Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is typically very mild and often asymptomatic in children. A complication is the rare multisystem inflammatory syndrome in children (MIS-C) associated with COVID-19, presenting 4–6 weeks after infection as high fever, organ dysfunction, and strongly elevated markers of inflammation. The pathogenesis is unclear but has overlapping features with Kawasaki disease suggestive of vasculitis and a likely autoimmune etiology. We apply systems-level analyses of blood immune cells, cytokines, and autoantibodies in healthy children, children with Kawasaki disease enrolled prior to COVID-19, children infected with SARS-CoV-2, and children presenting with MIS-C. We find that the inflammatory response in MIS-C differs from the cytokine storm of severe acute COVID-19, shares several features with Kawasaki disease, but also differs from this condition with respect to T cell subsets, interleukin (IL)-17A, and biomarkers associated with arterial damage. Finally, autoantibody profiling suggests multiple autoantibodies that could be involved in the pathogenesis of MIS-C.

SUMMARY 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 hyperme-thylation. 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.

IntroductionNatural killer (NK) cells are characterized by cytolytic activity against susceptible target cells and by the secretion of cytokines, such as tumor necrosis factor ␣ (TNF-␣) and interferon ␥ (IFN-␥). NK cells discriminate between normal and abnormal cells (infected or transformed) through engagement and dynamic integration of multiple signaling pathways, which are initiated by germline-encoded receptors. [1][2][3] Healthy cells are protected from NK-cell-mediated lysis by expression of major histocompatibility complex (MHC) class I ligands for NK-cellinhibitory receptors. 1,4 However, de novo expression of ligands for NK-cell activation receptor NKG2D can trigger natural cytotoxicity against MHC class I ϩ target cells. 5,6 A number of structurally distinct receptors have been implicated in activation of NK-cell effector functions. It is not yet clear if any one receptor is necessary or sufficient to activate NK cells and to what extent activation receptors may be redundant. Activation receptors can be grouped in 3 categories: receptors that signal through immunoreceptor tyrosine-based activation motif (ITAM)-containing subunits (eg CD16, NKp46, NKp44), the DAP10-associated receptor NKG2D, and several other receptors (eg CD2, 2B4, DNAM-1) that signal by different pathways.CD16 (Fc␥RIII), a low-affinity receptor for IgG, is associated with the ITAM-containing Fc⑀RI ␥ chain and T-cell receptor (TCR) chain. NKp46 and NKp30 are associated with the TCR chain. 7 NKp44, KIR2DS, and CD94/NKG2C are associated with the ITAM-containing DAP12. Natural cytotoxicity receptors (NCRs), which include NKp46, NKp44, and NKp30, play a major role in NK-cell cytotoxicity against transformed cells. 8 Although ligands of NCRs have not been identified, antibodies against NCRs have been used to block lysis of tumor cells by interleukin 2 (IL-2)-activated and resting NK cells. [9][10][11] However, in the mouse, Syk/ ZAP70-independent natural cytotoxicity by NK cells was observed, implying that natural cytotoxicity can occur independently of ITAM-based activation signals. 12,13 NKG2D can signal through both DAP10 and DAP12 in mice, 14,15 whereas human NKG2D associates only with DAP10. [16][17][18] DAP10 is a signaling subunit that carries a phosphatidylinositol-3 kinase-binding motif. 16 Ligands for NKG2D, such as MICA and ULBP, are expressed on some tumor cells, and on infected or stressed cells. 19 Experiments have suggested that NKG2D signals are sufficient to activate NK-cell functions. [20][21][22] Lysis of certain tumor cells by resting NK cells and by IL-2-activated NK cells can be blocked by antibodies to NKG2D. 11,23 The importance of ligands for NKG2D in immune defense is underscored by strategies developed by viruses to interfere with their expression. 19,24,25 Several other receptors activate NK cells by signaling through their own cytoplasmic tail. 2B4 (CD244) recruits SAP and Fyn through cytoplasmic tyrosine-based motifs. 26,27 The ligand of 2B4 is CD48, which is expressed on hematopoietic cells. 28 CD2 and N...