Factor D.

Complement factor D (FD) is a serine protease that plays an essential role in the activation of the alternative pathway (AP) by cleaving complement factor B (FB) and generating the C3 convertases C3(H2 O)Bb and C3bBb. FD is produced mainly from adipose tissue and circulates in an activated form. On the contrary, the other serine proteases of the complement system are mainly synthesized in the liver. The activation mechanism of FD has long been unknown. Recently, a serendipitous discovery in the mechanism of FD activation has been provided by a generation of Masp1 gene knockout mice lacking both the serine protease MASP-1 and its alternative splicing variant MASP-3, designated MASP-1/3-deficient mice. Sera from the MASP-1/3-deficient mice had little-to-no lectin pathway (LP) and AP activity with circulating zymogen or proenzyme FD (pro-FD). Sera from patients with 3MC syndrome carrying mutations in the MASP1 gene also had circulating pro-FD, suggesting that MASP-1 and/or MASP-3 are involved in activation of FD. Here, we summarize the current knowledge of the mechanism of FD activation that was finally elucidated using the sera of mice monospecifically deficient for MASP-1 or MASP-3. Sera of the MASP-1-deficient mice lacked LP activity, but those of the MASP-3-deficient mice lacked AP activity with pro-FD. This review illustrates the pivotal role of MASP-3 in the physiological activation of the AP via activation of FD.

A novel complement inhibitor sMAP-FH targeting both the lectin and alternative complement pathways.

Inhibition of the complement activation has emerged as an option for treatment of a range of diseases. Activation of the lectin and alternative pathways (LP and AP, respectively) contribute to the deterioration of conditions in certain diseases such as ischemia-reperfusion injuries and age-related macular degeneration (AMD). In the current study, we generated dual complement inhibitors of the pathways MAp44-FH and sMAP-FH by fusing full-length MAp44 or small mannose-binding lectin-associated protein (sMAP), LP regulators, with the N-terminal five short consensus repeat (SCR) domains of complement factor H (SCR1/5-FH), an AP regulator. The murine forms of both fusion proteins formed a complex with endogenous mannose-binding lectin (MBL) or ficolin A in the circulation when administered in mice intraperitoneally. Multiple complement activation assays revealed that sMAP-FH had significantly higher inhibitory effects on activation of the LP and AP in vivo as well as in vitro compared to MAp44-FH. Human form of sMAP-FH also showed dual inhibitory effects on LP and AP activation in human sera. Our results indicate that the novel fusion protein sMAP-FH inhibits both the LP and AP activation in mice and in human sera, and could be an effective therapeutic agent for diseases in which both the LP and AP activation are significantly involved.

Cutting Edge: Role of MASP-3 in the Physiological Activation of Factor D of the Alternative Complement Pathway.

The complement system, a part of the innate immune system, can be activated via three different pathways. In the alternative pathway, a factor D (FD) plays essential roles in both the initiation and the amplification loop and circulates as an active form. Mannose-binding lectin-associated serine proteases (MASPs) are key enzymes of the lectin pathway, and MASP-1 and/or MASP-3 are reported to be involved in the activation of FD. In the current study, we generated mice monospecifically deficient for MASP-1 or MASP-3 and found that the sera of the MASP-1-deficient mice lacked lectin pathway activity, but those of the MASP-3-deficient mice lacked alternative pathway activity with a zymogen FD. Furthermore, the results indicate that MASP-3 but not MASP-1 activates the zymogen FD under physiological conditions and MASP-3 circulates predominantly as an active form. Therefore, our study illustrates that, in mice, MASP-3 orchestrates the overall complement reaction through the activation of FD.

Deconstructing the Lectin Pathway in the Pathogenesis of Experimental Inflammatory Arthritis: Essential Role of the Lectin Ficolin B and Mannose-Binding Protein-Associated Serine Protease 2.

