The+Complement_2023_2.pdf

The Complement Its Activation and Role in Innate Immunity Dr Felix N. Toka Professor, Veterinary Immunology & Virology Department of Biomedical Sciences Objectives of the topic Define the complement system and its role in innate immune responses Identify and understand the major activation pathways...

The Complement Its Activation and Role in Innate Immunity Dr Felix N. Toka Professor, Veterinary Immunology & Virology Department of Biomedical Sciences Objectives of the topic Define the complement system and its role in innate immune responses Identify and understand the major activation pathways of the mammalian complement system Describe the initiation of each complement system activation pathway Understand, in general terms, the arrangement of the biochemical cascade of the complement system activation Understand how complement system activation is regulated Understand the major outcomes of complement system activation and describe the mechanisms leading to them The Complement Discovered by Jules Bordet as a heat-labile component of normal serum that augmented the opsonisation of bacteria by a heat-stable component of serum (antibodies) This activity was said to “complement” the antibacterial activity of antibodies However, complement can be activated earlier in infection without antibodies What is complement? Complement is a system of proteins that is activated by the presence of pathogens It is made up of a large number of different plasma proteins that interact with one another (at least 30). Most of the protein components are inert enzymes that become active only after cleavage When activated, they interact sequentially forming a self-assembling enzymatic cascade and generating biologically active molecules important for full inflammatory response A key site for the activation of the complement is the surface of pathogens For the complement to be useful immunologically, it has to be activated There are 3 distinct pathways through which complement can be activated The 3 pathways depend on different types molecules for their initiation The 3 pathways converge to generate the same set of effector complement components There are 3 ways in which the complement system participates in protection against pathogens 1 2 3 First way of protection Some components of the complement act as chemoattractants to recruit phagocytic cells to sites of complement activation Second way of protection Generation of large amounts of activated complement proteins that bind covalently to the pathogens opsonizing them for phagocytosis Third way of protection The final components of the complement activation kill (damage) microbes by creating pores in their membranes Apart from its role in innate immunity, the complement also participates in activating the adaptive immunity: a. Opsonization by complement components allows uptake of microbes by antigen presenting cells (APC) such as dendritic cells b. B cells have complement receptors for certain complement proteins which enhance B cell response to complement coated microbes Nomenclature of complement components Native components are designated the letter C and also have a simple number designation e.g., C1, C2, C3 etc. Unfortunately, components were numbered not in numerical order but according to the order of discovery The order of discovery is C1, C4, C2,C3, C5, C6, C7, C8, C9 The products of complement cleavage are designated by addition of a lower case letter The smaller cleavage fragment is designated “a” The larger cleavage fragment is designated “b” e.g., C4 is cleaved to C4a and C4b; C4b is larger fragment An exception: for C2, the larger fragment is C2a – the one with enzymatic activity Regarding the complement components of the alternative pathway, instead of letter C and numbers, they are designated by different capital letters and are called factors e.g., Factor B or Factor D The cleavage products are designated lower case letters “a or b” e.g., larger fragment of Factor B cleavage is Bb and smaller is Ba Initiation of complement activation 1. The classical pathway is initiated by binding of C1q (first protein of the cascade) to the surface of a pathogen C1q binds to the surface of pathogens in 3 ways 1. binds directly to the surface of bacterial components (proteins, lipoteichoic acid) 2. Binds to C-reactive protein (an acute phase protein) that initially binds to phosphocholine residues on bacterial polysaccharides 3. Binds to antigen-antibody complex on pathogen surface (most effective initiation) 2. The lectin pathway is initiated by binding of carbohydrate-binding proteins to carbohydrates on the surface of pathogens The carbohydrate-binding proteins include: