SOCS3 control the activity of NF-κB induced by HSP70 via degradation of MyD88-adapter-like protein (Mal) in IPEC-J2 cells.

Hyperthermia in pigs induces suppressor of cytokine signaling (SOCS) 3 and SOCS4 expression in intestinal gut and causes disruption of inflammation cytokine production. These changes may affect the development of inflammatory bowel disease in heat-stressed pigs. However, the mechanisms are not well understood. Accordingly, in this study, we examined the roles of SOCS members in regulation of the nuclear factor (NF)-κB pathway and heat shock protein (HSP) 70-mediated cytokine induction in 293T human embryonic kidney cells and IPEC-J2 porcine small intestinal epithelial cells. Ectopic expression of HSP70 significantly modulated NF-κB activity (p ≤ .05). Moreover, co-expression of SOCS3 or SOCS4 with HSP70 reduced NF-κB activity, which was abolished by SOCS3 or SOCS4 knockdown with short hairpin RNA. Interestingly, MyD88-adaptor-like (Mal) protein was downregulated in cells expressing SOCS3 but not in cells expressing SOCS4. In addition, SOCS3 but not SOCS4 negatively regulated the activity of NF-κB induced by HSP70 overexpression via degradation of Mal. These findings may facilitate the development of novel SOCS3-based therapeutic strategies to control heat stress-related disorders in pigs.

Molecular cloning and functional studies on magang goose toll-like receptor 5.

Disease outbreaks heavily impact the economic viability of animal industries. Little is known about the mechanisms of immune system-related diseases in geese. Toll-like receptors (TLRs) play a major role in the anti-inflammatory immunity process in most animal species, but they have not been studied in the Magang goose. To elucidate the role of TLRs, reverse transcription polymerase chain reaction (RT-PCR) and PCR amplification of cDNA ends (Smart RACE) were used to clone the Magang goose TLR5 gene (mgTLR5). The full-length cDNA of mgTLR5 was 2967 bp in length, including a 5'-terminal untranslated region (UTR) of 215 bp, a 3'-terminal UTR of 384 bp, and an open reading frame of 2583 bp that encodes a protein of 860 amino acids. Structurally, mgTLR5 has a toll/interleukin-receptor (TIR) domain, a transmembrane domain, and seven leucine-rich repeats (LRRs) domains. Homology alignment of TLR5 and its TIR domains with other species revealed that mgTLR5 shared 98 % and 81.3 % of sequence similarity with white goose TLR5 and chicken TLR5, respectively. Quantitative RT-PCR showed that the mgTLR5 gene of the goose is widely expressed in all tested tissues, with the highest expression in the kidney and spleen. The increase in NF-κB promoter activity stimulated by flagellin was dependent on mgTLR5 expression in 293 T cells. Salmonella pullorum and flagellin significantly upregulated the expression of TLR5, IL-8, and IL-1 mRNA in peripheral blood mononucleotide cells of Magang goose cultured in vitro. Stimulation by S. pullorum for 24 h upregulated mgTLR5 expression in the cecum and kidney. We conclude that Magang goose TLR5 is a functional TLR5 homologue of the protein in other species and plays an important role in bacterial recognition.

Goose toll-like receptor 3 (TLR3) mediated IFN-γ and IL-6 in anti-H5N1 avian influenza virus response.

Induction of the innate immune pathways is critical for early anti-viral defense. How geese recognize viral molecules and activate these pathways is not well understood. In mammals, Toll-like receptor 3 (TLR3) recognizes double-stranded RNA. Activation of TLR3 induces the activation of NF-кB and the production of type-I interferon. In this study, the goose TLR3 gene was cloned using rapid amplification of cDNA ends. Goose TLR3 encoded an 896-amino-acid protein, containing a signal secretion peptide, 14 extracellular leucine-rich repeat domains, a transmembrane domain, a Toll/interleukin-1 receptor signaling domain, and shared 46.7-84.4% homology with other species. Tissue expression of goose TLR3 varied markedly and was highest in the pancreas and lowest in the skin. Human embryonic kidney 293 cells transfected with goose TLR3 and NF-κB-luciferase-containing plasmids responded significantly to poly i:c. The expression of TLR3, IL-6 and IFN-γ mRNA, but not IL-1 mRNA, was significantly upregulated after poly i:c or high pathogenic avian influenza virus (H5N1) stimulation in goose peripheral blood mononuclear cells cultured in vitro. Furthermore, geese infected with H5N1 showed significant upregulation of TLR3, especially in the lung and brain. We conclude that goose TLR3 is a functional TLR3 homologue of the protein in other species and plays an important role in virus recognition.