Avinash Bhandoola

Thymopoiesis independent of common lymphoid progenitors

Early T lineage progenitors (ETPs) in the thymus are thought to develop from common lymphoid progenitors (CLPs) in the bone marrow (BM). We compared thymic ETPs to BM CLPs in mice and found that they differed in several respects. Thymic ETPs were not interleukin 7 (IL-7)–responsive and generated B lineage progeny with delayed kinetics, whereas BM CLPs were IL-7–responsive and rapidly generated B cells. ETPs sustained production of T lineage progeny for longer periods of time than BM CLPs. Analysis of Ikaros-deficient mice that exhibit ongoing thymopoiesis without B lymphopoeisis revealed near-normal frequencies of thymic ETPs, yet undetectable numbers of BM CLPs. We conclude that ETPs can develop via a CLP-independent pathway.

Deletion of the developmentally essential gene ATR in adult mice leads to age-related phenotypes and stem cell loss.

Developmental abnormalities, cancer, and premature aging each have been linked to defects in the DNA damage response (DDR). Mutations in the ATR checkpoint regulator cause developmental defects in mice (pregastrulation lethality) and humans (Seckel syndrome). Here we show that eliminating ATR in adult mice leads to defects in tissue homeostasis and the rapid appearance of age-related phenotypes, such as hair graying, alopecia, kyphosis, osteoporosis, thymic involution, fibrosis, and other abnormalities. Histological and genetic analyses indicate that ATR deletion causes acute cellular loss in tissues in which continuous cell proliferation is required for maintenance. Importantly, thymic involution, alopecia, and hair graying in ATR knockout mice were associated with dramatic reductions in tissue-specific stem and progenitor cells and exhaustion of tissue renewal and homeostatic capacity. In aggregate, these studies suggest that reduced regenerative capacity in adults via deletion of a developmentally essential DDR gene is sufficient to cause the premature appearance of age-related phenotypes.

IL25 elicits a multipotent progenitor cell population that promotes TH2 cytokine responses

CD4+ T helper 2 (TH2) cells secrete interleukin (IL)4, IL5 and IL13, and are required for immunity to gastrointestinal helminth infections. However, TH2 cells also promote chronic inflammation associated with asthma and allergic disorders. The non-haematopoietic-cell-derived cytokines thymic stromal lymphopoietin, IL33 and IL25 (also known as IL17E) have been implicated in inducing TH2 cell-dependent inflammation at mucosal sites, but how these cytokines influence innate immune responses remains poorly defined. Here we show that IL25, a member of the IL17 cytokine family, promotes the accumulation of a lineage-negative (Lin-) multipotent progenitor (MPP) cell population in the gut-associated lymphoid tissue that promotes TH2 cytokine responses. The IL25-elicited cell population, termed MPPtype2 cells, was defined by the expression of Sca-1 (also known as Ly6a) and intermediate expression of c-Kit (c-Kitint), and exhibited multipotent capacity, giving rise to cells of monocyte/macrophage and granulocyte lineages both in vitro and in vivo. Progeny of MPPtype2 cells were competent antigen presenting cells, and adoptive transfer of MPPtype2 cells could promote TH2 cytokine responses and confer protective immunity to helminth infection in normally susceptible Il25-/- mice. The ability of IL25 to induce the emergence of an MPPtype2 cell population identifies a link between the IL17 cytokine family and extramedullary haematopoiesis, and suggests a previously unrecognized innate immune pathway that promotes TH2 cytokine responses at mucosal sites.

The earliest thymic progenitors for T cells possess myeloid lineage potential

There exists controversy over the nature of haematopoietic progenitors of T cells. Most T cells develop in the thymus, but the lineage potential of thymus-colonizing progenitors is unknown. One approach to resolving this question is to determine the lineage potentials of the earliest thymic progenitors (ETPs). Previous work has shown that ETPs possess T and natural killer lymphoid potentials, and rare subsets of ETPs also possess B lymphoid potential, suggesting an origin from lymphoid-restricted progenitor cells. However, whether ETPs also possess myeloid potential is unknown. Here we show that nearly all ETPs in adult mice possess both T and myeloid potential in clonal assays. The existence of progenitors possessing T and myeloid potential within the thymus is incompatible with the current dominant model of haematopoiesis, in which T cells are proposed to arise from lymphoid-. Our results indicate that alternative models for lineage commitment during haematopoiesis must be considered.

Notch signaling controls the generation and differentiation of early T lineage progenitors

Signaling by the transmembrane receptor Notch is critical for T lineage development, but progenitor subsets that first receive Notch signals have not been defined. Here we identify an immature subset of early T lineage progenitors (ETPs) in the thymus that expressed the tyrosine kinase receptor Flt3 and had preserved B lineage potential at low progenitor frequency. Notch signaling was active in ETPs and was required for generation of the ETP population. Additionally, Notch signals contributed to the subsequent differentiation of ETPs. In contrast, multipotent hematopoietic progenitors circulated in the blood even in the absence of Notch signaling, suggesting that critical Notch signals during early T lineage development are delivered early after thymic entry.

Canonical notch signaling is dispensable for the maintenance of adult hematopoietic stem cells.