mannose binding lectin (MBL) – a lectin that binds mannose-containing carbohydrates on microbial surface Ficolins that bind N-acetylglucosamine on surfaces of some microbes 3. The alternative pathway can be initiated by binding of spontaneously derived plasma C3b to pathogen surfaces Overview of the complement cascade In each pathway, sequential reactions lead to generation of a protein complex called C3 convertase The C3 convertase remains bound to the surface of the pathogen Then, C3 convertase cleaves C3 to yield large amounts of C3b (the main effector molecule of the complement system) and C3a (a mediator of inflammation) The C3b molecule acts as an opsonin – binds covalently to the pathogen surface A pathogen opsonized with C3b becomes a target for destruction through phagocytosis C3b also binds C3 convertase to produce C5 convertase which converts C5 into C5a (an inflammatory peptide) and C5b C5b initiates the last event in the complement system activation The activation culminates in formation of the membrane attack complex (MAC) Activation of complement through the Classical Pathway In most domestic animal species, IgM and certain isotypes of IgG are the most potent activators of the classical complement pathway When one IgM or several IgG molecules bind to an antigen, the Fc portion of the antibody can bind the complement component C1 (C1qr2s2) The backbone of C1 is made up of C1q C1q has six globular head domains connected by linear tail domains Associated with tail domains of C1q are two copies of C1r and two copies of C1s (serine proteases) block C1 is activated when two globular heads of C1q bind two adjacent Fc regions of antibodies bound to an antigen C1q binding to antibody activates the protease activity of C1r C1r cleaves itself Active C1r then cleaves the adjacent C1s molecules, which activates the serine protease activity of C1s Activated C1s cleaves the complement protein C4 to C4a and C4b C4b covalently attaches (through reaction with hydroxyl or amine groups) to the surface of the microbe (or Ab-Ag complex) Microbe C1s, cleaves C2 into C2b and the larger C2a C2a fragment then binds C4b forming a complex C4b2a on the surface of the microbe C4b2a is the classical pathway C3 convertase Microbe The C3 convertase cleaves C3 (the most abundant complement protein) into C3a and C3b C3b, the largest fragment attaches to the surfaces of microbes Attachment of C3b to the surface of the microbe is called opsonisation – it tags the microbe for destruction by phagocytes – this achieves one goal of complement activation Microbe C3b forms a complex with C3 convertase (C4b2a) to form the classical pathway C5 convertase (C4b2a3b) C5 convertase cleaves C5 into C5a and C5b C5b remains associated with the C4b2a3b complex and plays a role in formation of the membrane attack complex (MAC) Activation of the Classical Pathway Activation of the complement through the lectin pathway It is an independent innate immune response Lectins bind particular carbohydrate (sugar) residues on pathogens MBL and Ficolins are found in plasma and can recognize microbialspecific carbohydrates on the surface of microbes MBL is a tetrameric member of the C-type lectin family and has structural similarities with C1q MBL recognizes mannose, fucose, and Nacetylglucosamine that are present in the cell walls of bacteria and fungi, as well as on some protozoa and in envelopes of some viruses When MBL binds to a microbial cell wall, conformational changes occur which promote activation of MBL-associated serine proteases (MASPs) via autocatalysis MASPs then cleave C4 and C2 to C4a and C4b, and C2a and C2b, respectively C4b complexes with C2a to create the C3 convertase (C4b2a), which is assembled on the surface of the microbe ? C4b2a cleaves C3, into C3a and C3b leading to deposition of C3b on the surface of the microbes C4b2a complexes with C3b to create C4b2a3b, a C5 convertase, which initiates the terminal membrane attack pathway Activation of the complement through the alternative pathway It does not require recognition of a microbial surface by antibodies or lectins It utilizes the nonspecific, low-level hydrolysis of C3 C3 is the most abundant complement protein and a key player in all pathways Spontaneous cleavage of C3 produces C3a and C3b components C3b binds to a microbial surface or is degraded If C3b binds to a healthy host cell membrane, it is rapidly degraded to prevent any further complement activation This degradation occurs because sialic acid residues on cell membranes promote binding of C3b to factor H (a negative regulator of complement activation) Factor H and factor I, inactivate and degrade inappropriately bound C3b Hence, on those surfaces on