IgA Receptor Research Articles

C‐Jun activating binding protein 1 binds to the IgA receptor and modulates protein levels of FcαRI and FcRγ‐chain

The receptor for IgA, FcalphaRI or CD89, is expressed on myeloid cells and can trigger phagocytosis, tumor cell lysis, and release of inflammatory mediators. These functions critically depend on the associated FcR gamma-chain; however, some biological functions, like receptor internalization, are solely mediated by FcalphaRI alpha-chain. Little is known as to how FcalphaRI regulates these processes and the FcalphaRI intracellular domain does not contain recognized signalling motifs. We searched for associating proteins and identified c-Jun activating binding protein 1 (JAB1) as a binding partner specifically for FcalphaRI. We found increased FcalphaRI surface expression after ectopic expression of JAB1 as well as diminished protein levels of total FcR gamma-chain levels after JAB1 knock-down. These data functionally link JAB1 with controlling protein expression levels of FcalphaRI-FcR gamma-chain protein complex.

MAL2 Selectively Regulates Polymeric IgA Receptor Delivery from the Golgi to the Plasma Membrane in WIF-B Cells

Myelin and lymphocyte protein 2 (MAL2) has been identified as a hepatic transcytotic regulator that mediates delivery from basolateral endosomes to the subapical compartment (SAC). However, overexpression of polymeric immunoglobulin A-receptor (pIgA-R) in polarized, hepatic WIF-B cells led to the dramatic redistribution of MAL2 into the Golgi and all the transcytotic intermediates occupied by the receptor. Although overexpressed hemagglutinin and dipeptidylpeptidase IV (DPPIV) distributed to the same compartments, MAL2 distributions did not change indicating the effect is selective. Cycloheximide treatment led to decreased pIgA-R and MAL2 intracellular staining, first in the Golgi then the SAC, suggesting they were apically delivered and that MAL2 was mediating the process. This was tested in Clone 9 cells (that lack endogenous MAL2). When expressed alone, pIgA-R was restricted to the Golgi whereas when coexpressed with MAL2, it distributed to the surface, was internalized and delivered to MAL2-positive puncta. In contrast, DPPIV distributions were independent of MAL2. Surface delivery of newly synthesized pIgA-R, but not DPPIV, was enhanced greater than ninefold by MAL2 coexpression. In WIF-B cells where MAL2 expression was knocked down, pIgA-R, but not DPPIV, was retained in the Golgi and its basolateral delivery was impaired. Thus, in addition to its role in transcytosis, MAL2 also regulates pIgA-R delivery from the Golgi to the plasma membrane.

Requirement of the cytoplasmic portion for dimer formation of Fcα/μ receptor expressed on cell surface

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

Endocytosis of FcαR is clathrin and dynamin dependent, but its cytoplasmic domain is not required

FcalphaR, the Fc receptor for IgA, is essential for IgA-mediated immune responses. Previous studies have shown that IgA and IgA immune complexes can be rapidly endocytosed by FcalphaR. However, the underlying mechanism remains unclear. Here, we investigated the endocytic pathway of FcalphaR in monocytic cell line, U937, that naturally express FcalphaR and in transfected Chinese hamster ovary (CHO), COS-7 and Hela cells. By using selective chemical inhibitors of different endocytic pathways, overexpression of dominant-negative mutants of Eps15 and knockdown of clathrin heavy chain (CHC) via RNA interference, we demonstrated that endocytosis of FcalphaR was through a clathrin-mediated pathway. The endocytosed FcalphaR went into Rab5- and Rab11-positive endosomes. However, endocytosis of FcalphaR could not be blocked by a dominant-negative mutant of Rab5. We also demonstrated that endocytosis of FcalphaR was dynamin-dependent by overexpressing a dominant-negative mutant of dynamin. The potential endocytic motif for FcalphaR was also examined. Unexpectedly, we found that the entire cytoplasmic domain of FcalphaR was not required for the endocytic process of FcalphaR. We conclude that endocytosis of FcalphaR is clathrin- and dynamin-dependent, but is not regulated by Rab5, and the endocytic motif is not located in the cytoplasmic domain of FcalphaR.

