Proliferation of memory-phenotype (CD44hi) CD8+ cells induced by infectious agents can be mimicked by injection of type I interferon (IFN I) and by IFN I-inducing agents such as lipopolysaccharide and Poly I:C; such proliferation does not affect naive T cells and appears to be TCR independent. Since IFN I inhibits proliferation in vitro, IFN I-induced proliferation of CD8+ cells in vivo presumably occurs indirectly through production of secondary cytokines, e.g., interleukin-2 (IL-2) or IL-15. We show here that, unlike IL-2, IL-15 closely mimics the effects of IFN I in causing strong and selective stimulation of memory-phenotype CD44hi CD8+ (but not CD4+) cells in vivo; similar specificity applies to purified T cells in vitro and correlates with much higher expression of IL-2Rbeta on CD8+ cells than on CD4+ cells.

T cell proliferation in vivo is presumed to reflect a T cell receptor (TCR)-mediated polyclonal response directed to various environmental antigens. However, the massive proliferation of T cells seen in viral infections is suggestive of a bystander reaction driven by cytokines instead of the TCR. In mice, T cell proliferation in viral infections preferentially affected the CD44hi subset of CD8+ cells and was mimicked by injection of polyinosinic-polycytidylic acid [poly(I:C)], an inducer of type I interferon (IFN I), and also by purified IFN I; such proliferation was not associated with up-regulation of CD69 or CD25 expression, which implies that TCR signaling was not involved. IFN I [poly(I:C)]-stimulated CD8+ cells survived for prolonged periods in vivo and displayed the same phenotype as did long-lived antigen-specific CD8+ cells. IFN I also potentiated the clonal expansion and survival of CD8+ cells responding to specific antigen. Production of IFN I may thus play an important role in the generation and maintenance of specific memory.

SUMMARY Recent studies suggest a hierarchical model in which lineage-determining factors act in a collaborative manner to select and prime cell-specific enhancers, thereby enabling signal-dependent transcription factors to bind and function in a cell type-specific manner. Consistent with this model, TLR4 signaling primarily regulates macrophage gene expression through a pre-existing enhancer landscape. However, TLR4 signaling also induces priming of ~3000 enhancer-like regions de novo, enabling visualization of intermediates in enhancer selection and activation. Unexpectedly, we find that enhancer transcription precedes local mono- and di-methylation of histone H3 lysine 4 (H3K4me1/2). H3K4 methylation at de novo enhancers is primarily dependent on the histone methyltransferases Mll1, Mll2/4 and Mll3, and is significantly reduced by inhibition of RNA polymerase II elongation. Collectively, these findings suggest an essential role of enhancer transcription in H3K4me1/2 deposition at de novo enhancers that is independent of potential functions of the resulting eRNA transcripts.

