Heinrich Schlums

Cytomegalovirus Infection Drives Adaptive Epigenetic Diversification of NK Cells with Altered Signaling and Effector Function

The mechanisms underlying human natural killer (NK) cell phenotypic and functional heterogeneity are unknown. Here, we describe the emergence of diverse subsets of human NK cells selectively lacking expression of signaling proteins after human cytomegalovirus (HCMV) infection. The absence of B and myeloid cell-related signaling protein expression in these NK cell subsets correlated with promoter DNA hypermethylation. Genome-wide DNA methylation patterns were strikingly similar between HCMV-associated adaptive NK cells and cytotoxic effector T cells but differed from those of canonical NK cells. Functional interrogation demonstrated altered cytokine responsiveness in adaptive NK cells that was linked to reduced expression of the transcription factor PLZF. Furthermore, subsets of adaptive NK cells demonstrated significantly reduced functional responses to activated autologous T cells. The present results uncover a spectrum of epigenetically unique adaptive NK cell subsets that diversify in response to viral infection and have distinct functional capabilities compared to canonical NK cell subsets.

Molecular Mechanisms of Natural Killer Cell Activation

With an array of activating and inhibitory receptors, natural killer (NK) cells can specifically eradicate infected and transformed cells. Target cell killing is achieved through directed release of lytic granules. Recognition of target cells also induces production of chemokines and cytokines that can coordinate immune responses. Upon contact with susceptible cells, a multiplicity of activating receptors can induce signals for adhesion. Engagement of the integrin leukocyte functional antigen-1 mediates firm adhesion, provides signals for granule polarization and orchestrates the structure of an immunological synapse that facilitates efficient target cell killing. Other activating receptors apart from leukocyte functional antigen-1 signal for lytic granule exocytosis, a process that requires overcoming a threshold for activation of phospholipase C-γ, which in turn induces STIM1- and ORAI1-dependent store-operated Ca2+ entry as well as exocytosis mediated by the SNARE-containing protein syntaxin-11 and regulators thereof. Cytokine and chemokine release follows a different secretory pathway which also requires phospholipase C-γ activation and store-operated Ca2+ entry. Recent studies of human NK cells have provided insights into a hierarchy of effector functions that result in graded responses by NK cell populations. Responses display cellular heterogeneity and are influenced by environmental cues. This review highlights recent knowledge gained on the molecular pathways for and regulation of NK cell activation.

CD56dimCD57+NKG2C+ NK cell expansion is associated with reduced leukemia relapse after reduced intensity HCT.

We have recently described a specialized subset of human natural killer (NK) cells with a CD56dimCD57+NKG2C+ phenotype that expand specifically in response to cytomegalovirus (CMV) reactivation in hematopoietic cell transplant (HCT) recipients and exhibit properties characteristic of adaptive immunity. We hypothesize that these cells mediate relapse protection and improve post-HCT outcomes. In 674 allogeneic HCT recipients, we found that those who reactivated CMV had lower leukemia relapse (26% (17–35%), P=0.05) and superior disease-free survival (DFS) (55% (45–65%) P=0.04) 1 year after reduced intensity conditioning (RIC) compared with CMV seronegative recipients who experienced higher relapse rates (35% (27–43%)) and lower DFS (46% (38–54%)). This protective effect was independent of age and graft-vs-host disease and was not observed in recipients who received myeloablative regimens. Analysis of the reconstituting NK cells demonstrated that CMV reactivation is associated with both higher frequencies and greater absolute numbers of CD56dimCD57+NKG2C+ NK cells, particularly after RIC HCT. Furthermore, expansion of these cells at 6 months posttransplant independently trended toward a lower 2-year relapse risk. Together, our data suggest that the protective effect of CMV reactivation on posttransplant relapse is in part driven by adaptive NK cell responses.

