Characterization and expression profiling of buffalo IFN-lambda family.

Interferon lambda (IFN-λ) is an important type III interferon triggered mainly by viral infection. IFN-λ binds to their heterodimeric receptors and signals through JAK-STAT pathways similar to type I IFN. In this study, we deduced the buffalo IFN-λ sequences through the polymerase chain reaction, and then studied IFN-λ's expression patterns in different tissues, and post induction with poly I:C and live MRSA using RT-qPCR. The full-length sequences of buffalo IFN-λ3, IFN-λ receptors, and a transcript variant of IFN-λ4 were determined. IFN-λ1 is identified as a pseudogene. Virus response elements and a recombination hotspot factor was observed in the regulatory region of IFN-λ. The IFN-λ3 expressed highest in lungs and monocytes but IFN-λ4 did not. The expression of Interferon Lambda Receptor 1 was tissue specific, while Interleukin 10 Receptor subunit beta was ubiquitous. Following poly I:C induction, IFN-λ3 expression was primarily observed in epithelial cells as opposed to fibroblasts, displaying cell type-dependent expression. The cytosolic RNA sensors were expressed highest in endometrial epithelial cells, whereas the endosomal receptor was higher in fibroblasts. 2',5'-oligoadenylate synthetase expressed higher in fibroblasts, myxoma resistance protein 1 and IFN-stimulated gene 56 in epithelial cells, displaying cell-specific antiviral response of the interferon stimulated genes (ISGs). The endometrial epithelial cells expressed IFN-λ3 after live S. aureus infection indicating its importance in bacterial infection. The induction of IFN-λ3 was S. aureus isolate specific at the same multiplicity of infection (MOI). This study elucidates the IFN-λ sequences, diverse expression patterns revealing tissue specificity, and specificity in response to poly I:C and bacterial stimuli, emphasising its crucial role in innate immune response modulation.

Poly I:C stimulation in-vitro as a marker for an antiviral response in different cell types generated from Buffalo (Bubalus bubalis).

The innate immune system is activated upon virus invasion of a host cell by recognizing viral component, such as dsRNA through specific receptors, resulting in the production of type- I IFNs, which confer an antiviral state within the invaded as well as surrounding cells. In the present study, fibroblast, monocyte and macrophage cells derived from water Buffalo (Bubalus bubalis) were exposed to a synthetic dsRNA analogue, poly I:C to mimic viral invasion in each cell type. Recognition of poly I:C through cytosolic helicase receptors RIG-I and MDA5 molecule lead to the activation of the RLR pathway, subsequently activating the MAVS-IRF3/7 cascade and the production of antiviral effector molecule like IFNß and ISGs. Within the different cell types, we identified variability in RLR receptor and IFNß expression after poly I:C administration. Fibroblasts responded quickly and strongly with IFNß production, followed by macrophages and monocytes. Despite absolute expression variability among different cell types the expression trend of RLRs pathway genes were similar. Length of poly I:C molecule also influence IFNß expression in response of RLR pathway. Short (LMW) poly I:C induce stronger IFN-ß expression in myeloid (macrophage and monocyte) cells. In contrast long (HMW) poly I:C preferably elicit higher IFNß expression in non-myeloid (fibroblast) cell. Therefore, MDA5 and RIG-1 plays an indispensable role in eliciting antiviral response in non- immune (fibroblast) host cell. Thus, stimulation of RLR pathway with suitable and potentially cell-type specific agonist molecules successfully elicit antiviral state in the host animal, with fibroblasts conferring a stronger antiviral state compared with the monocytes and macrophages.