During human CMV infection, there is a preferential expansion of natural killer (NK) cells expressing the activating CD94-NKG2C receptor complex, implicating this receptor in the recognition of CMV-infected cells. We hypothesized that NK cells expanded in response to pathogens will be marked by expression of CD57, a carbohydrate antigen expressed on highly mature cells within the CD56

During mouse cytomegalovirus (CMV) infection, a population of Ly49H ؉ natural killer (NK) cells expands and is responsible for disease clearance through the induction of a "memory NK-cell response." Whether similar events occur in human CMV infection is unknown. In the present study, we characterized the kinetics of the NK-cell response to CMV reactivation in human recipients after hematopoietic cell transplantation. During acute infection, NKG2C ؉ NK cells expanded and were potent producers of IFN␥. NKG2C ؉ NK cells predominately expressed killer cell immunoglobulin-like receptor, and self-killer cell immunoglobulinlike receptors were required for robust IFN␥ production. During the first year after transplantation, CMV reactivation induced a more mature phenotype characterized by an increase in CD56 dim NK cells. Strikingly, increased frequencies of NKG2C ؉ NK cells persisted and continued to increase in recipients who reactivated CMV, whereas these cells remained at low frequency in recipients without CMV reactivation. Persisting NKG2C ؉ NK cells lacked NKG2A, expressed CD158b, preferentially acquired CD57, and were potent producers of IFN␥ during the first year after transplantation. Recipients who reactivated CMV also expressed higher amounts of IFN␥, T-bet, and IL-15R␣ mRNA transcripts. Our findings support the emerging concept that CMV-induced innate memory-cell populations may contribute to malignant disease relapse protection and infectious disease control long after transplantation. (Blood. 2012; 119(11):2665-2674) IntroductionNatural killer (NK) cells are important effectors during the host innate immune response to viral infections. They can recognize and eliminate virally infected cells, interact with dendritic cells, and produce a range of cytokines and chemokines (eg, IFN␥, TNF-␣, MIP-1␣, MIP-1␤, and RANTES) that recruit and modulate the adaptive immune response.Under normal homeostatic conditions, a balance of activating and inhibitory signals tightly controls NK-cell function. 1,2 The best-characterized inhibitory receptors are the inhibitory killer cell immunoglobulin-like receptors (KIRs) that recognize allelic epitopes present on certain class I human leukocyte antigens (HLAs) and the C-type lectin-like receptor NKG2A, which recognizes the nonclassic class I HLA, HLA-E. 3,4 Activating signals are mediated by receptor families, including activating KIR, NKG2C, NKG2D, the natural cytotoxicity receptors (NKp30, NKp44, and NKp46), CD16, and CD244. 1 When self HLA is down-regulated, cells are susceptible to NK-cell lysis because of the lack of ligands for the inhibitory receptors, a phenomenon known as the "missing self" hypothesis. 5,6 Human cytomegalovirus (CMV), a member of the Herpesviridae family, causes asymptomatic or mild illness in healthy people. 7 CMV remains latent in infected hosts and, by adulthood, approximately 60% of people in the United States are seropositive for CMV. 8 However, for patients immunosuppressed due to HIV infection or solid organ or hematopoietic call transplantatio...