SummaryOn the basis of their surface markers, T lymphocytes are divided into subsets of "naive" and "memory cells". We have defined the interrelationship and relative life spans of naive and memory T cells by examining the surface markers on murine T cells incorporating bromodeoxyuridine, a DNA precursor, given in the drinking water. Three findings are reported. First, using a new method we show that the release of newly formed naive T cells from the unmanipulated thymus is very low (confirming the findings of others with surgical approaches). Second, in thymectomized mice, T cells with a naive phenotype remain in interphase for prolonged periods; however, some of these cells divide and retain (or regain) their "naive" markers. Third, most T cells with a memory phenotype divide rapidly, but others remain in interphase for many weeks. Collectively, the data indicate that long-lived T cells have multiple phenotypes and contain a mixture of memory cells, naive (virgin) cells, and memory cells masquerading as naive cells.O n the basis of their surface markers, T cells are divided into two broad groups of cells. In young life, most CD4 + and CD8 + T cells express high molecular weight isoforms ofCD45 (CD45RA/B/C) (1-7), a low density of CD44 (Pgp-1) (8, 9), and a high density of L-selectin (MEL-14) (10, 11). These cells are thought to be naive (virgin) cells that have remained in interphase since their release from the thymus (12, 13). Another population of T cells displays a reciprocal phenotype, i.e., CD45R 1~ CD44 hi L-selectin 1~ Since this phenotype is shared by T cells carrying memory to defined antigens (1--4, 8, 11), the T cells with this phenotype in normal animals are considered to be memory cells primed to various environmental antigens. T cells with a memory phenotype are rare in young life but are conspicuous in old age (14-17).Although it is well accepted that memory T cells arise from naive precursor cells (2,18,19), the interrelationship and relative life spans of these two types of T cells is controversial. In the case of naive T cells in rodents, many groups regard these cells as short-lived cells that either die within a few weeks of leaving the thymus or switch to memory cells (11). By contrast, other workers view naive T cells as potentially long-lived cells that can remain in interphase for many months (20). Similar uncertainty applies to the life span of memory cells. Until recently, it was generally assumed that memory to defined antigens is carried by a population of resting longlived lymphocytes (21). Now, however, there is increasing evidence that memory cells are short-lived unless the cells retain contact with residual antigen (22).In view of this controversy, the question arises whether the subsets of naive-and memory-phenotype T cells in normal animals differ in their rate of turnover. Direct evidence on this topic is sparse. Thus, although it is well established that T cells divide infrequently in vivo at a population level (13, 23), information on the relative turnover rates of naive ...

Memory T cells can be divided into central memory T cell (T CM cell) and effector memory T cell (T EM cell) subsets based on homing characteristics and effector functions. Whether T EM and T CM cells represent interconnected or distinct lineages is unclear, although the present paradigm suggests that T EM and T CM cells follow a linear differentiation pathway from naive T cells to effector T cells to T EM cells to T CM cells. We show here that naive T cell precursor frequency profoundly influenced the pathway along which CD8 + memory T cells developed. At low precursor frequency, those T EM cells generated represented a stable cell lineage that failed to further differentiate into T CM cells. These findings do not adhere to the present dogma regarding memory T cell generation and provide a means for identifying factors controlling memory T cell lineage commitment.Based on homing characteristics and effector functions, at least two types of memory T cells have been described in CD4 + and CD8 + T cell populations. The original descriptions of central and effector memory T cells suggested that central memory T cells (T CM cells) reside in lymphoid organs and express CCR7 and CD62L, whereas effector memory T cells (T EM cells) reside mainly in nonlymphoid tissues, do not express CCR7 or CD62L and have immediate effector functions 1-3 . This raised the question of how T CM cells and T EM cells are generated and whether each is the product of interdependent or separate lineages.Three models of differentiation have been proposed, with the first being that T CM cells provide a continual source of T EM cells. This model is based on the findings that memory CCR7 + T cells in short-term in vitro culture can lose expression of this chemokine receptor and in the process become functionally competent1 ,4 . Analysis of the T cell receptor (TCR) repertoire of human blood memory CD8 + T cells has suggested an additional possibility in which T CM and T EM cells represent mostly separate lineages 5 . In contrast, an alternative model has indicated that over time T EM cells convert to T CM cells 6 . This conclusion was derived from analysis of TCR-transgenic CD8 + memory T cells specific for lymphocytic choriomeningitis virus (LCMV) glycoprotein 33 (gp33) that had been separated by virtue of

The two tandem bromodomains of the BET proteins enable chromatin binding to facilitate transcription. Drugs that inhibit both bromodomains equally have shown efficacy in certain malignant and inflammatory conditions. To explore the individual functional contributions of the first (BD1) and second (BD2) bromodomains in biology and therapy, we developed selective BD1 and BD2 inhibitors. We found that steady-state gene expression primarily requires BD1 whereas the rapid increase of gene expression induced by inflammatory stimuli requires both BD1 and BD2 of all BET proteins. BD1 inhibitors phenocopied the effects of pan-BET inhibitors in cancer models whereas BD2 inhibitors were predominantly effective in models of inflammatory and autoimmune disease. These insights into the differential requirement of BD1 and BD2 for the maintenance and induction of gene expression may guide future BET targeted therapies.