Complement plays an important role in the pathogenesis of rheumatoid arthritis. Although the alternative pathway (AP) is known to play a key pathogenic role in models of rheumatoid arthritis, the importance of the lectin pathway (LP) pattern recognition molecules such as ficolin (FCN) A, FCN B, and collectin (CL)-11, as well as the activating enzyme mannose-binding lectin-associated serine protease-2 (MASP-2), are less well understood. We show in this article that FCN A-/- and CL-11-/- mice are fully susceptible to collagen Ab-induced arthritis (CAIA). In contrast, FCN B-/- and MASP-2-/-/sMAp-/- mice are substantially protected, with clinical disease activity decreased significantly (p < 0.05) by 47 and 70%, respectively. Histopathology scores, C3, factor D, FCN B deposition, and infiltration of synovial macrophages and neutrophils were similarly decreased in FCN B-/- and MASP-2-/-/sMAp-/- mice. Our data support that FCN B plays an important role in the development of CAIA, likely through ligand recognition in the joint and MASP activation, and that MASP-2 also contributes to the development of CAIA, likely in a C4-independent manner. Decreased AP activity in the sera from FCN B-/- and MASP-2-/-/sMAp-/- mice with arthritis on adherent anti-collagen Abs also support the hypothesis that pathogenic Abs, as well as additional inflammation-related ligands, are recognized by the LP and operate in vivo to activate complement. Finally, we also speculate that the residual disease seen in our studies is driven by the AP and/or the C2/C4 bypass pathway via the direct cleavage of C3 through an LP-dependent mechanism.

Lectin pathway effector enzyme mannan-binding lectin-associated serine protease-2 can activate native complement C3 in absence of C4 and/or C2.

All 3 activation pathways of complement-the classic pathway (CP), the alternative pathway, and the lectin pathway (LP)- converge into a common central event: the cleavage and activation of the abundant third complement component, C3, via formation of C3-activating enzymes (C3 convertases). The fourth complement component, C4, and the second component, C2, are indispensable constituents of the C3 convertase complex, C4bC2a, which is formed by both the CP and the LP. Whereas in the absence of C4, CP can no longer activate C3, LP retains a residual but physiologically critical capacity to convert native C3 into its activation fragments, C3a and C3b. This residual C4 and/or C2 bypass route is dependent on LP-specific mannan-binding lectin-associated serine protease-2. By using various serum sources with defined complement deficiencies, we demonstrate that, under physiologic conditions LP-specific C4 and/or C2 bypass activation of C3 is mediated by direct cleavage of native C3 by mannan-binding lectin-associated serine protease-2 bound to LP-activation complexes captured on ligand-coated surfaces.-Yaseen, S., Demopulos, G., Dudler, T., Yabuki, M., Wood, C. L., Cummings, W. J., Tjoelker, L. W., Fujita, T., Sacks, S., Garred, P., Andrew, P., Sim, R. B., Lachmann, P. J., Wallis, R., Lynch, N., Schwaeble, W. J. Lectin pathway effector enzyme mannan-binding lectin-associated serine protease-2 can activate native complement C3 in absence of C4 and/or C2.

Mannan-Binding Lectin-Associated Serine Protease 1/3 Cleavage of Pro-Factor D into Factor D In Vivo and Attenuation of Collagen Antibody-Induced Arthritis through Their Targeted Inhibition by RNA Interference-Mediated Gene Silencing.

The complement system is proposed to play an important role in the pathogenesis of rheumatoid arthritis (RA). The complement system mannan-binding lectin-associated serine proteases (MASP)-1/3 cleave pro-factor D (proDf; inactive) into Df (active), but it is unknown where this cleavage occurs and whether inhibition of MASP-1/3 is a relevant therapeutic strategy for RA. In the present study, we show that the cleavage of proDf into Df by MASP-1/3 can occur in the circulation and that inhibition of MASP-1/3 by gene silencing is sufficient to ameliorate collagen Ab-induced arthritis in mice. Specifically, to examine the cleavage of proDf into Df, MASP-1/3-producing Df-/- liver tissue (donor) was transplanted under the kidney capsule of MASP-1/3-/- (recipient) mice. Five weeks after the liver transplantation, cleaved Df was present in the circulation of MASP-1/3-/- mice. To determine the individual effects of MASP-1/3 and Df gene silencing on collagen Ab-induced arthritis, mice were injected with scrambled, MASP-1/3-targeted, or Df-targeted small interfering RNAs (siRNAs). The mRNA levels for MASP-1 and -3 decreased in the liver to 62 and 58%, respectively, in mice injected with MASP-1/3 siRNAs, and Df mRNA decreased to 53% in the adipose tissue of mice injected with Df siRNAs; additionally, circulating MASP-1/3 and Df protein levels were decreased. In mice injected with both siRNAs the clinical disease activity, histopathologic injury scores, C3 deposition, and synovial macrophage/neutrophil infiltration were significantly decreased. Thus, MASP-1/3 represent a new therapeutic target for the treatment of RA, likely through both direct effects on the lectin pathway and indirectly through the alternative pathway.

