Benjamin D. McDonald

A committed precursor to innate lymphoid cells

Innate lymphoid cells (ILCs) specialize in the rapid secretion of polarized sets of cytokines and chemokines to combat infection and promote tissue repair at mucosal barriers. Their diversity and similarities with previously characterized natural killer (NK) cells and lymphoid tissue inducers (LTi) have prompted a provisional classification of all innate lymphocytes into groups 1, 2 and 3 solely on the basis of cytokine properties, but their developmental pathways and lineage relationships remain elusive. Here we identify and characterize a novel subset of lymphoid precursors in mouse fetal liver and adult bone marrow that transiently express high amounts of PLZF, a transcription factor previously associated with NK T cell development, by using lineage tracing and transfer studies. PLZFhigh cells were committed ILC progenitors with multiple ILC1, ILC2 and ILC3 potential at the clonal level. They excluded classical LTi and NK cells, but included a peculiar subset of NK1.1+DX5− ‘NK-like’ cells residing in the liver. Deletion of PLZF markedly altered the development of several ILC subsets, but not LTi or NK cells. PLZFhigh precursors also expressed high amounts of ID2 and GATA3, as well as TOX, a known regulator of PLZF-independent NK and LTi lineages. These findings establish novel lineage relationships between ILC, NK and LTi cells, and identify the common precursor to ILCs, termed ILCP. They also reveal the broad, defining role of PLZF in the differentiation of innate lymphocytes.

PLZF expression maps the early stages of ILC1 lineage development

Significance Diverse populations of group 1 innate lymphocytes, which exert critical early cytolytic functions against virally infected cells, have recently been discovered, raising issues of lineage relationships. We used expression of the transcription factor promyelocytic leukaemia zinc finger (PLZF) to identify the developmental intermediates of innate lymphoid cells type 1 (ILC1s), a subset of innate lymphoid cells that are particularly abundant in the liver, and demonstrated that this lineage arises from a distinct precursor, but that its development partially overlaps with established classical NK stages. Using microarray analysis, we defined a set of PLZF-dependent genes that may contribute to lineage divergence between ILC1s and classical NK cells. Among the variety of tissue-resident NK-like populations recently distinguished from recirculating classical NK (cNK) cells, liver innate lymphoid cells (ILC) type 1 (ILC1s) have been shown to represent a distinct lineage that originates from a novel promyelocytic leukaemia zinc finger (PLZF)-expressing ILC precursor (ILCP) strictly committed to the ILC1, ILC2, and ILC3 lineages. Here, using PLZF-reporter mice and cell transfer assays, we studied the developmental progression of ILC1s and demonstrated substantial overlap with stages previously ascribed to the cNK lineage, including pre–pro-NK, pre-NK precursor (pre-NKP), refined NKP (rNKP), and immature NK (iNK). Although they originated from different precursors, the ILC1 and cNK lineages followed a parallel progression at early stages and diverged later at the iNK stage, with a striking predominance of ILC1s over cNKs early in ontogeny. Although a limited set of ILC1 genes depended on PLZF for expression, characteristically including Il7r, most of these genes were also differentially expressed between ILC1s and cNKs, indicating that PLZF together with other, yet to be defined, factors contribute to the divergence between these lineages.

Elevated T Cell Receptor Signaling Identifies a Thymic Precursor to the TCRαβ+CD4−CD8β− Intraepithelial Lymphocyte Lineage

The origin and developmental pathway of intestinal T cell receptor αβ(+) CD4(-)CD8β(-) intraepithelial lymphocytes (unconventional iIELs), a major population of innate-like resident cytolytic T cells, have remained elusive. By cloning and expressing several TCRs isolated from unconventional iIELs, we identified immature CD4(lo)CD8(lo)(DP(lo))CD69(hi)PD-1(hi) thymocytes as the earliest postsignaling precursors for these cells. Although these precursors displayed multiple signs of elevated TCR signaling, a sizeable fraction of them escaped deletion to selectively engage in unconventional iIEL differentiation. Conversely, TCRs cloned from DP(lo)CD69(hi)PD-1(hi) thymocytes, a population enriched in autoreactive thymocytes, selectively gave rise to unconventional iIELs upon transgenic expression. Thus, the unconventional iIEL precursor overlaps with the DP(lo) population undergoing negative selection, indicating that, concomitant with the downregulation of both CD4 and CD8 coreceptors, a balance between apoptosis and survival signals results in outcomes as divergent as clonal deletion and differentiation to the unconventional iIEL lineage.