A Monomeric Chicken IgY Receptor Binds IgY with 2:1 Stoichiometry

IgY is the principal serum antibody in birds and reptiles, and an IgY-like molecule was the evolutionary precursor of both mammalian IgG and IgE. A receptor for IgY on chicken monocytes, chicken leukocyte receptor AB1 (CHIR-AB1), lies in the avian leukocyte receptor cluster rather than the classical Fc receptor cluster where the genes for mammalian IgE and IgG receptors are found. IgG and IgE receptors bind to the lower hinge region of their respective antibodies with 1:1 stoichiometry, whereas the myeloid receptor for IgA, FcalphaRI, and the IgG homeostasis receptor, FcRn, which are found in the mammalian leukocyte receptor cluster, bind with 2:1 stoichiometry between the heavy chain constant domains 2 and 3 of each heavy chain. In this paper, the extracellular domain of CHIR-AB1 was expressed in a soluble form and shown to be a monomer that binds to IgY-Fc with 2:1 stoichiometry. The two binding sites have similar affinities: K(a)(1) = 7.22 +/- 0.22 x 10(5) m(-1) and K(a)(2) = 3.63 +/- 1.03 x 10(6) m(-1) (comparable with the values reported for IgA binding to its receptor). The affinity constants for IgY and IgY-Fc binding to immobilized CHIR-AB1 are 9.07 +/- 0.07 x 10(7) and 6.11 +/- 0.02 x 10(8) m(-1), respectively, in agreement with values obtained for IgY binding to chicken monocyte cells and comparable with reported values for human IgA binding to neutrophils. Although the binding site for CHIR-AB1 on IgY is not known, the data reported here with a monomeric receptor binding to IgY at two sites with low affinity suggest an IgA-like interaction.

Relationship between Epstein-Barr virus infection and nasopharyngeal carcinoma pathogenesis

Nasopharyngeal carcinoma (NPC) is one of the common cancers among Chinese living in South China, Taiwan, Singapore, and several other countries or regions in distinct areas. The etiological factors have not been clearly identified yet. So far, no major gene related with hereditary factor has been identified in NPC carcinogenesis; however, some environmental factors, such as consumption of salted fish and long-term exposure to sulfuric-acid vapor, have been tentatively linked to NPC induction, while research has proposed that there is a close association between Epstein-Barr virus (EBV) and NPC pathogenesis. To investigate the relationship between NPC and EBV, we have established ten NPC cell lines. After extensive investigation, we conclude that EBV may establish an infection only in nasopharyngeal neoplastic cells, not in metaplastic epithelial cells, through the IgA receptor (secretory component protein)-mediated endocytosis. Our observations indicate that EBV plays an important role in enhancement of NPC progression, but is involved in neither the initiation nor the promotion of NPC pathogenesis.

Negative regulation of IgA production by the Fc&[alpha]/&[mu]R, an Fc receptor for IgA and IgM (39.13)

Abstract Fc&[alpha]/&[mu] receptor (Fc&[alpha]/&[mu]R) is a novel Fc receptor for both IgA and IgM. Fc&[alpha]/&[mu]R transcript is expressed in the gut as well as spleen and lymph node. The immunohistological analysis showed that Fc&[alpha]/&[mu]R is highly expressed on follicular dendritic cells (FDCs) in germinal centers and at lesser extent on B cells of Peyer's patches. Since Peyer's patches are known as major inductive sites of IgA-producing cells, we investigated the roles of Fc&[alpha]/&[mu]R in IgA production. We found that Fc&[alpha]/&[mu]R-deficient mice showed elevated serum, but not fecal, IgA. Moreover, after oral administration of T- independent antigens, Fc&[alpha]/&[mu]R-deficient mice showed elevated antigen-specific serum, but not fecal, IgA production. Fc&[alpha]/&[mu]R-deficient mice showed comparable half life of serum IgA to wild type mice, indicating that the production of serum IgA is increased in Fc&[alpha]/&[mu]R-deficient mice. Indeed, IgA-positive B cells were increased in Peyer's patches, mecenteric lymph nodes and spleen in Fc&[alpha]/&[mu]R-deficient mice. Bone marrow chimeric mice demonstrated that serum IgA was elevated in mice whose non-hematopoietic, but not hematopoietic, cells lack Fc&[alpha]/&[mu]R expresssion. These results suggested that Fc&[alpha]/&[mu]R expressed on FDC in Peyer's patches may regulate IgA production against intragastrically administered antigens. The molecular and cellular basis of these phenomena are under investigation.