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IntroductionNatural killer (NK) cells comprise 5% to 20% of human peripheral blood lymphoid cells and are a critical component of the immune system, providing protection against viral infections and contributing to tumor immune surveillance. NK-cell activity is regulated by an intricate balance of signals transmitted by inhibitory and activating receptors. 1,2 Functionally distinct NK-cell subsets can be defined based on the level of CD56 and CD16 coexpression. 3 CD56 bright CD16 Ϫ NK cells produce abundant IFN-␥ in response to stimulation with interleukin (IL)-12 and proliferate robustly when cultured in IL-2, whereas CD56 dim CD16 ϩ NK cells are more cytolytic and produce significant amounts of cytokine when their activating receptors are engaged. 4 CD56 dim CD16 ϩ NK cells are considered mature NK cells and are differentiated from the immature CD56 bright CD16 -NK-cell subset. This is further supported by recent data demonstrating the dynamics of expression of the killer immunoglobulin-like receptors (KIR), CD57, CD94, and CD62L expression on the CD56 dim CD16 ϩ NK cells as they mature from CD56 bright CD16 -NK-cell precursors. [5][6][7][8][9] T-cell immunoglobulin-and mucin domain-containing (Tim)-3 is a member of Tim family of receptors of which there are 3 in humans (Tim-1, Tim-3, and Tim-4). 10 These molecules are involved in diverse metabolic and immunoregulatory processes. 11 Tim-3 is a type I transmembrane protein that contains no defined signaling motifs in its cytoplasmic domain, but it has been implicated both in activation and inhibition of immune responses 12,13 and in the induction of apoptosis of Tim-3-bearing cells through interactions with galectin-9. 14 Tim-3 is expressed on CD4 ϩ T cells, dendritic cells, monocytes, 15-17 CD8 ϩ T cells, 18,19 and NK cells. 20 In a comparison of lymphocyte populations in healthy human subjects, the highest transcription of the gene encoding Tim-3 was observed in NK cells. 21 There is evidence that engagement of Tim-3 on mouse T cells with the ligand galectin-9 promotes aggregation, leading to the death of T-helper 1 cells and the selective loss of interferon (IFN)-␥-producing T cells. 14 On human T cells, the expression of Tim-3 regulates cell proliferation and IFN-␥ secretion. 19,21,22 We and others have observed that increased amounts of Tim-3 on T cells during HIV, hepatitis C virus, and other chronic viral infections correlated with T-cell dysfunction, suggesting that Tim-3 is part of a negative regulatory pathway. 19,[23][24][25] In this study, we investigated the expression of Tim-3 on human NK cells and its regulation by cytokines, and we provide evidence for the role of Tim-3 in the restraint of NK cell-mediated cytotoxicity in healthy individuals. Methods Primary cells and cell linesPeripheral blood mononuclear cells (PBMCs) of healthy individuals were obtained from the Stanford Blood Bank. Cord blood PMBCs were obtained The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, ...

The presence of the envelope glycoprotein Env in HIV-1 virions is essential for infectivity. To date, the molecular mechanism by which Env is packaged into virions has been largely unknown. Here, we show that TIP47 (tail-interacting protein of 47 kDa), which has been shown to interact with Env, also binds the MA (matrix) domain of HIV-1 Gag protein and that these three proteins form a ternary complex. Mutations in Gag that abrogate interaction with TIP47 inhibit Env incorporation and virion infectivity as well as colocalization between Gag and Env. We also show that TIP47 silencing impairs Env incorporation and infectivity and abolishes coimmunoprecipitation of Gag with Env. In contrast, overexpression of TIP47 increases Env packaging. Last, we demonstrate that TIP47 can interact simultaneously with Env and Gag. Taken together, our results show that TIP47 is a cellular cofactor that plays an essential role in Env incorporation, allowing the encounter and the physical association between HIV-1 Gag and Env proteins during the viral assembly process.envelope incorporation ͉ HIV-1 envelope glycoprotein ͉ matrix domain ͉ HIV-1 assembly ͉ Gag precursor T he incorporation of the HIV-1 envelope glycoprotein Env into virions confers infectivity to HIV-1 particles, making it a key step in the viral life cycle. Env is synthesized as a precursor, gp160, which is cleaved to give the surface subunit SUgp120 and the transmembrane subunit TMgp41. This SUgp120͞TMgp41 complex is then incorporated into budding virions. SUgp120 binds to the CD4 receptor of cells and then to the coreceptor, initiating HIV-1 entry, whereas TMgp41 induces the membrane fusion between viral and cellular lipid bilayers (1, 2).Recent studies have elucidated most of the molecular mechanisms involved in the budding of HIV and other retroviruses (3). However, the mechanisms that govern the incorporation of Env into viral particles, conferring infectivity to these particles, remain largely unknown. Many studies suggest that the cytoplasmic domain (CD) of TMgp41 (TMgp41 CD) and the MA (matrix) domain of the Gag protein, a protein that is closely associated with the virion membrane, together with putative cellular cofactors, play a central role in this mechanism. Two models have been suggested for HIV-1 Env packaging. The first one, the ''active packaging'' model, proposes that a specific interaction occurs between Gag and Env proteins. This model is based on the observation that many deletions and substitutions in the MA domain and in the TMgp41 CD abolish the packaging of Env into Gag particles, whereas particle release remains unaffected (4-17). This model is also supported by an in vitro study that showed direct binding between HIV-1 MA and TMgp41 CD peptides (18) and by the fact that HIV-1 Env functions to drive basolateral budding of Gag in polarized cells (19). In the second model, the ''passive packaging'' model, Env is thought to be passively incorporated into nascent virions at the plasma membrane. HIV-1 Gag particles are able to accommodate heterolog...

