Human NK cell deficiency as a result of biallelic mutations in MCM10.

Human natural killer cell deficiency (NKD) arises from inborn errors of immunity that lead to impaired NK cell development, function, or both. Through the understanding of the biological perturbations in individuals with NKD, requirements for the generation of terminally mature functional innate effector cells can be elucidated. Here, we report a cause of NKD resulting from compound heterozygous mutations in minichromosomal maintenance complex member 10 (MCM10) that impaired NK cell maturation in a child with fatal susceptibility to CMV. MCM10 has not been previously associated with monogenic disease and plays a critical role in the activation and function of the eukaryotic DNA replisome. Through evaluation of patient primary fibroblasts, modeling patient mutations in fibroblast cell lines, and MCM10 knockdown in human NK cell lines, we have shown that loss of MCM10 function leads to impaired cell cycle progression and induction of DNA damage-response pathways. By modeling MCM10 deficiency in primary NK cell precursors, including patient-derived induced pluripotent stem cells, we further demonstrated that MCM10 is required for NK cell terminal maturation and acquisition of immunological system function. Together, these data define MCM10 as an NKD gene and provide biological insight into the requirement for the DNA replisome in human NK cell maturation and function.

Comparison of the Performance of Skin Prick, ImmunoCAP, and ISAC Tests in the Diagnosis of Patients with Allergy.

BACKGROUND: Allergy is diagnosed from typical symptoms, and tests are performed to incriminate the suspected precipitant. Skin prick tests (SPTs) are commonly performed, inexpensive, and give immediate results. Laboratory tests (ImmunoCAP) for serum allergen-specific IgE antibodies are usually performed more selectively. The immuno-solid phase allergen chip (ISAC) enables testing for specific IgE against multiple allergen components in a multiplex assay. METHODS: We retrospectively analysed clinic letters, case notes, and laboratory results of 118 patients attending the National Adult Allergy Service at the University Hospital of Wales who presented diagnostic difficulty, to evaluate which testing strategy (SPT, ImmunoCAP, or ISAC) was the most appropriate to use to confirm the diagnosis in these complex patients, evaluated in a "real-life" clinical service setting. RESULTS: In patients with nut allergy, the detection rates of SPTs (56%) and ISAC (65%) were lower than those of ImmunoCAP (71%). ISAC had a higher detection rate (88%) than ImmunoCAP (69%) or SPT (33%) in the diagnosis of oral allergy syndrome. ImmunoCAP test results identified all 9 patients with anaphylaxis due to wheat allergy (100%), whereas ISAC was positive in only 6 of these 9 (67%). CONCLUSIONS: In this difficult diagnostic group, the ImmunoCAP test should be the preferred single test for possible allergy to nuts, wheat, other specific foods, and anaphylaxis of any cause. In these conditions, SPT and ISAC tests give comparable results. The most useful single test for oral allergy syndrome is ISAC, and SPT should be the preferred test for latex allergy.

Early acquisition of cytolytic function and transcriptional changes in a primary CD8+ T-cell response in vivo.

Functional studies show that programming of CD8+ T cells occurs early after initial antigen encounter within as little as 2 hours. To define the molecular basis of these events, we transferred TCR transgenic T cells from F5 Rag-/- mice into naive recipients and stimulated them with recombinant vaccinia expressing the immunodominant influenza epitope NP366-374. Transcription in epitope-specific cytotoxic T lymphocytes (CTLs) was analyzed using Affymetrix 430 2.0 GeneChips and quantitative polymerase chain reaction (PCR). We demonstrated an early transcriptional burst with the greatest number of genes reaching peak expression 12 hours after stimulation. Using in vivo cytotoxicity assays we demonstrated that early up-regulation of cytolytic genes was accompanied by acquisition of killing capacity within 24 hours of stimulation. However, T-cell proliferation was not observed until 48 hours. We therefore conclude that clonal expansion rather than acquisition of effector function is the rate-limiting step in the development of a primary CTL response.

The role of CTLs in persistent viral infection: cytolytic gene expression in CD8+ lymphocytes distinguishes between individuals with a high or low proviral load of human T cell lymphotropic virus type 1.

The proviral load in human T cell lymphotropic virus type 1 (HTLV-1) infection is typically constant in each infected host, but varies by >1000-fold between hosts and is strongly correlated with the risk of HTLV-1-associated inflammatory disease. However, the factors that determine an individual's HTLV-1 proviral load remain uncertain. Experimental evidence from studies of host genetics, viral genetics, and lymphocyte function and theoretical considerations suggest that a major determinant of the equilibrium proviral load is the CD8+ T cell response to HTLV-1. In this study, we tested the hypothesis that the gene expression profile in circulating CD8+ and CD4+ lymphocytes distinguishes between individuals with a low proviral load of HTLV-1 and those with a high proviral load. We show that circulating CD8+ lymphocytes from individuals with a low HTLV-1 proviral load overexpressed a core group of nine genes with strong functional coherence: eight of the nine genes encode granzymes or other proteins involved in cell-mediated lysis or Ag recognition. We conclude that successful suppression of the HTLV-1 proviral load is associated with strong cytotoxic CD8+ lymphocyte activity in the peripheral blood.