Molecular Mechanisms Underlying Transcription Factor Foxp1 Mediated Regulation Of Regulatory T Cell Homeostasis And Suppressive Function

Regulatory T cell(Treg cell)is a subpopulation of T cells that regulate immune homeostasis and self-tolerance,which exerts their immunosuppressive functions by suppressing effector T cells,and they are closely related to the development of infection,autoimmune disease and tumor.Generally,the Treg cells we mentioned refer to CD4+Foxp3+Treg cells,in which transcription factor Foxp3 is indispensable for Treg cell development,maintenance and suppressive function.In both human and mice,the dysfunctional mutation or deletion of Foxp3 is sufficient to drive lethal systemic autoimmunityStudies have suggested that Foxp3 alone is insufficient to determine the Treg cell differentiation and function though it has important roles in Treg cells;instead,Treg cell differentiation and functional maintenance need a well-orchestrated transcriptional network,including the interactions between Foxp3 and its partners.A large number of Foxp3 partners have been identified as transcription-related proteins,including transcription factors NFATc2,Runxl,Bcll lb,Foxp1,Foxp4,GATA-3,STAT3,Ikaros(Tkzfl),Aiolos(Ikzf3),Ets and Cnot3.Among these Foxp3 partners,Foxpl is one of the P sub-family members of the forkhead box(FOX)transcription factor family as is Foxp3,they have conserved forkhead domain and can bind to the same DNA motif,moreover,they can form heterodimers.However,the role of Foxpl in Treg cells and the relationship between Foxpl and Foxp3 in Treg cells are not understood yet.Based on the expression levels of cell surface marker CD44 and CD62L,Treg cells can be divided into CD44lowCD62Lhigh resting Treg(rTreg)and CD44highCD62Llow activated Treg(aTreg)cells.Compared with rTreg cells,aTreg cells have more spontaneous proliferation and are generally considered as potent suppressors in the anti-inflammatory responses.Instead,rTreg cells act mainly as a reserve pool that is responsible for continuous Treg cell supply.Therefore,the regulation of rTreg and aTreg cell homeostasis and the maintenance of rTreg cell pool is of great importance as well.Studies have shown that transcription factors including Blimp-1,IRF4,Myb,NF-κB c-Rel,Foxol and Nrpl are important for aTreg differentiation and migration,neverthless,the transcriptional regulation of rTreg cell quiescence maintenance and Treg cell homeostasis are poorly understood.Therefore,the transcriptional regulation of Treg cell activation and differentiation is still of great interest in the study of Treg cells.Based on the scientific questions above,we mainly focused on the following two aspects:1.Foxpl mediated regulation of Treg cell homeostasis and the related molecular mechanisms.Firstly,we dissected the unique expression pattern of Foxpl in rTreg and aTreg cells.We found rTreg cells expressed high levels of Foxpl,in which Foxp1A is the major isoform.However in aTreg cells,though the expression level of Foxp1D is relatively increased,total Foxpl expression is at a low level.Secondly,we found that Foxpl does not affect the generation of Treg cells in the thymus;instead,it affects the differentiation and maintenance of iTreg cells.Then,through dynamically comparing Treg cell activation in mice of different ages,we found Foxpl is essential for maintemaning Treg cell quiescence and inhibiting aTreg cell differentiaton.Moreover,through combing Foxp1f/foxp3Cre/+female mice and Foxp1 acute deletion model,we demonstrated that Foxpl regulates Treg cell quiescence in an intrinsic manner.Mechanistically,by analyzing cell proliferation,apoptosis and RNA sequencing data,we found Foxpl possibly maintains Treg cell quiescence through inhibiting cell proliferation and energy metabolism,which is quite similar to the mechanism involved in CD8+ T cell quiescence regulation.2.Foxpl mediated regulation of Treg cell suppressive function and the related molecular mechanisms.Firstly,we analyzed T cell activation in mice of different ages under hoemostatic conditions,and we found young mice with Foxpl-deficient Treg cells have already exhibited lymphadenopathy,splenomagly and more conventional T cell activation;meanwhile the inflammation in older mice with Foxpl-deficient Treg cells is more severe.Next,we used two different inflammation models to study the function of Foxpl-deficient Treg cells in anti-inflammation response.We found both in dextran sulfate sodium(DSS)induced colitis model and experimental autoimmune encephalomyelitis(EAE)model,the suppressive function of Foxpl-deficient Treg cells were greatly impaired.Mechanistically,we demonstrated that both Foxp3 and Ctla4 are direct target genes of Foxpl,and Foxpl helps maintain Treg suppressive function partially through control stable Foxp3 expression.Moreover,we took Ctla4 as an example and proved the synergistic regulation by Foxpl and Foxp3 in Treg cells.Taken together,our study demonstrates that Foxpl plays critical roles in both maintaining Treg cell quiescence during homeostasis and regulating Treg suppressive function and that the related molecular mechanisms,improving the transcriptional regulatory network in Treg cells.Besides,Foxpl and Foxp3 synergistically regulate key genes in Treg cells,which provide new insights in mechanisms involving transcriptional regulation of Treg cells.