ABSTRACT
Human IL-10 (hIL-10) signaling is mediated by receptors consisting of two subunits, IL-10 receptor 1 (IL-10R1) and IL-10 receptor 2. Two common variants of the IL-10R1 (Ser 138 Gly (single-nucleotide polymorphism 3, SNP3) and Gly 330 Arg (SNP4)) are associated with diverse disease phenotypes. Viral homologs to hIL-10, such as cmvIL-10, utilize the same IL-10 receptor complex as part of viral immune evasion strategies. For the present study we hypothesized that IL-10R1 variants alter the ability of viral IL-10 to utilize the IL-10R1 signaling pathway. HeLa cell clones expressing different IL-10R1 haplotypes (WT or any variant) were incubated with hIL-10 or cmvIL-10. In cells expressing IL-10R1-WT, cmvIL-10 (both non-glycosylated- and HeLa-expressed) resulted in equal or slightly stronger STAT3 phosphorylation compared with hIL-10. In clones expressing IL-10R1-SNP3, IL-10R1-SNP4 or IL-10R1-SNP3+4, the cmvIL-10 showed significantly less STAT3 phosphorylation, especially when HeLa-expressed cytokines were used. Time course experiments demonstrated a slower kinetic of cmvIL-10 STAT3 activation through the variant IL-10R1. Similarly, IL-10R1 variants decreased the cmvIL-10-induced SOCS3 and signaling lymphocytic activation molecule mRNA expression. These data suggest that the IL-10R1 variants differentially reduce the signaling activity of cmvIL-10 and thereby may affect CMV's ability to escape from the host's immune surveillance.
Subject(s)
Interleukin-10 Receptor alpha Subunit/immunology , Viral Proteins/immunology , Haplotypes , HeLa Cells , Humans , Interleukin-10 Receptor alpha Subunit/genetics , Interleukin-10 Receptor alpha Subunit/metabolism , Kinetics , Ligands , Lymphocyte Activation/immunology , Mutation/genetics , Phosphorylation , STAT3 Transcription Factor/metabolism , Suppressor of Cytokine Signaling 3 Protein , Suppressor of Cytokine Signaling Proteins/genetics , Suppressor of Cytokine Signaling Proteins/metabolism , Viral Proteins/genetics , Viral Proteins/metabolismABSTRACT
Botulinum neurotoxin C1 (BoNt C1) and its corresponding gene were detected in seven aquatic habitats covering a range of low (LR) to high risk (HR) avian botulism outbreak areas during a study period of 10 months. Toxin and gene in sediment and avian faecal samples were analysed before (in situ) and after cultivation (in vitro) by a newly adapted ELISA, the common mouse bioassay and by a recently described nested PCR protocol. BoNt C1 gene fragments were detected in 74% and 83% of all investigated sediment samples by in situ PCR and in vitro PCR, respectively, at comparable frequencies in HR and LR areas. Similar high values were also observed for faecal samples. No BoNt C1 could be detected in the sediment in situ, while 53% and 56% of all cultivated samples contained BoNt C1 as detected in the mouse bioassay and the ELISA, respectively. The percentage of BoNt C1 positive cultivated samples was significantly higher (2-fold) in HR areas than in LR areas. Hence, our data clearly indicate an increased ratio of potentially BoNt C1 producing clostridia to BoNt C1 genes as the frequency or likelihood of botulinum epizootics increases in the environment. In addition, the good correlation between the results from the ELISA and the mouse bioassay for all sediment and faecal samples (r=0.90, p<0.001, n=121) indicates a high potential for the ELISA to reduce/replace the mouse bioassay for the detection of BoNt C1 in environmental samples.