IgA1 antibodies specific for outer membrane protein PorA modulate the interaction between Neisseria meningitidis and the epithelium

Despite high carriage rates of Neisseria meningitidis, incidence of meningococcal disease remains low, partially due to development of natural immunity. We have previously demonstrated an inverse relationship between salivary anti-meningococcal IgA and disease incidence, but little is known about the contribution of IgA to immunity at mucosal surfaces. Here we show strong immunoreactivity by human salivary IgA against the meningococcal outer membrane porin, PorA. Monomeric anti-PorA IgA1 (humanized chimeric antibodies) but not IgG increased the association of unencapsulated serogroup B N. meningitidis (H44/76) with Chang (conjunctival) but not with either Detroit (pharyngeal) cells or with A549 (alveolar) epithelial cells. Association of encapsulated N. meningitidis was not increased. Epithelial binding of IgA was Fc fragment dependent and not inhibited by IgM. Together these data suggest the presence of a specific epithelial IgA receptor that could influence the effect of both naturally acquired and vaccine induced IgA antibodies at the epithelial surface.

Human Fcα/μR and pIgR distribute differently in intestinal tissues

Fcα/μR, an Fc receptor for both IgA and IgM, is close to pIgR in gene location and has an Ig-like domain homologous to pIgR. In this study, we generated monoclonal antibodies and determined Fcα/μR and pIgR distributions in human intestinal tissues. Immunohistochemistry analysis showed that, unlike pIgR, which was expressed on epithelial cells of intestinal tissues, Fcα/μR was expressed mainly in the intestinal lamina propria and germinal centers of some lymphoid follicles. Double label immunohistochemistry analysis showed that Fcα/μR was expressed on intestinal macrophages and plasma cells. Fcα/μR was also expressed in Paneth cells. Similar expression patterns for Fcα/μR were observed in the small intestine, appendix, colon and rectum. These results indicate for the first time that Fcα/μR protein is expressed by human intestinal tissues. The different tissue distribution patterns indicate that Fcα/μR and pIgR have different roles in intestinal immunity.

IgA-specific proteins of pathogenic bacteria

Data on structure and specificity of bacterial IgA receptors (IgA-binding M-like proteins Arp4 and Sir22 from hemolytic streptococci of serogroup A, beta-antigen from hemolytic streptococci of serogroup B, and SSL family proteins from Staphylococcus aureus) are surveyed in this review. The principal conclusion derived from comparison is the fact that all bacterial receptors bind the same site in the IgA molecule overlapping with the binding site of endogenous human IgA receptor CD89. We assume that this site, consisting of spatially close amino acid strands Leu257-Gly259 in domain Calpha2 and Pro440-Phe443 in domain Calpha3, is subject to conformational rearrangement induced by the binding of antigen in the IgA active site.

