I-Motif Structures Formed in the Human c-MYC Promoter Are Highly Dynamic–Insights into Sequence Redundancy and I-Motif Stability

The GC-rich nuclease hypersensitivity element III1 (NHE III1) of the c-MYC promoter largely controls the transcriptional activity of the c-MYC oncogene. The C-rich strand in this region can form I-motif DNA secondary structures. We determined the folding pattern of the major I-motif formed in the NHE III1, which can be formed at near-neutral pH. While we find that the I-motif formed in the four 3′ consecutive runs of cytosines appears to be the most favored, our results demonstrate that the C-rich strand of the c-MYC NHE III1 exhibits a high degree of dynamic equilibration. Using a trisubstituted oligomer of this region, we determined the formation of two equilibrating loop isomers, one of which contains a flipped-out cytosine. Our results indicate that the intercalative cytosine+–cytosine base pairs are not always necessary for an intramolecular I-motif. The dynamic character of the c-MYC I-motif is intrinsic to the NHE III1 sequence and appears to provide stability to the c-MYC I-motif

High-Density SNP Mapping of the HLA Region Identifies Multiple Independent Susceptibility Loci Associated with Selective IgA Deficiency

Selective IgA deficiency (IgAD; serum IgA<0.07 g/l) is the most common form of human primary immune deficiency, affecting approximately 1∶600 individuals in populations of Northern European ancestry. The polygenic nature of IgAD is underscored by the recent identification of several new risk genes in a genome-wide association study. Among the characterized susceptibility loci, the association with specific HLA haplotypes represents the major genetic risk factor for IgAD. Despite the robust association, the nature and location of the causal variants in the HLA region remains unknown. To better characterize the association signal in this region, we performed a high-density SNP mapping of the HLA locus and imputed the genotypes of common HLA-B, -DRB1, and -DQB1 alleles in a combined sample of 772 IgAD patients and 1,976 matched controls from 3 independent European populations. We confirmed the complex nature of the association with the HLA locus, which is the result of multiple effects spanning the entire HLA region. The primary association signal mapped to the HLA-DQB1*02 allele in the HLA Class II region (combined P = 7.69×10−57; OR = 2.80) resulting from the combined independent effects of the HLA-B*0801-DRB1*0301-DQB1*02 and -DRB1*0701-DQB1*02 haplotypes, while additional secondary signals were associated with the DRB1*0102 (combined P = 5.86×10−17; OR = 4.28) and the DRB1*1501 (combined P = 2.24×10−35; OR = 0.13) alleles. Despite the strong population-specific frequencies of HLA alleles, we found a remarkable conservation of these effects regardless of the ethnic background, which supports the use of large multi-ethnic populations to characterize shared genetic association signals in the HLA region. We also provide evidence for the location of association signals within the specific extended haplotypes, which will guide future sequencing studies aimed at characterizing the precise functional variants contributing to disease pathogenesis

Immunologic Adverse Effects of Biologics for the Treatment of Atopy

The use of biologic agents as therapies for atopic diseases such as asthma and atopic dermatitis has increased greatly in recent years. The biological agents used to treat atopic diseases are for the most part monoclonal antibodies that suppress the immune response and reduce inflammation by targeting particular cytokines or other molecules involved in Th1, Th2, or Th17 immune reactions. Various side effects and rare complications have been reported from these agents. In this review, we discuss mechanisms of various adverse effects for the biologic agents currently in use or in development for atopic and inflammatory diseases. Monoclonal antibodies targeting the Th1 and Th17 pathways have been associated with significant side effects, partially due to their ability to cause significant impairment in immune responses to pathogens because of the immunologic alterations that they produce. Biologicals targeting Th2-mediated inflammation have had fewer reported side effects, though many are new and emerging drugs whose adverse effects may remain to be fully elucidated with more use. Therefore, continued long-term safety monitoring is required. As with all therapies, the risks associated with side effects of biologics must be balanced against the benefits these drugs offer for treating atopic diseases. One of the most apparent benefits is the steroid-sparing effect of well-chosen biologic therapy used to treat severe atopic disease. In contrast with the quite favorable safety profile of currently available biologics that target the Th2-mediated immune response, chronic systemic corticosteroid use is associated with significant side effects, many of which impact the majority of patients who are placed on long-term steroid therapy

Expression of 5-lipoxygenase (5-LOX) in T lymphocytes

5-lipoxygenase (5-LOX) is the key enzyme responsible for the synthesis of the biologically active leukotrienes. Its presence has been reported in cells of the myeloid lineage and B lymphocytes but has not been formally defined in T lymphocytes. In this study, we provide evidence for 5-LOX expression on both transcriptional and translational levels in highly purified peripheral blood T cells as well as in human T lymphoblastoid cell lines (MOLT4 and Jurkat). Messenger RNA (mRNA) of 5-LOX was amplified by conventional reverse transcription–polymerase chain reaction (RT-PCR; MOLT4 and Jurkat cells) and by in situ RT-PCR (T lymphocytes). 5-LOX protein expression was confirmed by Western blot and immunofluorescence studies. 5-LOX was present primarily in the cytoplasm with some nuclear localization and was translocated to the nuclear periphery after culture in a mitosis-supporting medium. Fluorescence-activated cell sorter analysis of different T-lymphocyte populations, including CD4, CD8, CD45RO, CD45RA, T helper type 2, and T-cell receptor-αβ and -γδ expressing cells, did not identify a differential distribution of the enzyme. Purified peripheral blood T lymphocytes were incapable of synthesizing leukotrienes in the absence of exogenous arachidonic acid. Jurkat cells produced leukotriene C4 and a small amount of leukotriene B4 in response to CD3–CD28 cross-linking. This synthesis was abolished by two inhibitors of leukotriene synthesis, MK-886 and AA-861. The presence of 5-LOX in T lymphocytes but the absence of endogenous lipoxygenase metabolite production compared to Jurkat cells may constitute a fundamental difference between resting peripheral lymphocytes and leukaemic cells