Gain-of-function experiments have demonstrated the potential of Notch signals to expand primitive hematopoietic progenitors, but whether Notch physiologically regulates hematopoietic stem cell (HSC) homeostasis in vivo is unclear. To answer this question, we evaluated the effect of global deficiencies of canonical Notch signaling in rigorous HSC assays. Hematopoietic progenitors expressing dominant-negative Mastermind-like1 (DNMAML), a potent inhibitor of Notch-mediated transcriptional activation, achieved stable long-term reconstitution of irradiated hosts and showed a normal frequency of progenitor fractions enriched for long-term HSCs. Similar results were observed with cells lacking CSL/RBPJ, a DNA-binding factor that is required for canonical Notch signaling. Notch-deprived progenitors provided normal long-term reconstitution after secondary competitive transplantation. Furthermore, Notch target genes were expressed at low levels in primitive hematopoietic progenitors. Taken together, these results rule out an essential physiological role for cell-autonomous canonical Notch signals in HSC maintenance.

Adaptation of Innate Lymphoid Cells to a Micronutrient Deficiency Promotes Type 2 Barrier Immunity

How the immune system adapts to malnutrition to sustain immunity at barrier surfaces, such as the intestine, remains unclear. Vitamin A deficiency is one of the most common micronutrient deficiencies and is associated with profound defects in adaptive immunity. Here, we found that type 3 innate lymphoid cells (ILC3s) are severely diminished in vitamin A–deficient settings, which results in compromised immunity to acute bacterial infection. However, vitamin A deprivation paradoxically resulted in dramatic expansion of interleukin-13 (IL-13)–producing ILC2s and resistance to nematode infection in mice, which revealed that ILCs are primary sensors of dietary stress. Further, these data indicate that, during malnutrition, a switch to innate type 2 immunity may represent a powerful adaptation of the immune system to promote host survival in the face of ongoing barrier challenges. Vitamin A deficiency alters the balance of innate immune cells in the gut, promoting resistance to nematode infection. An Immune Response to Malnutrition Mucosal surfaces, such as those lining the intestine, are in constant contact with potentially pathogenic microbes, including bacteria and parasitic worms. This necessitates so-called barrier immunity, which is mediated in part by innate lymphoid cells, subsets of which combat specific types of infection. Although malnutrition has been associated with immunosuppression, Spencer et al. (p. 432) now show that vitamin A deficiency selectively activates one branch of barrier immunity. Vitamin A deficiency in mice enhanced immunity to chronic worm infections by increasing the levels of one subset of innate lymphoid cells lacking the corresponding retinoic acid receptor. In contrast, another innate lymphoid cell subset that carries the vitamin A receptor and is important for bacterial immunity was depleted. Thus, the immune system can adapt its response to dietary stress, thereby promoting host survival.

Circulating hematopoietic progenitors with T lineage potential

The thymus is seeded via the blood, but the identity of hematopoietic progenitors with access to the circulation in adult mice is unknown. We report here that the only progenitors in blood with efficient T lineage potential were lineage negative with high expression of stem cell antigen 1 and c-Kit (LSK cells). The blood LSK population, like its counterpart in the bone marrow, contained hematopoietic stem cells and nonrenewing, multipotent progenitors, including early lymphoid progenitors and CD62L+ cells previously described as efficient T lineage progenitors. Common lymphoid progenitors could not be identified in the circulation, suggesting they are not physiological T lineage precursors. We conclude that blood LSK cells are the principal circulating progenitors with T lineage potential.

A critical role for TCF-1 in T-lineage specification and differentiation

The vertebrate thymus provides an inductive environment for T-cell development. Within the mouse thymus, Notch signals are indispensable for imposing the T-cell fate on multipotential haematopoietic progenitors, but the downstream effectors that impart T-lineage specification and commitment are not well understood. Here we show that a transcription factor, T-cell factor 1 (TCF-1; also known as transcription factor 7, T-cell specific, TCF7), is a critical regulator in T-cell specification. TCF-1 is highly expressed in the earliest thymic progenitors, and its expression is upregulated by Notch signals. Most importantly, when TCF-1 is forcibly expressed in bone marrow (BM) progenitors, it drives the development of T-lineage cells in the absence of T-inductive Notch1 signals. Further characterization of these TCF-1-induced cells revealed expression of many T-lineage genes, including T-cell-specific transcription factors Gata3 and Bcl11b, and components of the T-cell receptor. Our data suggest a model where Notch signals induce TCF-1, and TCF-1 in turn imprints the T-cell fate by upregulating expression of T-cell essential genes.

The Earliest Step in B Lineage Differentiation from Common Lymphoid Progenitors Is Critically Dependent upon Interleukin 7

Little is known about the signals that promote early B lineage differentiation from common lymphoid progenitors (CLPs). Using a stromal-free culture system, we show that interleukin (IL)-7 is sufficient to promote the in vitro differentiation of CLPs into B220+ CD19+ B lineage progenitors. Consistent with current models of early B cell development, surface expression of B220 was initiated before CD19 and was accompanied by the loss of T lineage potential. To address whether IL-7 receptor (R) activity is essential for early B lineage development in vivo, we examined the frequencies of CLPs and downstream pre–pro- and pro-B cells in adult mice lacking either the α chain or the common gamma chain (γc) of the IL-7R. The data indicate that although γc −/− mice have normal frequencies of CLPs, both γc −/− and IL-7Rα−/− mice lack detectable numbers of all downstream early B lineage precursors, including pre–pro-B cells. These findings challenge previous notions regarding the point in B cell development affected by the loss of IL-7R signaling and suggest that IL-7 plays a key and requisite role during the earliest phases of B cell development.