Cytoplasmic domain of human Fcalpha/mu receptor is required for ligand internalization

The Fcalpha/mu receptor (Fcα/μR), a type I transmembrane protein, is an immunoglobulin Fc receptor for both IgA and IgM. Its functions in immune defense are not clear at present. In this work, human Fcα/μR was expressed in CHO, 293T, and COS-7 cells to study its biochemical functions. Fcα/μR expressed by CHO and 293T was only in monomer form in cytoplasma and the monomeric receptor could not bind IgA or IgM. In comparison, Fcα/μR expressed by COS-7 cells had both monomer and dimer forms. The binding assay showed that Fcα/μR expressed by COS-7 cells could bind IgM strongly and IgA weakly, implying that dimeric receptor could be expressed on cell membrane and functioned. The bound IgM could be internalized and the internalization was abolished when the cytoplasmic domain of Fcα/μR was truncated. Therefore, the cytoplasmic portion of human Fcα/μR is required in the internalization.

A new strategy to understand how HIV infects women: identification of a window of vulnerability during the menstrual cycle

Although 85% of new HIV cases are due to sexual transmission from men to women little attention is being paid to the immune system in the female reproductive tract (FRT) and to how it meets the conflicting challenges of protecting from pathogens and permitting procreation. As a new approach we have tried to envision how HIV evades FRT mucosal immune protection and have been led to the unexpected conclusion that in a normal menstrual cycle there is a window of vulnerability (7-10 days following ovulation) in which the potential for viral infectivity in the FRT is enhanced. During that period aspects of the innate humoral and cell-mediated immune systems are suppressed by sex hormones to optimize conditions for procreation. Suppression occurs in the upper (Fallopian tubes uterus endocervix) and lower (ectocervix and vagina) FRT and coincides with the recruitment of potentially infectable cells and upregulation of coreceptors essential for viral uptake. Implications of these findings are that the entire FRT is a potential target for HIV infection immune cells and antibodies in blood are not surrogate markers for immune protection in the FRT and immune protection against HIV will require an understanding of the hormone-induced regulation of humoral cellmediated and innate immune systems throughout the FRT. (excerpt)

Functional expression of IgA receptor FcαRI on human platelets

FcalphaRI (CD89) is a human IgA FcR expressed on cells of myeloid lineage such as neutrophils, monocytes, tissue macrophages, eosinophils, and subpopulations of dendritic cells. FcalphaRI mediates cell activation through Src family kinases and downstream tyrosine-based phosphorylation pathways. However, the role of IgA and the expression and role of its cognate receptor FcalphaRI (CD89) in platelet activation are undefined. In the current study, we demonstrate that human platelets express FcalphaRI mRNAs and proteins. Furthermore, we show that the platelet FcalphaRI is associated with the FcR gamma-chain, and cross-linking of FcalphaRI leads to Syk phosphorylation. Clustering of FcalphaRI induces pre-mRNA splicing and protein production of tissue factor and IL-1beta, suggesting novel roles for human platelet FcalphaRI and serum IgA in thrombosis and inflammation.

Inside-Out Regulation of FcαRI (CD89) Depends on PP2A

To achieve a correct cellular immune response toward pathogens, interaction between FcR and their ligands must be regulated. The Fc receptor for IgA, FcalphaRI, is pivotal for the inflammatory responses against IgA-opsonized pathogens. Cytokine-induced inside-out signaling through the intracellular FcalphaRI tail is important for FcalphaRI-IgA binding. However, the underlying molecular mechanism governing this process is not well understood. In this study, we report that PP2A can act as a molecular switch in FcalphaRI activation. PP2A binds to the intracellular tail of FcalphaRI and, upon cytokine stimulation, PP2A becomes activated. Subsequently, FcalphaRI is dephosphorylated on intracellular Serine 263, which we could link to receptor activation. PP2A inhibition, in contrast, decreased FcalphaRI ligand binding capacity in transfected cells but also in eosinophils and monocytes. Interestingly, PP2A activity was found crucial for IgA-mediated binding and phagocytosis of Neisseria meningitidis. The present findings demonstrate PP2A involvement as a molecular mechanism for FcalphaRI ligand binding regulation, a key step in initiating an immune response.