Mannan binding lectin-associated serine protease-2 (MASP-2) critically contributes to post-ischemic brain injury independent of MASP-1.

BACKGROUND: Complement activation via the lectin activation pathway (LP) has been identified as the key mechanism behind post-ischemic tissue inflammation causing ischemia-reperfusion injury (IRI) which can significantly impact the clinical outcome of ischemic disease. This work defines the contributions of each of the three LP-associated enzymes-mannan-binding lectin-associated serine protease (MASP)-1, MASP-2, and MASP-3-to ischemic brain injury in experimental mouse models of stroke. METHODS: Focal cerebral ischemia was induced in wild-type (WT) mice or mice deficient for defined complement components by transient middle cerebral artery occlusion (tMCAO) or three-vessel occlusion (3VO). The inhibitory MASP-2 antibody was administered systemically 7 and 3.5 days before and at reperfusion in WT mice in order to assure an effective MASP-2 inhibition throughout the study. Forty-eight hours after ischemia, neurological deficits and infarct volumes were assessed. C3 deposition and microglia/macrophage morphology were detected by immunohistochemical, immunofluorescence, and confocal analyses. RESULTS: MASP-2-deficient mice (MASP-2(-/-)) and WT mice treated with an antibody that blocks MASP-2 activity had significantly reduced neurological deficits and histopathological damage after transient ischemia and reperfusion compared to WT or control-treated mice. Surprisingly, MASP-1/3(-/-) mice were not protected, while mice deficient in factor B (fB(-/-)) showed reduced neurological deficits compared to WT mice. Consistent with behavioral and histological data, MASP-2(-/-) had attenuated C3 deposition and presented with a significantly higher proportion of ramified, surveying microglia in contrast to the hypertrophic pro-inflammatory microglia/macrophage phenotype seen in the ischemic brain tissue of WT mice. CONCLUSIONS: This work demonstrates the essential role of the low-abundant MASP-2 in the mediation of cerebral ischemia-reperfusion injury and demonstrates that targeting MASP-2 by an inhibitory therapeutic antibody markedly improved the neurological and histopathological outcome after focal cerebral ischemia. These results contribute to identifying the key lectin pathway component driving brain tissue injury following cerebral ischemia and call for a revision of the presently widely accepted view that MASP-1 is an essential activator of the lectin pathway effector component MASP-2.

Granular C3 Dermatosis.

There has been no previous systematic study of bullous skin diseases with granular basement membrane zone deposition exclusively of C3. In this study we collected 20 such patients, none of whom showed cutaneous vasculitis histopathologically. Oral dapsone and topical steroids were effective. Various serological tests detected no autoantibodies or autoantigens. Direct immunofluorescence for various complement components revealed deposition only of C3 and C5-C9, indicating that no known complement pathways were involved. Studies of in situ hybridization and micro-dissection with quantitative RT-PCR revealed a slight reduction in expression of C3 in patient epidermis. These patients may represent a new disease entity, for which we propose the term "granular C3 dermatosis". The mechanism for granular C3 deposition in these patients is unknown, but it is possible that the condition is caused by autoantibodies to skin or aberrant C3 expression in epidermal keratinocytes.

Evolution of the lectin-complement pathway and its role in innate immunity.

Discrimination between self and non-self by lectins (carbohydrate-binding proteins) is a strategy of innate immunity that is found in both vertebrates and invertebrates. In vertebrates, immune recognition mediated by ficolins (lectins that consist of a fibrinogen-like and a collagen-like domain), as well as by mannose-binding lectins, triggers the activation of the complement system, which results in the activation of novel serine proteases. The presence of a similar lectin-based complement system in ascidians, our closest invertebrate relatives, indicates that the complement system probably had a pivotal role in innate immunity before the evolution of an adaptive immune system in jawed vertebrates.

Mitochondria and the lectin pathway of complement.

