Lymphatic endothelial cells (LECs) induce peripheral tolerance by direct presentation to CD8 T cells (T CD8) IntroductionIt has been well established that intrinsic peripheral tolerance in self-reactive T cells occurs through anergy or deletion. Early work demonstrated that anergy in vitro was because of lack of CD28 costimulation, 1 which also led to deletional tolerance in vivo. 2,3 However, in other models, CD28 costimulation was required for tolerance induction. 4,5 In addition, induction of peripheral deletion and/or anergy in vivo could be reversed by costimulation through CD27, 4-1BB, and OX40. 6,7 While these costimulatory pathways operate at distinct points in the response of T cells to foreign antigens, they all induce IL-2 production, [8][9][10][11] and are associated with up-regulation of antiapoptotic molecules and enhanced survival. 10,[12][13][14] However, the basis for their reversal of tolerance induction has not been established.Inhibitory signals through programmed cell death 1 (PD-1) and B-and T-lymphocyte attenuator (BTLA) receptors, via their ligands programmed cell death-1 ligand 1 (B7-H1; also known as PD-L1) and herpesvirus entry mediator (HVEM), also have been reported to diminish T-cell accumulation and/or acquisition of effector activity in in vitro 15 and in vivo [16][17][18][19][20] models of tolerance. Interfering with these pathways enables self-reactive T cells to accumulate in secondary lymphoid organs and become fully differentiated effectors that cause autoimmunity. [16][17][18][19] Inhibitory signals through lymphocyte activation gene-3 (LAG-3) also diminish T-cell accumulation in peripheral tissue in vivo, 21 but a role for LAG-3 in CD8 T-cell (T CD8 ) tolerance induction in secondary lymphoid organs has not been established. In response to foreign antigens, signaling via these inhibitory pathways is associated with inhibition of IL-2 production [22][23][24] and diminished expression of antiapoptotic molecules. 23 However, it has yet to be clearly established how a lack of costimulation and inhibitory signaling are related to one another during peripheral tolerance induction. Finally, the cells that express the ligands for these inhibitory receptors during peripheral tolerance induction in vivo have yet to be identified.Peripheral tolerance has classically been ascribed to dendritic cells (DCs) that cross-present self-antigen acquired from peripheral tissues. 25 More recently, it has been demonstrated that it can also be mediated via direct presentation by 3 different lymph node (LN) stromal cell (LNSC) populations, including extrathymic Aireexpressing cells, 26 fibroblastic reticular cells (FRCs), 27 and lymphatic endothelial cells (LECs). 28 We previously reported that LECs directly present an epitope derived from tyrosinase, a melanocyte differentiation protein that is recognized by T CD8 recovered from melanoma and vitiligo patients, and induce peripheral tolerance through deletion of tyrosinase-specific T CD8 . 28 Here, we determined the roles of both costimulatory and ...

SUMMARY It is now established that Bcl11b specifies T cell fate. Here we show that in developing T-cells the Bcl11b enhancer repositioned from the lamina to the nuclear interior. Our search for factors that relocalized the Bcl11b enhancer identified a non-coding RNA named ThymoD (Thymocyte Differentiation Factor). ThymoD-deficient mice displayed a block at the onset of T cell development and developed lymphoid malignancies. We found that ThymoD transcription promoted demethylation at CTCF bound sites and activated cohesin-dependent looping to reposition the Bcl11b enhancer from the lamina to the nuclear interior and to juxtapose the Bcl11b enhancer and promoter into a single loop domain. These large-scale changes in nuclear architecture were associated with the deposition of activating epigenetic marks across the loop domain, plausibly facilitating phase separation. These data indicate how during developmental progression and tumor suppression non-coding transcription orchestrates chromatin folding and compartmentalization to direct with high precision enhancer-promoter communication.

BACKGROUND Severe combined immunodeficiency (SCID) is characterized by arrested T-lymphocyte production and by B-lymphocyte dysfunction, which result in life-threatening infections. Early diagnosis of SCID through population-based screening of newborns can aid clinical management and help improve outcomes; it also permits the identification of previously unknown factors that are essential for lymphocyte development in humans. METHODS SCID was detected in a newborn before the onset of infections by means of screening of T-cell–receptor excision circles, a biomarker for thymic output. On confirmation of the condition, the affected infant was treated with allogeneic hematopoietic stem-cell transplantation. Exome sequencing in the patient and parents was followed by functional analysis of a prioritized candidate gene with the use of human hematopoietic stem cells and zebrafish embryos. RESULTS The infant had “leaky” SCID (i.e., a form of SCID in which a minimal degree of immune function is preserved), as well as craniofacial and dermal abnormalities and the absence of a corpus callosum; his immune deficit was fully corrected by hematopoietic stem-cell transplantation. Exome sequencing revealed a heterozygous de novo missense mutation, p.N441K, in BCL11B. The resulting BCL11B protein had dominant negative activity, which abrogated the ability of wild-type BCL11B to bind DNA, thereby arresting development of the T-cell lineage and disrupting hematopoietic stem-cell migration; this revealed a previously unknown function of BCL11B. The patient’s abnormalities, when recapitulated in bcl11ba-deficient zebrafish, were reversed by ectopic expression of functionally intact human BCL11B but not mutant human BCL11B. CONCLUSIONS Newborn screening facilitated the identification and treatment of a previously unknown cause of human SCID. Coupling exome sequencing with an evaluation of candidate genes in human hematopoietic stem cells and in zebrafish revealed that a constitutional BCL11B mutation caused human multisystem anomalies with SCID and also revealed a prethymic role for BCL11B in hematopoietic progenitors. (Funded by the National Institutes of Health and others.)

The c-Myb transcription factor is required for normal adult hematopoiesis. However, the embryonic lethality of Myb-null mutations has been an impediment to identifying roles for c-Myb during lymphocyte development. We have used tissue-specific inactivation of the Myb locus in early progenitor cells to demonstrate that c-Myb is absolutely required for the differentiation of CD19+ B-lineage cells and B cell differentiation is profoundly blocked beyond the pre-pro-B cell stage in Mybf/f Mb1-cre mice. We demonstrate that c-Myb is required for the intrinsic survival of CD19+ pro-B cells as well as the proper expression of the α-chain of the IL-7 receptor (CD127) and Ebf1. However, survival of c-Myb-deficient CD19+ pro-B cells cannot be rescued by transduction with CD127-producing retrovirus, suggesting that c-Myb controls a survival pathway independent of CD127. Furthermore, c-Myb-deficient progenitor cells inefficiently generate CD19+ B-lineage cells during stromal cell culture but this process can be partially rescued with exogenous Ebf1. Thus, c-Myb does not appear to be required for commitment to B cell differentiation but is crucial for B cell differentiation to the CD19+ pro-B cell stage as well as survival of CD19+ pro-B cells. Surprisingly, forced c-Myb expression in lymphoid-primed multipotent progenitors favors differentiation toward the myeloid lineage, suggesting that proper c-Myb expression is crucial for B-lineage development.