The Glomerular Response to IgA Deposition in IgA Nephropathy

Compelling evidence points to a role for IgA receptors in the pathogenesis of IgA nephropathy. The soluble form of the type I IgA receptor (FcalphaRI or CD89) forms complexes with IgA that can be found in patients' serum and that initiate the disease in CD89 transgenic mice. A nonclassic IgA receptor, identified as the transferrin receptor (TfR), is highly expressed in patients' mesangium and colocalizes with IgA deposits. TfR preferentially binds polymeric IgA1 complexes, but not monomeric IgA1 or IgA2. The TfR-IgA1 interaction is dependent on carbohydrate moieties because hypoglycosylated IgA1 has superior binding to TfR than normally glycosylated IgA1. Polymeric IgA1 binding enhances mesangial cell TfR expression and results in cell proliferation and inflammatory and profibrogenic cytokine and chemokine production, suggesting a pivotal role in mesangial cell proliferation, matrix expansion, and recruitment of inflammatory cells. We propose that, as a second event, activation of the classic, FcRgamma-associated transmembrane FcalphaRI expressed on circulating myeloid leukocytes takes place. FcalphaRI/gamma2 cross-linking in human FcalphaRI transgenic animals promotes disease progression by enhancing leukocyte chemotaxis and cytokine production, and IgA immune complexes from IgA nephropathy patients induce FcalphaRI-dependent cell activation. This review therefore details the functional consequences of IgA/receptor interactions and discusses proposed mechanisms to explain the development and chronicity of the disease.

Unusual biochemical features and follicular dendritic cell expression of human Fcα/μ receptor

The Fc receptor for IgA and IgM (Fcalpha/muR) is of particular interest because it can bind antibodies of both IgM and IgA isotypes and thus may play a pivotal role in systemic and mucosal immunity. Using IgM and IgA ligands and newly generated Fcalpha/muR specific monoclonal antibodies we have defined biochemical features and cellular distribution of the human Fcalpha/muR. Both recombinant and native forms of human Fcalpha/muR are expressed on the cell surface as remarkably stable homodimeric transmembrane glycoproteins that can bind specifically polymeric IgM or IgA. The only human B cells to express Fcalpha/muR, albeit at very low levels, are found in the pre-germinal center subpopulation defined by the IgD+/CD38+ phenotype. Hence the expression pattern differs from that of the mouse wherein Fcalpha/muR is expressed by both circulating and resident B cell populations. Significantly, the predominant cell type expressing the Fcalpha/muR in humans is the follicular dendritic cell of germinal centers. The Fcalpha/muR may thus function in antigen presentation and B cell selection in the germinal center response.

The crystal structure of staphylococcal superantigen‐like protein 11 in complex with sialyl Lewis X reveals the mechanism for cell binding and immune inhibition

Staphylococcus aureus is a major pathogen that produces a family of 14 staphylococcal superantigen-like (SSL) proteins, which are structurally similar to superantigens but do not stimulate T cells. SSL11 is one member of the family that is found in all staphylococcal strains. Recombinant SSL11 bound to granulocytes and monocytes through a sialic acid-dependent mechanism and was rapidly internalized. SSL11 also bound to sialic acid-containing glycoproteins, such as the Fc receptor for IgA (FcalphaRI) and P-selectin glycoprotein ligand-1 (PSGL-1), and inhibited neutrophil attachment to a P-selectin-coated surface. Biosensor analysis of two SSL11 alleles binding to sialyl Lewis X [sLe(x)- Neu5Acalpha2-3Galbeta1-4(Fuc1-3)GlcNAc] coupled to bovine serum albumin gave dissociation constants of 0.7 and 7 mum respectively. Binding of SSL11 to a glycan array revealed specificity for glycans containing the trisaccharide sialyllactosamine (sLacNac - Neu5Acalpha2-3Galbeta1-4GlcNAc). A 1.6 A resolution crystal structure of SSL11 complexed with sLe(x) revealed a discrete binding site in the C-terminal beta-grasp domain, with predominant interactions with the sialic acid and galactose residues. A single amino acid mutation in the carbohydrate binding site abolished all SSL11 binding. Thus, SSL11 is a staphylococcal protein that targets myeloid cells by binding sialyllactosamine-containing glycoproteins.

