ABSTRACT
Coronin 1 is a leukocyte specific regulator of Ca(2+)-dependent signaling and is essential for the survival of peripheral T lymphocytes, but its role in B cells is unknown. In this study, we show that coronin 1 is essential for intracellular Ca(2+) mobilization and proliferation upon triggering of the BCR. However, the presence of costimulatory signals rendered coronin 1 dispensable for B cell signaling, consistent with the generation of normal immune responses against a variety of Ags in coronin 1-deficient mice. We conclude that coronin 1, while being essential for T cell function and survival, is dispensable for B cell function in vivo.
Subject(s)
B-Lymphocytes/immunology , Calcium Signaling/immunology , Calcium/metabolism , Immunoglobulin M/immunology , Microfilament Proteins/immunology , Animals , Blotting, Western , Enzyme-Linked Immunosorbent Assay , Flow Cytometry , Fluorescent Antibody Technique , Immunohistochemistry , Mice , Mice, Transgenic , Microfilament Proteins/metabolism , Microscopy, ConfocalABSTRACT
T cell homeostasis is essential for the functioning of the vertebrate immune system, but the intracellular signals required for T cell homeostasis are largely unknown. We here report that the WD-repeat protein family member coronin-1, encoded by the gene Coro1a, is essential in the mouse for T cell survival through its promotion of Ca2+ mobilization from intracellular stores. Upon T cell receptor triggering, coronin-1 was essential for the generation of inositol-1,4,5-trisphosphate from phosphatidylinositol-4,5-bisphosphate. The absence of coronin-1, although it did not affect T cell development, resulted in a profound defect in Ca2+ mobilization, interleukin-2 production, T cell proliferation and T cell survival. We conclude that coronin-1, through activation of Ca2+ release from intracellular stores, is an essential regulator of peripheral lymphocyte survival.
Subject(s)
Calcium/metabolism , Inositol 1,4,5-Trisphosphate/biosynthesis , Microfilament Proteins/physiology , Receptors, Antigen, T-Cell/metabolism , T-Lymphocytes/immunology , Actins/metabolism , Animals , Calcium Signaling/genetics , Calcium Signaling/physiology , Cell Survival/genetics , Cell Survival/immunology , Intracellular Fluid/immunology , Intracellular Fluid/metabolism , Lymphocyte Activation/genetics , Lymphocyte Activation/immunology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , Microfilament Proteins/deficiency , Microfilament Proteins/genetics , Receptors, Antigen, T-Cell/physiology , T-Lymphocytes/metabolism , Type C Phospholipases/metabolismABSTRACT
Macrophages are crucial for innate immunity, apoptosis, and tissue remodeling, processes that rely on the capacity of macrophages to internalize and process cargo through phagocytosis. Coronin 1, a member of the WD repeat protein family of coronins specifically expressed in leukocytes, was originally identified as a molecule that is recruited to mycobacterial phagosomes and prevents the delivery of mycobacteria to lysosomes, allowing these to survive within phagosomes. However, a role for coronin 1 in mycobacterial pathogenesis has been disputed in favor for its role in mediating phagocytosis and cell motility. In this study, a role for coronin 1 in actin-mediated cellular processes was addressed using RNA interference in the murine macrophage cell line J774. It is shown that the absence of coronin 1 in J774 macrophages expressing small interfering RNA constructs specific for coronin 1 does not affect phagocytosis, macropinocytosis, cell locomotion, or regulation of NADPH oxidase activity. However, in coronin 1-negative J774 cells, internalized mycobacteria were rapidly transferred to lysosomes and killed. Therefore, these results show that in J774 cells coronin 1 has a specific role in modulating phagosome-lysosome transport upon mycobacterial infection and that it is dispensable for most F-actin-mediated cytoskeletal rearrangements.
Subject(s)
Actins/metabolism , Macrophages/cytology , Macrophages/microbiology , Microfilament Proteins/metabolism , Mycobacterium/physiology , RNA Interference , Animals , Cell Line , Chemotaxis/drug effects , Clone Cells , Epidermal Growth Factor/pharmacology , Erythrocytes/cytology , Erythrocytes/drug effects , Gene Expression Regulation/drug effects , Macrophage Activation/drug effects , Macrophages/drug effects , Macrophages/enzymology , Mice , Microbial Viability/drug effects , Microfilament Proteins/genetics , Mycobacterium/cytology , Mycobacterium/drug effects , NADPH Oxidases/metabolism , Phagocytosis/drug effects , Pinocytosis/drug effects , Protein Transport/drug effects , Pseudopodia/drug effects , Pseudopodia/metabolism , RNA Interference/drug effects , RNA, Small Interfering/metabolism , SheepABSTRACT
Pathogenic mycobacteria survive within macrophages by avoiding lysosomal delivery, instead residing in mycobacterial phagosomes. Upon infection, the leukocyte-specific protein coronin 1 is actively recruited to mycobacterial phagosomes, where it blocks lysosomal delivery by an unknown mechanism. Analysis of macrophages from coronin 1-deficient mice showed that coronin 1 is dispensable for F-actin-dependent processes such as phagocytosis, motility, and membrane ruffling. However, upon mycobacterial infection, coronin 1 was required for activation of the Ca(2+)-dependent phosphatase calcineurin, thereby blocking lysosomal delivery of mycobacteria. In the absence of coronin 1, calcineurin activity did not occur, resulting in lysosomal delivery and killing of mycobacteria. Furthermore, blocking calcineurin activation with cyclosporin A or FK506 led to lysosomal delivery and intracellular mycobacterial killing. These results demonstrate a role for coronin 1 in activating Ca(2+) dependent signaling processes in macrophages and reveal a function for calcineurin in the regulation of phagosome-lysosome fusion upon mycobacterial infection.