Mitochondria, the powerhouses of our cells, are remnants of a eubacterial endosymbiont. Notwithstanding the evolutionary time that has passed since the initial endosymbiotic event, mitochondria have retained many hallmarks of their eubacterial origin. Recent studies have indicated that during perturbations of normal homeostasis, such as following acute trauma leading to massive necrosis and release of mitochondria, the immune system might mistake symbiont for enemy and initiate an inappropriate immune response. The innate immune system is the first line of defense against invading microbial pathogens, and as such is the primary suspect in the recognition of mitochondria-derived danger-associated molecular patterns and initiation of an aberrant response. Conversely, innate immune mechanisms are also central to noninflammatory clearance of innocuous agents. Here we investigated the role of a central humoral component of innate immunity, the lectin pathway of complement, in recognition of mitochondria in vitro and in vivo. We found that the soluble pattern recognition molecules, mannan-binding lectin (MBL), L-ficolin, and M-ficolin, were able to recognize mitochondria. Furthermore, MBL in complex with MBL-associated serine protease 2 (MASP-2) was able to activate the lectin pathway and deposit C4 onto mitochondria, suggesting that these molecules are involved either in homeostatic clearance of mitochondria or in induction of untoward inflammatory reactions. We found that following mitochondrial challenge, C3 was consumed in vivo in the absence of overt inflammation, indicating a potential role of complement in noninflammatory clearance of mitochondria. Thus, we report here the first indication of involvement of the lectin pathway in mitochondrial immune handling.

The lectin pathway of complement activation is a critical component of the innate immune response to pneumococcal infection.

The complement system plays a key role in host defense against pneumococcal infection. Three different pathways, the classical, alternative and lectin pathways, mediate complement activation. While there is limited information available on the roles of the classical and the alternative activation pathways of complement in fighting streptococcal infection, little is known about the role of the lectin pathway, mainly due to the lack of appropriate experimental models of lectin pathway deficiency. We have recently established a mouse strain deficient of the lectin pathway effector enzyme mannan-binding lectin associated serine protease-2 (MASP-2) and shown that this mouse strain is unable to form the lectin pathway specific C3 and C5 convertases. Here we report that MASP-2 deficient mice (which can still activate complement via the classical pathway and the alternative pathway) are highly susceptible to pneumococcal infection and fail to opsonize Streptococcus pneumoniae in the none-immune host. This defect in complement opsonisation severely compromises pathogen clearance in the lectin pathway deficient host. Using sera from mice and humans with defined complement deficiencies, we demonstrate that mouse ficolin A, human L-ficolin, and collectin 11 in both species, but not mannan-binding lectin (MBL), are the pattern recognition molecules that drive lectin pathway activation on the surface of S. pneumoniae. We further show that pneumococcal opsonisation via the lectin pathway can proceed in the absence of C4. This study corroborates the essential function of MASP-2 in the lectin pathway and highlights the importance of MBL-independent lectin pathway activation in the host defense against pneumococci.

Mannose-binding lectin-associated serine protease-1 is a significant contributor to coagulation in a murine model of occlusive thrombosis.

Bleeding disorders and thrombotic complications constitute a major cause of death and disability worldwide. Although it is known that the complement and coagulation systems interact, no studies have investigated the specific role or mechanisms of lectin-mediated coagulation in vivo. FeCl(3) treatment resulted in intra-arterial occlusive thrombogenesis within 10 min in wild-type (WT) and C2/factor B-null mice. In contrast, mannose-binding lectin (MBL)-null and MBL-associated serine protease (MASP)-1/-3 knockout (KO) mice had significantly decreased FeCl(3)-induced thrombogenesis. Reconstitution with recombinant human (rh) MBL restored FeCl(3)-induced thrombogenesis in MBL-null mice to levels comparable to WT mice, suggesting a significant role of the MBL/MASP complex for in vivo coagulation. Additionally, whole blood aggregation demonstrated increased MBL/MASP complex-dependent platelet aggregation. In vitro, MBL/MASP complexes were captured on mannan-coated plates, and cleavage of a chromogenic thrombin substrate (S2238) was measured. We observed no significant differences in S2238 cleavage between WT, C2/factor B-null, MBL-A(-/-), or MBL-C(-/-) sera; however, MBL-null or MASP-1/-3 KO mouse sera demonstrated significantly decreased S2238 cleavage. rhMBL alone failed to cleave S2238, but cleavage was restored when rMASP-1 was added to either MASP-1/-3 KO sera or rhMBL. Taken together, these findings indicate that MBL/MASP complexes, and specifically MASP-1, play a key role in thrombus formation in vitro and in vivo.

