Innate immunity enables an organism to respond rapidly to invading microorganisms. To do this, innate immune system uses a variety of receptors or intracellular proteins that can recognize a microbial pathogen by the motifs pathogen-associated molecular patterns (PAMPs). Pathogen-associated motifs include mannans in the yeast cell wall, formylated peptides and various bacterial cell-wall components such as lipopolysaccharide (LPS), lipopeptides, peptidoglycans and teichoic acids. Bacterial DNA is also a potent inducer of innate immunity. Bacterial DNA contains increased frequency of CpG dinucleotide motifs in particular sequence contexts (PuPuCpGPyPy), which are greatly reduced in frequency in the mammalian genome, and where it occurs, the cytosine commonly is methylated. DNA containing unmethylated CpG motifs is immunostimulatory and able to activate cellular component of innate immunity, including macrophages and dendritic cells (DC) as well as B cells. Thus, DNA containing unmethylated CpG motif in sequences of PuPuCpGPyPy is also called ISS-DNA. Activation of immune cells by ISS-DNA appears to require uptake into the cells by receptor- or pinocytosis-mediated endocytosis and endosome acidification, suggesting that the recognition structure is either endosome-associated or cytoplasmic. Recent studies suggested that Toll-like receptor (TLR) 9 and adapter protein MyD88 are required for ISS-DNA to induce immune cells to produce interlukin-12, and for activation of NF-kB by ISS-ODN. The studies from our group suggested that catalytic subunit of DNA-dependent protein kinase (DNAPKcs) and IkB kinase (IKK) are required for the activation of innate immunity by ISS-DNA. ISS-DNA activates DNA-PKcs, which in turn activates IKK, leading to NF-kB activation. Null mutations in either the DNA-PKcs- or IKKb gene were found to selectively impair the ability of mice or their macrophages, to respond to ISS-DNA with inducible cytokine gene expression. TLR9 is on cell membrane while DNA-PKcs is in cytoplasm and nucleus. The connection between TLR9 and DNA-PKcs is not apparent. To further understand the molecular mechanisms of activation of innate immunity by ISS-DNA, we will investigate the relationship between TLR9-signaling pathway and DNA-PK by using biochemical and genetic means. Moreover, we will investigate if the regulatory subunits of DNA-PK, Ku70 and Ku80, and their associated factors are required for the innate response to ISS-DNA. Furthermore, we will identify new molecules that are required for activation of innate immunity by ISS-DNA.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Special Emphasis Panel (ZRG1-SSS-F (01))
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Winter, David B
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Brown University
Schools of Medicine
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