Enhanced humoral immune responses against T-independent antigens in Fc alpha/muR-deficient mice.

IgM is an antibody class common to all vertebrates that plays a primary role in host defenses against infection. Binding of IgM with an antigen initiates the complement cascade, accelerating cellular and humoral immune responses. However, the functional role of the Fc receptor for IgM in such immune responses remains obscure. Here we show that mice deficient in Fc alpha/muR, an Fc receptor for IgM expressed on B cells and follicular dendritic cells (FDCs), have enhanced germinal center formation and affinity maturation and memory induction of IgG3(+) B cells after immunization with T-independent (TI) antigens. Moreover, Fc alpha/muR-deficient mice show prolonged antigen retention by marginal zone B (MZB) cells and FDCs. In vitro studies demonstrate that interaction of the IgM immune complex with Fc alpha/muR partly suppress TI antigen retention by MZB cells. We further show that downregulation of complement receptor (CR)1 and CR2 or complement deprivation by in vivo injection with anti-CR1/2 antibody or cobra venom factor attenuates antigen retention by MZB cells and germinal center formation after immunization with TI antigens in Fc alpha/muR(-/-) mice. Taken together, these results suggest that Fc alpha/muR negatively regulates TI antigen retention by MZB cells and FDCs, leading to suppression of humoral immune responses against T-independent antigens.

Identification of the Fcalpha/muR isoform specifically expressed in the kidney tubules.

Fcalpha/muR is expressed not only in lymphoid, but also in non-lymphoid organs, including kidney. However, molecular and functional characteristics of Fcalpha/muR, particularly expressed in non-lymphoid organs, have remained unclear. Here we identified an isoform of Fcalpha/muR in the murine kidney on the C57BL/6J background. The kidney expressed only the isoform, which was not expressed in other lymphoid and non-lymphoid organs, and this isoform binds both IgA and IgM. Immunohistochemical analyses suggested that the Fcalpha/muR isoform was expressed in the tubular epithelial cells, but not in the glomeruli. This was confirmed by flow cytometry analysis of isolated tubular epithelial cells and by RT-PCR analyses using the separately excised glomerular and tubular regions by the laser microdissection system. These results suggest that Fcalpha/muR may not be involved in IgA deposition in glomerular mesangium cells in IgA nephropathy. Rather, it may play an important role in immunity in the renal tubular regions.

Isolation and characterization of naïve follicular dendritic cells.

Follicular dendritic cells (FDC) are specialized antigen-presenting cells to cognate B cells in the follicle of the lymphoid tissues. FDC also support survival and proliferation of the B cells, leading to the germinal center formation. FDC therefore play a central role in humoral immune responses. However, molecular and functional characteristics of FDC are largely unknown, because it is difficult to isolate and analyze FDC due to a very small number of FDC in the lymphoid tissues and the fragility by mechanical and chemical stresses in vitro. In this report, we established a novel method for FDC isolation from the spleen of naïve mice by flow cytometry and analyzed the phenotypical and functional characteristics. The isolated FDC, which accounted for ~0.2% of the spleen cells of naïve mice, were CD45(-), FDC-M2(+), and ICAM-1(+), and supported the survival and LPS-induced proliferation of B cells. We also showed that a neutralizing antibody against B cell activating factor TNF family (BAFF) suppressed FDC-dependent B cell proliferation in the presence of LPS, but not survival, demonstrating the evidence that FDC-derived BAFF is involved in B cell proliferation.

Requirement of the cytoplasmic portion for dimer formation of Fcalpha/micro receptor expressed on cell surface.

Fcalpha/mu receptor (Fcalpha/muR), an Fc receptor for IgA and IgM, is the only Fc receptor for IgM identified on hematopoietic cells in human and rodents and for IgA in rodents. Fcalpha/microR is a type 1 transmembrane protein containing one immunoglobulin-like domain in the extracellular portion. Both human and mouse Fcalpha/microR mediate endocytosis of the ligands IgA and IgM, for which the cytoplasmic portion of Fcalpha/microR is responsible. However, molecular characteristics of Fcalpha/muR involved in the function have been incompletely understood. Here, we show that both monomeric and dimeric Fcalpha/microR are expressed in a mouse B cell line BCL1-B20 and BW5147 or Ba/F3 transfectants stably expressing Fcalpha/microR. We also show that the dimeric, but not monomeric, Fcalpha/microR is preferentially localized to the cell surface of the transfectants. BW5147 transfectant expressing mutant Fcalpha/microR lacking the cytoplasmic portion expressed only the monomeric Fcalpha/microR. These results suggest that the cytoplasmic portion is required for the dimer formation and thus for efficient cell surface expression of Fcalpha/microR.

Molecular characteristics of IgA and IgM Fc binding to the Fcalpha/muR.

Fcalpha/mu receptor (Fcalpha/muR), a novel Fc receptor for IgA and IgM, is a type I transmembrane protein with an immunoglobulin (Ig)-like domain in the extracellular portion. Although IgA and IgM bind to Fcalpha/muR, the molecular and structural characteristics of the ligand-receptor interactions have been undetermined. Here, we developed twelve monoclonal antibodies (mAbs) against murine Fcalpha/muR by immunizing mice deficient in Fcalpha/muR gene. Eight mAbs totally or partially blocked IgA and IgM bindings to Fcalpha/muR. These blocking mAbs bound to a peptide derived from the Ig-like domain of murine Fcalpha/muR, which is conserved not only in human and rat Fcalpha/muR but also in polymeric Ig receptor (poly-IgR), another Fc receptor for IgA and IgM. These results suggest that IgA and IgM bind to an epitope in the conserved amino acids in the Ig-like domain of Fcalpha/muR as well as poly-IgR.