Functional Study Of Autophagy Related Gene ATGs In B Cell Development And Germinal Center Reaction

Autophagy and autophagy related genes play an important role in the development of immune cells and the regulation of various immune processes.ATG7 and ATG10 are indispensable parts in the formation and elongation of autophagosomes.Basal level of autophagy is required to maintain certain cellular functions.Autophagy enables recycling of macromolecules that serve as a source of building blocks and energy required for cell survival under stress.Many studies have shown that autophagy related genes play important roles out of the autophagy pathway.The robust humoral immune response requires the formation of germinal center(GC)structures.The proliferating B-cells undergo immunoglobin genes somatic hypermutation(SHM)and class switch recombination(CSR)in the GC to produce high affinity antibodies of different isotype classes,along with generation of long-lived antibody secreting cells(ACSs)and memory B cells.Therefore,the roles of autophagy and autophagy genes ATG7 and ATG10 in the B cell development and GC response deserve further study.In this study,first we checked the expression profiles of ATG7 and ATG10 in human and mouse B-cell lineages and found both were highly upregulated in GC B cells.Secondly,to study the role of ATG7 and ATG10 in B cell development,we used mice with ATG7 or ATG10 deletion at the pro-B late stage to study the formation of various B cell subsets at different developmental stages.The results showed that the proportion of mature B cells in bone marrow and peripheral organs decreased significantly after the deletion of ATG7 and ATG10.In addition,it was found that B1 a B cells in the peritoneal cavity were severely decreased by ATG7 or ATG10 deficiency.It is suggested that ATG7 and ATG10 are necessary for the maintenance of normal homeostasis of peripheral B cell subsets and the survival of B1 a B cells.Additionally,it was found that the proliferation and survival of ATG10-deficient B cells were impaired after the ligation of BCR receptor,which suggested that ATG10 was required for the proliferation and survival of B cells in response to BCR signaling.In GC reaction stage,we analyzed the T cell-dependent antibody responses in mice with ATG7 or ATG10 deletion at the GC B stage,and found that the lacking of ATG7 or ATG10 did not affect the formation of GC and Ig somatic hypermutation.In addition,unlike ATG7-deficient mice and wild-type mice treated with autophagy inhibitor 3-MA,CSR and antibody affinity maturation were impaired in TD antigen immunized ATG10-deficient mice.Furthermore,results showed that the key protein IRF4 and PAX5 were down-regulated in ATG10-deficient B cells,thus affecting the CSR process.These studies suggest that ATG10 plays a key role in GC responses and is involved in the production of high affinity antibodies through pathways other than classical autophagy process.In the post-GC stage,it was found that the secondary immune response were impaired and the number of long-lived ACSs were decreased after genetic ablation of ATG7 or ATG10.It is indicated that both ATG7 and ATG10 are involved in the immune memory response by affecting the survival of long-lived ACSs.Additionally,by establishing a mouse model of collagen-induced arthritis(CIA)in ATG7 conditional knockout mice,we found that the deletion of ATG7 at specific stage of B cells under pathological conditions could ameliorate the severity of the CIA by reducing the level of auto-antibodies,suggesting that ATG7 plays an important role in B cell-driven autoimmune diseases.This study is the first to find that ATG10,a key factor in the classical autophagy pathway,plays a citical role in humoral immune response,which highlights its function outside the autophagy process.In addition,new evidences are provided for emphasizing the importance of autophagy and autophagy protein ATG7 in humoral immune response.Moreover,the study provides more theoretical basis for potential clinical therapies targeting autophagy and autophagy genes.In the future,modulating B cell autophagy appears to be a beneficial therapeutic option for autoantibody-related autoimmune disease.