CD49a Expression Defines Tissue-Resident CD8+ T Cells Poised for Cytotoxic Function in Human Skin

SUMMARY Tissue‐resident memory T (Trm) cells form a heterogeneous population that provides localized protection against pathogens. Here, we identify CD49a as a marker that differentiates CD8+ Trm cells on a compartmental and functional basis. In human skin epithelia, CD8+CD49a+ Trm cells produced interferon‐&ggr;, whereas CD8+CD49a− Trm cells produced interleukin‐17 (IL‐17). In addition, CD8+CD49a+ Trm cells from healthy skin rapidly induced the expression of the effector molecules perforin and granzyme B when stimulated with IL‐15, thereby promoting a strong cytotoxic response. In skin from patients with vitiligo, where melanocytes are eradicated locally, CD8+CD49a+ Trm cells that constitutively expressed perforin and granzyme B accumulated both in the epidermis and dermis. Conversely, CD8+CD49a– Trm cells from psoriasis lesions predominantly generated IL‐17 responses that promote local inflammation in this skin disease. Overall, CD49a expression delineates CD8+ Trm cell specialization in human epithelial barriers and correlates with the effector cell balance found in distinct inflammatory skin diseases. Graphical Abstract Figure. No Caption available. HighlightsCD49a expression marks CD8+ Trm cells poised for IFN‐&ggr; production in human skinIL‐15 drives potent cytotoxic capacity in CD49a+ Trm cellsIL‐17 is preferentially produced by CD49a− CD8+ Trm cells in the skinCD49a+ versus CD49‐ Trm cell functional dichotomy is preserved in vitiligo and psoriasis &NA; Tissue‐resident memory T (Trm) cells provide localized adaptive immunity in peripheral tissues. Cheuk et al. identify cytotoxic CD49a+CD8+ Trm cells and IL‐17‐producing CD49a−CD8+ Trm cells in healthy human skin. The functional dichotomy of pathogenic Trm cells based on CD49a expression is preserved in focal skin diseases vitiligo and psoriasis.

Increased proportion of mature NK cells is associated with successful imatinib discontinuation in chronic myeloid leukemia

Recent studies suggest that a proportion of chronic myeloid leukemia (CML) patients in deep molecular remission can discontinue the tyrosine kinase inhibitor (TKI) treatment without disease relapse. In this multi-center, prospective clinical trial (EURO-SKI, NCT01596114) we analyzed the function and phenotype of T and NK cells and their relation to successful TKI cessation. Lymphocyte subclasses were measured from 100 imatinib-treated patients at baseline and 1 month after the discontinuation, and functional characterization of NK and T cells was done from 45 patients. The proportion of NK cells was associated with the molecular relapse-free survival as patients with higher than median NK-cell percentage at the time of drug discontinuation had better probability to stay in remission. Similar association was not found with T or B cells or their subsets. In non-relapsing patients the NK-cell phenotype was mature, whereas patients with more naïve CD56bright NK cells had decreased relapse-free survival. In addition, the TNF-α/IFN-γ cytokine secretion by NK cells correlated with the successful drug discontinuation. Our results highlight the role of NK cells in sustaining remission and strengthen the status of CML as an immunogenic tumor warranting novel clinical trials with immunomodulating agents.

Diversification and Functional Specialization of Human NK Cell Subsets.

Natural killer (NK) cells are lymphocytes that participate in different facets of immunity. They can act as innate sentinels through recognition and eradication of infected or transformed target cells, so-called immunosurveillance. In addition, they can contain immune responses through the killing of other activated immune cells, so-called immunoregulation. Furthermore, they instruct and regulate immune responses by producing pro-inflammatory cytokines such as IFN-γ, either upon direct target cell recognition or by relaying cytokine cues from various cell types. Recent studies in mouse and man have uncovered infection-associated expansions of NK cell subsets with specific receptor repertoires and diverse patterns of intracellular signaling molecule expression. Moreover, distinct attributes of NK cells in tissues, including tissue-resident subsets, are being further elucidated. Findings support an emerging theme of ever-increasing diversification and functional specialization among different NK cell subsets, with a functional dichotomy between subsets involved in immunoregulation or immunosurveillance. The epigenetic landscapes and transcriptional profiles of different NK cell subsets are providing insights into the molecular regulation of effector functions. Here, we review phenotypic, functional, and developmental characteristics of a spectrum of human NK cell subsets. We also discuss the molecular underpinnings of different NK cell subsets and their potential contributions to immunity as well as disease susceptibility.