Resting dendritic cells (DCs) are resident in most tissues and can be activated by environmental stimuli to mature into potent antigen-presenting cells. One important stimulus for DC activation is infection; DCs can be triggered through receptors that recognize microbial components directly or by contact with infectioninduced cytokines. We show here that murine DCs undergo phenotypic maturation upon exposure to type I interferons (type I IFNs) in vivo or in vitro. Moreover, DCs either derived from bone marrow cells in vitro or isolated from the spleens of normal animals express IFN-␣ and IFN-␤, suggesting that type I IFNs can act in an autocrine manner to activate DCs. Consistent with this idea, the ability to respond to type I IFN was required for the generation of fully activated DCs from bone marrow precursors, as DCs derived from the bone marrow of mice lacking a functional receptor for type I IFN had reduced expression of costimulatory and adhesion molecules and a diminished ability to stimulate naive T-cell proliferation compared with DCs derived from control bone marrow. IntroductionDendritic cells (DCs) are recognized as the key antigen-presenting cells (APCs) controlling the initiation of T cell-dependent immune responses. 1 DCs not only are the most potent APCs for activation of resting T cells, but can also regulate the type of response made, dictating the cytokines expressed by responding T cells. 2 Furthermore, DCs have the ability to down-regulate T-cell activation and may play a role in the induction of tolerance. [3][4][5] The type of response elicited by particular DCs is likely to reflect their developmental origin, anatomical location, and state of activation. 6 Immature (resting) DCs reside in most tissues and are efficient in binding and internalizing antigens. 1 However, these cells are relatively poor at presenting antigen and inducing T-cell activation, owing at least in part to their low cell surface expression of costimulatory and adhesion molecules. In response to a variety of stimuli (see below), DCs undergo a process that has been variably termed activation or maturation, during which they lose their capacity for antigen uptake and acquire potent T-cell stimulatory ability. This is associated with up-regulation of cell surface major histocompatibility complex (MHC) class II, costimulatory molecules (eg, CD80, CD86, CD40), and adhesion molecules (eg, CD54). 1 DC activation can be induced by signals that have been described as indicative of "danger," such as heat shock proteins, 7 mechanical manipulation, 8 or exposure to necrotic cells, 8,9 as well as components of the extracellular matrix, 10 interaction with activated (CD40 ligand-expressing)T cells, 11-14 and infection. DCs are particularly tuned to recognition of infection, as DC activation can be stimulated by exposure to whole pathogens (viruses or bacteria [15][16][17] One family of infection-induced cytokines that can activate DCs is the type I interferons (type I IFNs). Type I IFNs include a number of evolutionarily conserv...

Three distinct subtypes of dendritic cells (DC) are present in mouse spleen, separable as CD4−8α−, CD4+8α−, and CD4−8α+ DC. We have tested whether these represent stages of development or activation within one DC lineage, or whether they represent separate DC lineages. All three DC subtypes appear relatively mature by many criteria, but all retain a capacity to phagocytose particulate material in vivo. Although further maturation or activation could be induced by bacterially derived stimuli, phagocytic capacity was retained, and no DC subtype was converted to the other. Continuous elimination of CD4+8− DC by Ab depletion had no effect on the levels of the other DC subtypes. Bromodeoxyuridine labeling experiments indicated that all three DC subtypes have a rapid turnover (half-life, 1.5–2.9 days) in the spleen, with none being the precursor of another. The three DC subtypes showed different kinetics of development from bone marrow precursors. The CD8α+ spleen DC, apparently the most mature, displayed an extremely rapid turnover based on bromodeoxyuridine uptake and the fastest generation from bone marrow precursors. In conclusion, the three splenic DC subtypes behave as rapidly turning over products of three independent developmental streams.