Mice deficient in ficolin, a lectin complement pathway recognition molecule, are susceptible to Streptococcus pneumoniae infection.

Mannose-binding lectin (MBL) and ficolin are complexed with MBL-associated serine proteases, key enzymes of complement activation via the lectin pathway, and act as soluble pattern recognition molecules in the innate immune system. Although numerous reports have revealed the importance of MBL in infectious diseases and autoimmune disorders, the role of ficolin is still unclear. To define the specific role of ficolin in vivo, we generated model mice deficient in ficolins. The ficolin A (FcnA)-deficient (Fcna(-/-)) and FcnA/ficolin B double-deficient (Fcna(-/-)b(-/-)) mice lacked FcnA-mediated complement activation in the sera, because of the absence of complexes comprising FcnA and MBL-associated serine proteases. When the host defense was evaluated by transnasal infection with a Streptococcus pneumoniae strain, which was recognized by ficolins, but not by MBLs, the survival rate was significantly reduced in all three ficolin-deficient (Fcna(-/-), Fcnb(-/-), and Fcna(-/-)b(-/-)) mice compared with wild-type mice. Reconstitution of the FcnA-mediated lectin pathway in vivo improved survival rate in Fcna(-/-) but not in Fcna(-/-)b(-/-) mice, suggesting that both FcnA and ficolin B are essential in defense against S. pneumoniae. These results suggest that ficolins play a crucial role in innate immunity against pneumococcal infection through the lectin complement pathway.

Targeting of mannan-binding lectin-associated serine protease-2 confers protection from myocardial and gastrointestinal ischemia/reperfusion injury.

Complement research experienced a renaissance with the discovery of a third activation route, the lectin pathway. We developed a unique model of total lectin pathway deficiency, a mouse strain lacking mannan-binding lectin-associated serine protease-2 (MASP-2), and analyzed the role of MASP-2 in two models of postischemic reperfusion injury (IRI). In a model of transient myocardial IRI, MASP-2-deficient mice had significantly smaller infarct volumes than their wild-type littermates. Mice deficient in the downstream complement component C4 were not protected, suggesting the existence of a previously undescribed lectin pathway-dependent C4-bypass. Lectin pathway-mediated activation of C3 in the absence of C4 was demonstrated in vitro and shown to require MASP-2, C2, and MASP-1/3. MASP-2 deficiency also protects mice from gastrointestinal IRI, as do mAb-based inhibitors of MASP-2. The therapeutic effects of MASP-2 inhibition in this experimental model suggest the utility of anti-MASP-2 antibody therapy in reperfusion injury and other lectin pathway-mediated disorders.

The role of mannose-binding lectin-associated serine protease-3 in activation of the alternative complement pathway.

Mannose-binding lectin (MBL)-associated serine proteases (MASPs) are responsible for activation of the lectin complement pathway. Three types of MASPs (MASP-1, MASP-2, and MASP-3) are complexed with MBL and ficolins in serum. Although MASP-1 and MASP-2 are known to contribute to complement activation, the function of MASP-3 remains unclear. In this study, we investigated the mechanism of MASP-3 activation and its substrate using the recombinant mouse MASP-3 (rMASP-3) and several different types of MASP-deficient mice. A proenzyme rMASP-3 was obtained that was not autoactivated during preparation. The recombinant enzyme was activated by incubation with Staphylococcus aureus in the presence of MBL-A, but not MBL-C. In vivo studies revealed the phagocytic activities of MASP-1/3-deficient mice and all MASPs (MASP-1/2/3)-deficient mice against S. aureus and bacterial clearance in these mice were lower than those in wild-type and MASP-2-deficient mice. Sera from all MASPs-deficient mice showed significantly lower C3 deposition activity on the bacteria compared with that of wild-type serum, and addition of rMASP-3 to the deficient serum restored C3 deposition. The low C3 deposition in sera from all MASPs-deficient mice was probably caused by the low level factor B activation that was ameliorated by the addition of rMASP-3. Furthermore, rMASP-3 directly activated factors B and D in vitro. These results suggested that MASP-3 complexed with MBL is converted to an active form by incubation with bacterial targets, and that activated MASP-3 triggered the initial activation step of the alternative complement pathway.