Structural basis for evasion of IgA immunity by Staphylococcus aureus revealed in the complex of SSL7 with Fc of human IgA1.

Infection by Staphylococcus aureus can result in severe conditions such as septicemia, toxic shock, pneumonia, and endocarditis with antibiotic resistance and persistent nasal carriage in normal individuals being key drivers of the medical impact of this virulent pathogen. In both virulent infection and nasal colonization, S. aureus encounters the host immune system and produces a wide array of factors that frustrate host immunity. One in particular, the prototypical staphylococcal superantigen-like protein SSL7, potently binds IgA and C5, thereby inhibiting immune responses dependent on these major immune mediators. We report here the three-dimensional structure of the complex of SSL7 with Fc of human IgA1 at 3.2 A resolution. Two SSL7 molecules interact with the Fc (one per heavy chain) primarily at the junction between the Calpha2 and Calpha3 domains. The binding site on each IgA chain is extensive, with SSL7 shielding most of the lateral surface of the Calpha3 domain. However, the SSL7 molecules are positioned such that they should allow binding to secretory IgA. The key IgA residues interacting with SSL7 are also bound by the leukocyte IgA receptor, FcalphaRI (CD89), thereby explaining how SSL7 potently inhibits IgA-dependent cellular effector functions mediated by FcalphaRI, such as phagocytosis, degranulation, and respiratory burst. Thus, the ability of S. aureus to subvert IgA-mediated immunity is likely to facilitate survival in mucosal environments such as the nasal passage and may contribute to systemic infections.

Exogenous MAL Reroutes Selected Hepatic Apical Proteins into the Direct Pathway in WIF-B Cells

Unlike simple epithelial cells that directly target newly synthesized glycophosphatidylinositol (GPI)-anchored and single transmembrane domain (TMD) proteins from the trans-Golgi network to the apical membrane, hepatocytes use an indirect pathway: proteins are delivered to the basolateral domain and then selectively internalized and transcytosed to the apical plasma membrane. Myelin and lymphocyte protein (MAL) and MAL2 have been identified as regulators of direct and indirect apical delivery, respectively. Hepatocytes lack endogenous MAL consistent with the absence of direct apical targeting. Does MAL expression reroute hepatic apical residents into the direct pathway? We found that MAL expression in WIF-B cells induced the formation of cholesterol and glycosphingolipid-enriched Golgi domains that contained GPI-anchored and single TMD apical proteins; polymeric IgA receptor (pIgA-R), polytopic apical, and basolateral resident distributions were excluded. Basolateral delivery of newly synthesized apical residents was decreased in MAL-expressing cells concomitant with increased apical delivery; pIgA-R and basolateral resident delivery was unchanged. These data suggest that MAL rerouted selected hepatic apical proteins into the direct pathway.

Control of Germinal Center Reaction against T-Independent Antigens by the Fc(alpha)/(mu)Receptor (53.7)

Abstract Germinal center (GC) formation in the secondary lymphoid follicles is induced by immunization with T-independent (TI) as well as T-dependent (TD) antigens. However, structural and functional characteristics of the GC induced by TI antigens have not been elucidated. Here, we show that marginal zone B (MZB) cells and follicular dendritic cells (FDC) preferentially express the Fc receptor for IgM and IgA (Fc(alpha)/(mu)R) as well as the complement receptor CD21/35. Mice deficient in Fc(alpha)/(mu)R showed significantly enhanced GC formation in response to TI antigens, in which both B cells and FDC were involved. Fc(alpha)/(mu)R-deficient mice also showed enhanced TI antigen retention by MZB cells and FDC, CXCL13 expression and affinity maturation of IgG3. Depletion of complements by injection with cobra venom factor completely abrogated this phenotype. These results suggest that Fc(alpha)/(mu)R controls GC reactions and affinity maturation of antibodies against TI antigens by interacting with complement cascades.