Transcriptional regulation of Munc13-4 expression in cytotoxic lymphocytes is disrupted by an intronic mutation associated with a primary immunodeficiency

A conserved regulatory element in intron 1 of UNC13D regulates Munc13-4 expression.

Novel deep intronic and missense UNC13D mutations in familial haemophagocytic lymphohistiocytosis type 3.

Odile Beyne-Rauzy Sabine Brechignac Sylvestre Marechaux Dominique Vassilieff Olivier Ernst C eline Berthon Emmanuel Gyan Marie-Pierre Gourin François Dreyfus Pierre Fenaux Christian Rose Service d’H ematologie et de cardiologie Hôpital Saint Vincent de Paul, Universit e Catholique de Lille, Lille, Service de m edecine interne, CHU Toulouse-Purpan, Toulouse, GFM, Groupe Francophone des My elodysplasies, Hopital Avicenne, Paris (AP-HP), Hôpital Avicenne, Assistance Publique des Hôpitaux de Paris (AP-HP), Hôpital Cochin, AP-HP, Paris, Radiologie, Hôpital Huriez, CHU de Lille, H ematologie, CHU de Lille, Lille, , H ematologie, Hôpital Bretonneau, CHU de Tours, Tours, and CHU de Limoges, Limoge, France E-mail: rose.christian@ghicl.net

Systemic Lupus Erythematosus Immune Complexes Increase the Expression of SLAM Family Members CD319 (CRACC) and CD229 (LY-9) on Plasmacytoid Dendritic Cells and CD319 on CD56 dim NK Cells

Patients with systemic lupus erythematosus (SLE) display an activated type I IFN system due to unceasing IFN-α release from plasmacytoid dendritic cells (pDCs) stimulated by nucleic acid–containing immune complexes (ICs). NK cells strongly promote the IFN-α production by pDCs; therefore, we investigated surface molecules that could be involved in the pDC–NK cell cross-talk. In human PBMCs stimulated with RNA-containing ICs (RNA-ICs), the expression of the signaling lymphocyte activation molecule (SLAM) family receptors CD319 and CD229 on pDCs and CD319 on CD56dim NK cells was selectively increased. Upregulation of CD319 and CD229 on RNA-IC–stimulated pDCs was induced by NK cells or cytokines (e.g., GM-CSF, IL-3). IFN-α–producing pDCs displayed a higher expression of SLAM molecules compared with IFN-α− pDCs. With regard to signaling downstream of SLAM receptors, pDCs expressed SHIP-1, SHP-1, SHP-2, and CSK but lacked SLAM-associated protein (SAP) and Ewing’s sarcoma-activated transcript 2 (EAT2), indicating that these receptors may act as inhibitory receptors on pDCs. Furthermore, pDCs from patients with SLE had decreased expression of CD319 on pDCs and CD229 on CD56dim NK cells, but RNA-IC stimulation increased CD319 and CD229 expression. In conclusion, this study reveals that the expression of the SLAM receptors CD319 and CD229 is regulated on pDCs and NK cells by lupus ICs and that the expression of these receptors is specifically altered in SLE. These results, together with the observed genetic association between the SLAM locus and SLE, suggest a role for CD319 and CD229 in the SLE disease process.

The Past, Present, and Future of NK Cells in Hematopoietic Cell Transplantation and Adoptive Transfer

Hematopoietic cell transplantation (HCT) has been used as a part of cancer therapy for over half a decade. Beyond the necessity for donor-derived cells to reconstitute hematopoiesis after radiation and chemotherapy, immunologic reconstitution from allogeneic cells is important for the elimination of residual tumor cells. Natural killer (NK) cells are first among lymphocytes to reconstitute post-transplant and protect against cancer relapse. In this review, we provide a historical perspective on the role of NK cells in cancer control in the transplant setting and focus on current research aimed at improving NK cell responses for therapeutic benefit.