ABSTRACT
Plasmacytoid dendritic cells (pDC) are essential for immune competence. Here we show that pDC precursor differentiated from human CD34+ hematopoietic stem and progenitor cells (HSPC) has low surface expression of pDC markers, and has limited induction of type I interferon (IFN) and IL-6 upon TLR7 and TLR9 agonists treatment; by contrast, cGAS or RIG-I agonists-mediated activation is not altered. Importantly, after priming with type I and II IFN, these precursor pDCs attain a phenotype and functional activity similar to that of peripheral blood-derived pDCs. Data from CRISPR/Cas9-mediated genome editing of HSPCs further show that HSPC-pDCs with genetic modifications can be obtained, and that expression of the IFN-α receptor is essential for the optimal function, but dispensable for the differentiation, of HSPC-pDC percursor. Our results thus demonstrate the biological effects of IFNs for regulating pDC function, and provide the means of generating of gene-modified human pDCs.
Subject(s)
Antigens, CD34/metabolism , Dendritic Cells/metabolism , CRISPR-Cas Systems/genetics , Cell Differentiation/genetics , Cell Differentiation/physiology , Cells, Cultured , DEAD Box Protein 58/metabolism , Enzyme-Linked Immunosorbent Assay , Gene Editing , Humans , Interferon Type I/metabolism , Interleukin-6/metabolism , Nucleotidyltransferases/metabolism , Polymerase Chain Reaction , Receptor, Interferon alpha-beta/genetics , Receptor, Interferon alpha-beta/metabolism , Receptors, Immunologic , Toll-Like Receptor 7/agonists , Toll-Like Receptor 9/agonistsABSTRACT
Innate immune activation by macrophages is an essential part of host defence against infection. Cytosolic recognition of microbial DNA in macrophages leads to induction of interferons and cytokines through activation of cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING). Other host factors, including interferon-gamma inducible factor 16 (IFI16), have been proposed to contribute to immune activation by DNA. However, their relation to the cGAS-STING pathway is not clear. Here, we show that IFI16 functions in the cGAS-STING pathway on two distinct levels. Depletion of IFI16 in macrophages impairs cGAMP production on DNA stimulation, whereas overexpression of IFI16 amplifies the function of cGAS. Furthermore, IFI16 is vital for the downstream signalling stimulated by cGAMP, facilitating recruitment and activation of TANK-binding kinase 1 in STING complex. Collectively, our results suggest that IFI16 is essential for efficient sensing and signalling upon DNA challenge in macrophages to promote interferons and antiviral responses.
Subject(s)
DNA/metabolism , Macrophages/metabolism , Nuclear Proteins/metabolism , Nucleotides, Cyclic/metabolism , Phosphoproteins/metabolism , Cells, Cultured , Gene Expression Profiling , HEK293 Cells , Humans , Immunity, Innate/genetics , Interferons/immunology , Interferons/metabolism , Macrophages/immunology , Macrophages/virology , Membrane Proteins/genetics , Membrane Proteins/metabolism , Mutation , Nuclear Proteins/genetics , Nucleotidyltransferases/genetics , Nucleotidyltransferases/metabolism , Phosphoproteins/genetics , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , RNA Interference , Signal Transduction/genetics , THP-1 CellsABSTRACT
The startup of a wet retention pond designed for extended stormwater treatment was monitored by more than one year of continual measurement of hydraulic parameters, nutrients and quality parameters in the pond itself (pH, temperature, dissolved oxygen, turbidity). The data revealed that photosynthesis played an important role for dissolved oxygen and pH for most of the year. Another important observation was that the pond behaved more like a completely mixed reactor than like a plug flow reactor--even though the length to width ratio was as high as 4.5:1. The pond was equipped with sand filters and sorption filters whereby very good nutrient removal efficiencies were achieved.
