Structural Study Of Chicken CD8αα/pMHCI Complex And Analysis Of Specific α/βTCR Polymorphism

Cytotoxic T lymphocytes (CTL) immune response mediated by chicken CD8+T cell plays a crucial role in the process of eliminating virus infection. During the process, the T cell receptor (TCR) binds the MHCI molecule firstly, and then the TCR-pMHCI complex binds co-receptor CD8to activating the T cell. However, till now, researches of chicken CTL immunology are rarely reported, especially the structural studies of pMHCI and pMHCI-CD8. Compare with the mammals, chicken T cell research is more valuable because of the T cell containing the fifty percent y8T cell. Therefore, for promoting the research of CTL response relevant structure and disease’s prevention and control, we solved the structures of chicken CD8aa homodimer, CD8aa/pBF2*1501and CD8aa/pBF2*0401complexes.The structural analysis shows that the chicken CD8aa has a classical IgV domain, and the way of CD8aa homodimer’s formation is conserved in the evolution; There is a special species characterstic in the interaction of BF2and CD8aa molecules. In the CD8aa/pBF2*1501and CD8aa/pBF2*0401complexes, CD8al and CD8a2subunits make an equal contribution to the BF2molecules. And we found that the CD loop of CD8aa makes contact with the BF2molecules, and the affinity value between BF2molecules and the CD8αα is highest in all the known CD8aa/pMHCI complexes at the first time. Moreover, the number of the contacts, the residues invovling the interaction and the affinity are different between CD8aa/pBF2*1501and CD8aαα/pBF2*0401. After binding the CD8aa, the a3domain of pMHCI complex shifts towards the CD8aa, but the peptide binding is stable except some side chain shifts slightly. These results will help us understand the interaction mechanism between co-receptor CD8and BF2-peptide complex, and CTL immune responses in chicken.In order to identify and select the peptide-specific CD8+T cell, we screened peptides RY0808(RRREQTDY) and RRE (RREVHTYY) which can bind stably with BF2*1501from MDV and AIV. Firstly, we made the B15-MDV-tetramer and B15-AIV-tetramer and prepared the Mouse anti chicken CD8a antibodies of chicken. And then, CTL epitope peptides RY0808(RRREQTDY) and RRE (RREVHTYY) were used to immunize SPF chickens. Finally, we used the antibodies and tetramer combining the flow cytometry technology to value their immunological activity in vivo, and we proved their function for activating the T cell efficiently. In addition, we cloned and analyzed the TCR genes and got the stable αβTCR heterodimer in vitro, all these will be the basis of the TCR-related structural and functional research in the chicken CTL response.In summary, these research results will give insight into the immunological synapse interaction between the MHCI and CDS at atom level. It will help us understand the mechanism of CTL responses to virus infection.