The role of MASP-1/3 in complement activation.

The complement system, which consists of more than 30 plasma and cell surface proteins, is activated by three pathways: the classical, lectin, and alternative pathways, leading to the generation of opsonins and pathogen destruction. In the lectin pathway, mannose-binding lectin (MBL) and ficolins act as pattern recognition molecules for pathogens, resulting in the activation of MBL-associated serine proteases (MASPs: MASP-1, MASP-2, and MASP-3). Among these proteases, MASP-2 is a key enzyme that cleaves C4 and C2 to assemble a C3 convertase (C4b2a). However, the physiological function of MASP-1 and MASP-3 remains unclear. To investigate the roles of MASP-1 and MASP-3, we generated a MASP-1- and MASP-3-deficient (M1/3 KO) mouse model and found that the deficient mice lacked alternative pathway activation because factor D (Df) remained as a proenzyme in the serum. MASP-1 and MASP-3 were able to convert the proenzyme of Df to an active form in vitro. In addition, MASP-1 was able to activate MASP-2 and MASP-3 as C1r activates C1s. Thus, MASP-1 and MASP-3 seem to be involved in activation of both the lectin and alternative pathways.

Mouse ficolin B has an ability to form complexes with mannose-binding lectin-associated serine proteases and activate complement through the lectin pathway.

Ficolins are thought to be pathogen-associated-molecular-pattern-(PAMP-) recognition molecules that function to support innate immunity. Like mannose-binding lectins (MBLs), most mammalian ficolins form complexes with MBL-associated serine proteases (MASPs), leading to complement activation via the lectin pathway. However, the ability of murine ficolin B, a homologue of human M-ficolin, to perform this function is still controversial. The results of the present study show that ficolin B in mouse bone marrow is an oligomeric protein. Ficolin B, pulled down using GlcNAc-agarose, contained very low, but detectable, amounts of MASP-2 and small MBL-associated protein (sMAP) and showed detectable C4-deposition activity on immobilized N-acetylglucosamine. These biochemical features of ficolin B were confirmed using recombinant mouse ficolin B produced in CHO cells. Taken together, these results suggest that like other mammalian homologues, murine ficolin B has an ability to exert its function via the lectin pathway.

Essential role of complement mannose-binding lectin-associated serine proteases-1/3 in the murine collagen antibody-induced model of inflammatory arthritis.

Gene-targeted mice deficient in the complement mannose-binding lectin-associated serine protease-1 and -3 (MASP1/3(-/-)) express only the zymogen of factor D (pro-factor D [pro-Df]), a necessary component of the alternative pathway (AP). We used the murine collagen Ab-induced arthritis (CAIA) model, in which the AP is unique among complement pathways in being both necessary and sufficient for disease induction, to determine whether MASP-1/3 are required in vivo for the development of tissue injury. Disease activity scores, complement C3 tissue deposition in the joint, and histopathologic injury scores were markedly decreased in MASP1/3(-/-) as compared with wild-type (WT) mice. MASP-1 protein was immunochemically localized to synovial cells of knees of WT mice with arthritis. Pro-Df was present in both synovial cells and chondrocytes of knees of WT and MASP1/3(-/-) mice without arthritis, with increased amounts present in synovial cells of WT mice with CAIA. No conversion of pro-Df to mature Df was detectable in the serum of MASP1/3(-/-) mice during the evolution of CAIA. C3 activation and deposition as well as C5a generation induced in vitro by adherent anti-type II collagen mAbs were absent using sera from MASP1/3(-/-) mice under conditions in which only the AP was active. The addition of human Df fully reconstituted in vitro C3 activation and C5a generation using sera from MASP1/3(-/-) mice. Our studies demonstrate for the first time, to our knowledge, the absolute requirement for the activity of MASP-1 protein in autoimmune-associated inflammatory tissue injury in vivo through activation of the AP of complement by cleavage of pro-Df to mature Df.

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.