Molecular identification and characterisation of a novel chicken leukocyte immunoglobulin-like receptor A5.

1. Leukocyte immunoglobulin-like receptor A5 (LILRA5) is a key molecule that regulates the immune system. However, the LILRA5 gene has not been characterised in avian species, including chickens. The present study aimed to identify and functionally characterise LILRA5 identified from two genetically disparate chicken lines, viz., Marek's disease (MD)-resistant (R) line 6.3 and MD-susceptible (S) line 7.2. 2. Multiple sequence alignment and phylogenetic analyses confirmed that the identity and similarity homologies of amino acids of LILRA5 in chicken lines 6.3 and 7.2 ranged between 93% and 93.7%, whereas those between chicken and mammals ranged between 20.9% and 43.7% and 21.1% to 43.9%, respectively. The newly cloned LILRA5 from chicken lines 6.3 and 7.2 revealed high conservation and a close relationship with other known mammalian LILRA5 proteins. 3. The results indicated that LILRA5 from chicken lines 6.3 and 7.2 was associated with phosphorylation of Src kinases and protein tyrosine phosphatase non-receptor type 11 (SHP2), which play a regulatory role in immune functions. Moreover, the results demonstrated that LILRA5 in these lines was associated with the activation of major histocompatibility complex (MHC) class I and ß2-microglobulin and induced the expression of the transporter associated with antigen processing. In addition, LILRA5 in both chicken lines activated and induced Janus kinase (JAK)-signal transducer and the activator of transcription (STAT), nuclear factor kappa B (NF-κB), phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT) and the extracellular signal-regulated kinase (ERK)1/2 signalling pathways; toll-like receptors; and Th1-, Th2-, and Th17- cytokines. 4. The data suggested that LILRA5 has innate immune receptors essential for macrophage immune response and provide novel insights into the regulation of immunity and immunopathology.

Characterization and functional analyses of a novel chicken CD8α variant X1 (CD8α1).

We provide the first description of cloning and of structural and functional analysis of a novel variant in the chicken cluster of differentiation 8 alpha (CD8a) family, termed the CD8α X1 (CD8α1) gene. Multiple alignments of CD8α1 with known CD8α and CD8ß sequences of other species revealed relatively low conservation of AA residues involved in the specific and unique structural domains among CD8α genes. For example, cysteine residues that are involved in disulfide bonding to form the V domain are conserved. In contrast, the O-linked glycosylation sites (XPXX motif) are not found in the chicken CD8α1 sequence, and the A ß strand and complementarity-determining region 1 and 2 sequences are poorly conserved between chicken CD8α1 and avian CD8α. Furthermore, the alignment showed that the transmembrane regions show relatively high sequence similarity, whereas the cytoplasmic regions show relatively low similarity, indicating poor conservation. Moreover, the motif (CXCP) that is thought to be responsible for binding the p56 lymphocyte cell kinase subunit (p56) is missing in the CD8α1 sequence. The chicken CD8α1 genomic structure is similar to that of chicken CD8α, but their protein structures differ. Phylogenetic analysis showed that chicken CD8α1 grouped with known avian CD8α sequences but was somewhat distantly related to the CD8α molecules of other species. Moreover, we analyzed the signal transduction and cytokine response to CD8α1 treatment to determine the specific biological functions of chicken CD8α1 in immune cells. The results showed that chicken CD8α1 is a key regulator of the expression of genes that are associated and cooperate with transcription factors in the major histocompatibility complex class I and II promoter regions and activates Janus kinase (JAK) 1/2, signal transducer and activator of transcription (STAT), and suppressor of cytokine signaling (SOCS) 1 signaling-related genes. Immune cells that express functional CD8α1 induce proinflammatory cytokines as well as innate immune responses. Therefore, our data indicate that CD8α1 may have immunoregulatory activity by regulating the expression of proinflammatory or anti-inflammatory cytokines via its effect on immune cells.