFA.…”

“…Group settings were consistent with the description of nitric oxide production assay. Total RNA extraction and cDNA preparation were performed as previously reported ( 31 ). The primers INF-γ, IL-2, IL-4, IL-9, IL-10, IL-17, transforming growth factor β (TGF-β), IL-2R, inducible nitric oxide synthase (iNOS), and signal transducer and activator of transcription 5 (STAT5) for qPCR are listed in Supplementary Table 2 .…”

In vitro characterization of Haemonchus contortus trehalose-6-phosphate phosphatase and its immunomodulatory effects on peripheral blood mononuclear cells (PBMCs)

“…Previous studies have shown that various HcESP-derived protein molecules, such as rHcMTF-12 [ 54 ], rHCRD [ 37 ], rHc-GDC [ 40 ], rHcEF-1α [ 46 ], rHcES-15 [ 34 ], rMiro-1 [ 55 ], HCcyst-3 [ 56 ] and RHC-AK [ 57 ], can bind to PBMCs in vitro and regulate cellular immune function. Our studies showed that rHcGOB can bind to goat PBMCs and activate the IL-10/TGF-β/STAT3 pathway in PBMCs in vitro.…”

Advances in the Development of Anti-Haemonchus contortus Vaccines: Challenges, Opportunities, and Perspectives

“…Apart from the discrete functional potency of different constituents of HcESPs on goat PBMCs, a recent research on tropomyocin protein showed the suppressive potential of rHc-TpMy on goat PBMCs by suppressing various immune functions of host PBMCs [ 93 ]. Recently, we have explored a novel immune modulator from HcESPs, which improved the IFN-γ, IL-4, and IL-2 cytokines production of host immune cells [ 94 ]. In addition to that, ES proteins were found to have distinct immunomodulatory and suppressive regulatory properties during host–parasite interactions, which suggested that the discoveries of these novel antigens might be helpful in anti-parasitic vaccine development in the near future.…”