Immunological memory is a hallmark of the adaptive immune system. However, the ability to “remember” and respond more robustly against a second encounter with the same pathogen has been described in organisms lacking T and B cells. Recently, natural killer (NK) cells have been shown to mediate antigen-specific recall responses in several different model systems. Although NK cells do not rearrange the genes encoding their activating receptors, NK cells experience a selective education process during development, undergo a clonal-like expansion during virus infection, generate long-lived progeny (i.e. memory cells), and mediate more efficacious secondary responses against previously encountered pathogen – all characteristics previously ascribed only to T and B cells in mammals. This review describes past findings leading up to these new discoveries, summarizes the evidence for and characteristics of NK cell memory, and discusses the attempts and future challenges to identify these long-lived memory NK cell populations in humans.

BACKGROUND The eastern equine encephalitis (EEE) and Venezuelan equine encephalitis (VEE) viruses are pathogens that infect humans and horses in the Americas. Outbreaks of neurologic disease in humans and horses were reported in Panama from May through early August 2010. METHODS We performed antibody assays and tests to detect viral RNA and isolate the viruses in serum samples from hospitalized patients. Additional cases were identified with enhanced surveillance. RESULTS A total of 19 patients were hospitalized for encephalitis. Among them, 7 had confirmed EEE, 3 had VEE, and 1 was infected with both viruses; 3 patients died, 1 of whom had confirmed VEE. The clinical findings for patients with EEE included brain lesions, seizures that evolved to status epilepticus, and neurologic sequelae. An additional 99 suspected or probable cases of alphavirus infection were detected during active surveillance. In total, 13 cases were confirmed as EEE, along with 11 cases of VEE and 1 case of dual infection. A total of 50 cases in horses were confirmed as EEE and 8 as VEE; mixed etiologic factors were associated with 11 cases in horses. Phylogenetic analyses of isolates from 2 cases of equine infection with the EEE virus and 1 case of human infection with the VEE virus indicated that the viruses were of enzootic lineages previously identified in Panama rather than new introductions. CONCLUSIONS Cases of EEE in humans in Latin America may be the result of ecologic changes that increased human contact with enzootic transmission cycles, genetic changes in EEE viral strains that resulted in increased human virulence, or an altered host range. (Funded by the National Institutes of Health and the Secretaría Nacional de Ciencia, Tecnología e Innovación, Panama.)

Antitumor T cells are subject to multiple mechanisms of negative regulation1–3. Recent findings that innate lymphoid cells (ILCs) regulate adaptive T cell responses4–6 led us to examine the regulatory potential of ILCs in the context of cancer. We identified a unique ILC population that inhibits tumor-infiltrating lymphocytes (TILs) from high-grade serous tumors, defined their suppressive capacity in vitro, and performed a comprehensive analysis of their phenotype. Notably, the presence of this CD56+CD3− population in TIL cultures was associated with reduced T cell numbers, and further functional studies demonstrated that this population suppressed TIL expansion and altered TIL cytokine production. Transcriptome analysis and phenotypic characterization determined that regulatory CD56+CD3− cells exhibit low cytotoxic activity, produce IL-22, and have an expression profile that overlaps with those of natural killer (NK) cells and other ILCs. NKp46 was highly expressed by these cells, and addition of anti-NKp46 antibodies to TIL cultures abrogated the ability of these regulatory ILCs to suppress T cell expansion. Notably, the presence of these regulatory ILCs in TIL cultures corresponded with a striking reduction in the time to disease recurrence. These studies demonstrate that a previously uncharacterized ILC population regulates the activity and expansion of tumor-associated T cells.