Mannan-binding lectin—a soluble pattern recognition molecule

Abstract

The immune system must discriminate between self and non-self. Mannan-binding lectin (MBL) is a recognition molecule able to differentiate between the carbohydrates found on self glycoproteins and the carbohydrate patterns found on infectious non-self surfaces. It exists in a complex with MBL-associated serine proteases (MASPs). When MBL binds to suitable carbohydrate pattern it causes activation of MASPs leading to triggering of the complement cascade. This results in limiting the infection and the orchestration of subsequent adaptive immune response. The plasma concentration of MBL is determined by genetic polymorphisms. Deficiency of MBL is a risk factor for infection, especially when other functions in the immune system are also compromised. MBL has a potential to react against altered-self structures, as found on apoptotic cells, cancers or ischemic-reperfused tissue. The focus of the current review is to summarize the recent progress in our understanding of MBL functions.

Introduction

Central to tolerance, i.e., non-responsiveness, is the ability to distinguish between self, infectious non-self and altered self. Tolerance is present in the adaptive immune system due to a number of mechanisms. On the other hand tolerance is not induced in the innate immune system. Reactions against self are subdued due to low affinity binding to self structures or because of an inhibition of a response (Medzhitov and Janeway, 2002). Cells of the innate immune system may react when regulatory proteins are missing (missing-self surfaces). An example is the inhibition of NK cells by presence of MHC molecules. When surfaces are low for MHC this may lead to less inhibition and thus response can occur towards missing-self (Yokoyama and Scalzo, 2002). Another example is the lowering of cell signaling given to cells when certain sialic acid specific lectins (siglecs, e.g., CD22 and CD33-related Siglecs) react with sialic acid structures on the cell with which they are in contact (Crocker and Varki, 2001). Similarly, soluble proteins of the innate immune system may be deposited to a much greater extent on missing-self surfaces. Complement factors are better deposited on cells/microorganisms that do not posses enough complement control proteins (Sahu and Lambris, 2000).

A crosstalk exists between the innate and the adaptive immune system. It has been shown that the complement system may be involved in up or down regulation of antibody responses (Barrington et al., 2001, Pepys, 1972). A number of mechanisms may be implicated, e.g., the increased phagocytosis initiated by complement deposition may lead to enhanced presentation of peptides to the adaptive immune system and/or the cells phagocytosing complement coated microorganisms get a second stimulus as compared to situations where no complement is deposited. Humoral complement activators, like the mannan-binding lectin (MBL) described below may influence the immune response towards microorganisms or soluble antigens and will hence modulate the response toward infective agents.

MBL is a member of the innate immune system and initiates complement deposition on relevant surfaces (Turner, 2003). It is a plasma protein with the ability to distinguish between self and non-self by recognizing certain patterns of carbohydrate structures. This ability is based on a certain clustering of recognition sites within the target surface. MBL is an oligomer of polypeptide chains. Each chain is composed of a collagen-like region linked to a carbohydrate recognition domain (CRD) (Fig. 1). When MBL is fully assembled, the CRDs have the ability to react with patterns of carbohydrates. MBL thus binds to carbohydrates at two levels (Fig. 1): firstly, each CRD will recognize single oligosaccharide structures on the ligand. The dissociation constant of the binding between a single recombinant rat MBL-A CRD and a target monosaccharide was estimated to be 1.5×10⁻³ M by analyzing the chemical shifts via nuclear magnetic resonance at different monosaccharide concentrations (Iobst et al., 1994). Due to the initial description of MBL binding ligands, MBL was also termed mannose-binding lectin and mannose-binding protein. But it should be noted that MBL does not selectively recognize only mannose or its multimers. The CRDs bind with similar affinity to GlcNAc and fucose. Secondly, ligands with multiple possible binding sites will have to fit the geometry of oligomeric CRDs. This will lead to a clustering of several CRDs onto a surface and thus the strength of binding may be expressed as avidity. Scatchard plot analysis have demonstrated that the binding between native MBL and polyvalent carbohydrates or microorganisms occurs with a dissociation constants in the range of 10⁻⁹ M, i.e., comparable to the binding of good fitting antibodies to antigen (Kawasaki et al., 1983). MBL only obtains sufficient binding strength when a pattern corresponding to the spatial spanning of the CRDs of MBL are present, e.g., as in mannan.

The level of MBL in the blood is influenced by single nucleotide polymorphisms (SNPs) in the MBL gene (mbl2) (Turner, 2003). As illustrated in Fig. 2 there are polymorphisms in the promoter region (X or Y) and in exon 1 (B, C and D—A is the wild type). The mutations in exon 1 lead to disruption of the Gly-Xaa-Yaa pattern of the collagen region. Such a disordered collagen helix appears to act like a dominant feature and results in decreased circulating levels of MBL (Fig. 2). Due to linkage disequilibrium between the SNPs, seven distinct haplotypes exist in humans, four of which (YB, YC, YD and XA) dictate low serum levels (Fig. 2) (Lipscombe et al., 1995). Haplotype occurrences vary in different human populations (Lipscombe et al., 1996, Minchinton et al., 2002).

MBL in plasma exists in complex with MBL associated serine proteases (MASPs), MASP-1 (Matsushita and Fujita, 1992), MASP-2 (Thiel et al., 1997), and MASP-3 (Dahl et al., 2001) and with a smaller splice variant of MASP-2 called MAp19 (Schwaeble et al., 2002, Stover et al., 1999) or sMAP (Takahashi et al., 1999). As indicated in Fig. 1 these associated proteins are not equally distributed among the different oligomers of MBL.

The MASPs are activated when MBL binds to a fitting carbohydrate pattern. This is observed as a cleavage of the polypeptide chains of the MASPs resulting in an A and a B chain (Vorup-Jensen et al., 2000). Once cleaved, the MASPs become active enzymes. It is not yet clear what the preferred substrates are for the different MASPs. With regards to MASP-2 it is believed that complement factors C4 and C2 are the relevant substrates. Activated MASP-2 cleaves C4 20 times better than activated C1s, the other known efficient C4 cleaving enzyme, which is present in plasma in complex with C1q and C1r (C1 complex) (Rossi et al., 2001). MASP-1 has been shown to be able to cleave complement factors C3 and C2, coagulation factor XIII and fibrinogen, (Hajela et al., 2002) whereas no natural substrates are reported for MASP-3. All of the three MASPs have been shown to cleave various synthetic substrates (Presanis et al., 2004, Rossi et al., 2001). The activities of MASP-1 and MASP-2 may be inhibited by C1 esterase inhibitor (Ambrus et al., 2003, Petersen et al., 2000, Presanis et al., 2004).

The initiation of the complement system cascade leads to clearance of microorganisms via either formation of a membrane attack complex (MAC) in the bacterial membrane, opsonophagocytosis, and/or enhanced attraction/activation of inflammatory cells. As MBL interaction with cells may trigger a number of functions, several receptors for MBL were suggested but there is no concordance as to the most relevant molecules (for further details see Holmskov et al., 2003, Kishore and Reid, 2000).