Peripheral immune tolerance is generally thought to result from cross-presentation of tissue-derived proteins by quiescent tissue-resident dendritic cells to self-reactive T cells that have escaped thymic negative selection, leading to anergy or deletion. Recently, we and others have implicated the lymph node (LN) stroma in mediating CD8 T cell peripheral tolerance. We demonstrate that LN-resident lymphatic endothelial cells express multiple peripheral tissue antigens (PTAs) independent of the autoimmune regulator (Aire). They directly present an epitope derived from one of these, the melanocyte-specific protein tyrosinase, to tyrosinase-specific CD8 T cells, leading to their deletion. We also show that other LN stromal subpopulations express distinct PTAs by mechanisms that vary in their Aire dependence. These results establish lymphatic endothelial cells, and potentially other LN-resident cells, as systemic mediators of peripheral immune tolerance.

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 ...

SNF5/INI1 is a component of the ATP-dependent chromatin remodeling enzyme family SWI/SNF. Germ line mutations of INI1 have been identified in children with brain and renal rhabdoid tumors, indicating that INI1is a tumor suppressor. Here we report that disruption of Ini1 expression in mice results in early embryonic lethality. Ini1-null embryos die between 3.5 and 5.5 days postcoitum, and Ini1-null blastocysts fail to hatch, form the trophectoderm, or expand the inner cell mass when cultured in vitro. Furthermore, we report that approximately 15% of Ini1-heterozygous mice present with tumors, mostly undifferentiated or poorly differentiated sarcomas. Tumor formation is associated with a loss of heterozygocity at the Ini1 locus, characterizing Ini1 as a tumor suppressor in mice. Thus, Ini1 is essential for embryo viability and for repression of oncogenesis in the adult organism.The compact nature of chromatin structure presents a barrier to cellular processes that require access to DNA. A number of multiprotein complexes have been identified that share the ability to modify chromatin structure. These include the histone acetyltransferases and deacetylases, complexes which chemically modify the amino-terminal tails of histones by the addition or removal of acetyl groups, respectively, as well as a group of enzymes that utilize the energy derived from ATP hydrolysis to alter nucleosome structure (16,20,43,44,50). Included among these ATP-dependent chromatin remodeling enzymes is the SWI/SNF family of chromatin modifiers.SWI/SNF enzymes are large multisubunit enzymes of ϳ1 to 2 MDa. Yeast SWI/SNF genes were originally identified as being required for mating type switching or sucrose fermentation (4,32,42). Later work determined that SWI/SNF genes were required for the induction of a subset of yeast genes and that the SWI2/SNF2 protein possessed a DNA-stimulated ATPase activity (6,22,26,33,34,54). Mutations in SWI/SNF genes could be suppressed by mutations altering histone gene expression, histone structure, or nonhistone chromatin proteins, leading to the suggestion that these gene products facilitated transcriptional activation by altering chromatin structure (15,23,24).Human SWI/SNF (hSWI/SNF) complexes contain either the human BRM (hBRM) (hSNF2␣) or BRG1 (hSNF2␤) homologues of the yeast SWI2/SNF2 ATPase (7,19,30). Both yeast and human SWI/SNF complexes have been shown to possess nucleosome remodeling activity in vitro (8,17,25). Components of mammalian SWI/SNF complexes have been implicated in a variety of cellular processes, including gene activation and repression, development and differentiation, recombination and repair, and cell cycle control. There is evidence supporting a role for SWI/SNF in gene activation events mediated by nuclear hormone receptors, environmental stress, and viral infection (1,7,10,13,30). In contrast, SWI/SNF components also were shown to be involved in repression of c-fos and some E2F-regulated genes (31, 48). Both BRG1 and hBRM can interact with the retinoblastoma oncoprotein and indu...

ATP-dependent chromatin-remodeling complexes are conserved among all eukaryotes and function by altering nucleosome structure to allow cellular regulatory factors access to the DNA. Mammalian SWI-SNF complexes contain either of two highly conserved ATPase subunits: BRG1 or BRM. To identify cellular genes that require mammalian SWI-SNF complexes for the activation of gene expression, we have generated cell lines that inducibly express mutant forms of the BRG1 or BRM ATPases that are unable to bind and hydrolyze ATP. The mutant subunits physically associate with at least two endogenous members of mammalian SWI-SNF complexes, suggesting that nonfunctional, dominant negative complexes may be formed. We determined that expression of the mutant BRG1 or BRM proteins impaired the ability of cells to activate the endogenous stress response gene hsp70 in response to arsenite, a metabolic inhibitor, or cadmium, a heavy metal. Activation of hsp70 by heat stress, however, was unaffected. Activation of the heme oxygenase 1 promoter by arsenite or cadmium and activation of the cadmium-inducible metallothionein promoter also were unaffected by the expression of mutant SWI-SNF components. Analysis of a subset of constitutively expressed genes revealed no or minimal effects on transcript levels. We propose that the requirement for mammalian SWI-SNF complexes in gene activation events will be specific to individual genes and signaling pathways.The packaging of eukaryotic DNA into nucleosomes and higher order chromatin structure presents cells with a significant barrier to DNA utilization and necessitates mechanisms by which chromatin structure can be modified so that transcription can occur. Many multiprotein complexes with the ability to modify chromatin structure have been identified. These include histone acetyltransferases and deacetylases, which directly modify histone tail domains, and a class of energy-dependent enzymes that utilize ATP hydrolysis to alter nucleosome structure (reviewed in references 23, 30, 32, 34, 70, 83, and 84). The ATP-dependent chromatin remodeling complexes are conserved among eukaryotes, they share a related subunit that possesses DNA-stimulated ATPase activity, and each has been demonstrated to alter nucleosome structure in vitro in an ATP-dependent manner. Most of these complexes can be classified into two groups, those containing homologues of the yeast SWI2-SNF2 ATPase subunit, including yeast SWI-SNF (7, 12, 55), human SWI-SNF (hSWI-SNF) (24, 35, 82), yeast RSC (8), and Drosophila BRM complexes (54, 71), and those containing homologues of the Drosophila imitation-switch (ISWI) ATPase gene (16), including yeast ISW1 and ISW2 (76), human RSF (39), and the Drosophila NURF, CHRAC, and ACF complexes (25,75,78). A third group can be defined by Xenopus and human complexes containing the Mi2 protein, a related ATPase found in association with histone deacetylase activity (72,81,87,90).Although members of the ATP-dependent class of chromatin remodelers facilitate alterations in nucleosome structure in ...