Regulatory T (Treg) cells maintain immune homeostasis and prevent inflammatory and autoimmune responses. During development, thymocytes bearing a moderately self-reactive T cell receptor (TCR) can be selected to become Treg cells. Several observations suggest that also in the periphery mature Treg cells continuously receive self-reactive TCR signals. However, the importance of this inherent autoreactivity for Treg cell biology remains poorly defined. To address this open question, we genetically ablated the TCR of mature Treg cells in vivo. These experiments revealed that TCR-induced Treg lineage-defining Foxp3 expression and gene hypomethylation were uncoupled from TCR input in mature Treg cells. However, Treg cell homeostasis, cell-type-specific gene expression and suppressive function critically depend on continuous triggering of their TCR.

IntroductionB cells play essential roles during protective immune responses to invading pathogens. On encounter of foreign antigen and with cognate T-cell help, B lymphocytes proliferate and form distinct histologic structures, termed germinal center (GC). In the GC, they undergo somatic hypermutation and class-switch recombination. During somatic hypermutation, they introduce random mutations into their immunoglobulin variable regions while they exchange the heavy chain constant region during class-switch recombination to allow for different effector functions. After a selection process by antigen, B cells differentiate into memory B cells and plasma cells (PCs), which secrete antibodies. 1 The deregulation of this process is heavily implicated in human disease. Production of class-switched antibodies against self-antigens causes or contributes to various autoimmune syndromes and unrestrained B-cell proliferation and survival can result in lymphomas. 1,2 It is thought that the majority of human lymphomas derive from the GC, probably because the DNA damage inherent to the GC reaction facilitates mutations and chromosomal translocations. 1,3 Recently, the ubiquitin-editing enzyme A20, encoded by the tumor necrosis factor-␣-inducible gene 3 (TNFAIP3), has been associated with both autoimmunity and lymphomagenesis. Polymorphisms and mutations in or near the TNFAIP3 genomic locus have been linked with various human autoimmune syndromes with a strong humoral component, such as systemic lupus erythematosus (SLE), 4,5 rheumatoid arthritis, 6,7 and celiac disease. 8 Loss of A20 function through mutations, chromosomal deletions, and/or promoter methylation is a frequent event in several human lymphomas, 9-12 all of which are characterized by constitutive activation of nuclear factor-B (NF-B). 13 These factors regulate a plethora of genes encoding for proinflammatory mediators, antiapoptotic proteins, cell adhesion molecules and, for negative feedback control, inhibitory proteins, such as p100, IB␣, and A20. 14,15 During the transmission of NF-B activating signals from cell-surface receptors such as the B-cell receptor (BCR), CD40, or Toll-like receptors (TLRs), signal transduction occurs via the attachment of polyubiquitin chains to key proteins, including MALT1 or TRAF6. Polyubiquitin chains, linked via K63 or linear assembly, serve to recruit different kinase complexes. In the case of canonical NF-B, induced proximity allows the upstream kinase TAK1 to phosphorylate its target IKK2, which then effects NF-B activation. A20, whose transcription is induced by NF-B, dampens signaling through 2 main activities. First, as deubiquitinase A20 removes K63-linked polyubiquitin chains from essential signaling intermediates, such as TRAF6. Second, A20 induces, in concert with other proteins, degradation of some of its molecular targets, through addition of K48-linked ubiquitin chains. 14,16 Degradation of RIP1 limits TNF-induced signaling, 14 whereas degradation of the K63-chain-specific E2 ligases Ubc13/UbcH5c generally affects t...

By genetically ablating IκB kinase (IKK)-mediated NF-κB activation in the B cell lineage, and by analyzing a mouse mutant in which Igλ+ B cells are generated in the absence of rearrangements in Igk, we define two distinct, consecutive phases of early B cell development that differ in their dependence on IKK-mediated NF-κB signaling. During the first phase, in which NF-κB signaling is dispensable, predominantly Igκ+ B cells are generated and undergo efficient receptor editing. In the second phase, predominantly Igλ+ B cells are generated, whose development is ontogenetically timed to occur after Igk rearrangements. This second phase of development is dependent on NF-κB signals, which can be substituted by transgenic expression of the pro-survival factor Bcl2.

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