A negative feedback loop mediated by the Bcl6–cullin 3 complex limits Tfh cell differentiation

Bcl6 and E3 ligase cullin 3 complexes mediate negative feedback regulation during thymocyte development and T cell activation to restrain exaggerated Tfh responses.

Polarized effector programs for innate-like thymocytes

Distinct lineages of natural killer T cells arise in the thymus and are characterized by the signature transcription factors T-bet, GATA-3 or RORγt.

Microbial signals drive pre-leukaemic myeloproliferation in a Tet2 -deficient host

Somatic mutations in tet methylcytosinedioxygenase 2 (TET2), which encodes an epigenetic modifier enzyme, drive the development of haematopoietic malignancies1–7. In both humans and mice, TET2 deficiency leads to increased self-renewal of haematopoietic stem cells with a net developmental bias towards the myeloid lineage1,4,8,9. However, pre-leukaemic myeloproliferation (PMP) occurs in only a fraction of Tet2−/− mice8,9 and humans with TET2 mutations1,3,5–7, suggesting that extrinsic non-cell-autonomous factors are required for disease onset. Here we show that bacterial translocation and increased interleukin-6 production, resulting from dysfunction of the small-intestinal barrier, are critical for the development of PMP in mice that lack Tet2 expression in haematopoietic cells. Furthermore, in symptom-free Tet2−/− mice, PMP can be induced by disrupting intestinal barrier integrity, or in response to systemic bacterial stimuli such as the toll-like receptor 2 agonist. PMP was reversed by antibiotic treatment and failed to develop in germ-free Tet2−/− mice, which illustrates the importance of microbial signals in the development of this condition. Our findings demonstrate the requirement for microbial-dependent inflammation in the development of PMP and provide a mechanistic basis for the variation in PMP penetrance observed in Tet2−/− mice. This study will prompt new lines of investigation that may profoundly affect the prevention and management of haematopoietic malignancies.Microbial signals are crucial to the development of pre-leukaemic myeloproliferation, which can be induced by disrupting the intestinal barrier or by introducing systemic bacterial stimuli in Tet2-deficient mice.

Diverse developmental pathways of intestinal intraepithelial lymphocytes

The intestinal epithelial barrier is patrolled by resident intraepithelial lymphocytes (IELs) that are involved in host defence against pathogens, wound repair and homeostatic interactions with the epithelium, microbiota and nutrients. Intestinal IELs are one of the largest populations of lymphocytes in the body and comprise several distinct subsets, the identity and lineage relationships of which have long remained elusive. Here, we review advances in unravelling the complexity of intestinal IEL populations, which comprise conventional αβ T cell receptor (TCRαβ)+ subsets, unconventional TCRαβ+ and TCRγδ+ subsets, group 1 innate lymphoid cells (ILC1s) and ILC1-like cells. Although these intestinal IEL lineages have partially overlapping effector programmes and recognition properties, they have strikingly different developmental pathways. We suggest that evolutionary pressure has driven the recurrent generation of cytolytic effector lymphocytes to protect the intestinal epithelial layer, but they may also precipitate intestinal inflammatory disorders, such as coeliac disease.The intestine harbours a complex array of intraepithelial lymphocytes that patrol the epithelial layer and exert rapid cytolytic function. Here, the authors compare the different cytolytic populations and suggest that this diversity results from evolutionary pressure to protect this important barrier.