which complement activation is not wanted, spontaneously bound C3b is destroyed as fast as it is deposited. Microbial membranes and cell walls are typically rich in polysaccharides that do not contain sialic acid C3b bound to microbial surface cannot be degraded by Factor H and I Instead, C3b on a microbial surface binds Factor B to form C3bB complex C3bB complex is then cleaved by factor D Ba is released, leaving the active C3bBb C3bBb is the alternative C3 convertase Instead of low-level hydrolysis of C3, this alternative C3 convertase can efficiently cleave plasma C3 and create more C3b that gets deposited onto microbial surfaces Such behaviour of alternative C3 convertase (C3bBb) creates an amplification loop The alternative C3 convertase is unstable, but it is stabilized by Factor P (Properdin) Alternative C3 convertase cleaves C3 into C3a and C3b C3b complexes with the C3bBb to form C3bBb3b C3bBb3b is the alternative C5 convertase The alternative C5 convertase cleaves C5 producing C5b initiating the terminal MAC formation Terminal Membrane Attack Pathway The goal of this pathway is to construct a protein complex that makes a hole in the target membrane and thus can directly lyse and kill the microbe This protein complex is called the Membrane Attack Complex (MAC) C5 convertase (C4b2a3b) generated by the classical and lectin pathways, and C5 convertase (C3bBb3b), generated by the alternative pathway initiate formation of the MAC Regulation of complement activation Complement control proteins act to prevent the complement innate defense system from acting on inappropriate targets and also from acting in perpetuity Failure of control proteins to halt complement cascades can have devastating consequences The classical, lectin, alternative pathways can be stopped or downregulated at key points by soluble or membrane-associated complement control proteins Control proteins function to inhibit the protease activities or facilitate the degradation of activated complement complexes or convertases A summary of known complement control proteins and their function Complement control protein Location Pathway regulated Action C1 inhibitor Plasma, serum Classical and Lectin Inactivates C1r, C1s, and MASPs C4b-binding protein Plasma, serum Classical and Lectin Binds C4b; accelerates decay of the classical and lectin C3 convertase (C4b2a) Factor H Factor I Plasma, serum Alternative Binds C3b and accelerates decay Complement control protein Location Pathway regulated Action Decay-accelerating factor (CD55) Cell membrane of Alternative erythrocytes, neutrophils, lymphocytes, monocytes, platelets, and endothelial cells Accelerates decay of C3 convertase (C3bBb) Complement receptor I (CD35) Cell membrane of Classical, Lectin, erythrocytes (primates), and alternative neutrophils, lymphocytes, monocytes, and macrophages Binds C3b and C4b and inhibits alternative and classical/lectin C3 convertases Complement control protein Location Pathway regulated Membrane cofactor protein (CD46) Widely expressed on cell membrane of host cells Classical, lectin, and alternative Protectin (CD59) Predominantly expressed Terminal membrane on cell attack membrane of erythrocytes, leukocytes, and vascular endothelium Action Binds C3b and C4a and inhibits alternative and classical/lectin C3 Convertases Binds to C5b678 and prevents C9 recruitment and MAC formation An interesting fact is that venom from the cobra snake contains a C3b analogue (called cobra venom factor) that An interesting fact is that venom from the cobra snake contains can complex with factor Bb and factor P to form a stable a C3b analogue (called cobra venom factor) that can complex with factor Bb and venom factor P to form a stableBb). C3 convertase C3 convertase (cobra factor This convertase is (cobra venom factor Bb). This convertase is not recognized by the regulatory proteins factor H and I, and thus it will factor H and I, not recognized by the regulatory proteins continually hydrolyse C3 (and C5), leading to complement depletion and severe local tissue damage. and thus it will continually hydrolyse C3 (and C5), leading to complement depletion and severe local tissue damage. Rattle snake bite in a cat Severe tissue necrosis following cobra envenomation that required amputation above the knee. The person was an 11year-old boy, bitten two weeks earlier but treated only with antibiotics A summary of outcomes of complement activation Outcome Complement products Action Direct target lysis MAC Osmodysregulation and lysis of target cells Tissue inflammation C3a and C5a Activation of mast-cell degranulation leading to release of vasoactive amines (histamine and serotonin) Endothelial activation C3a and C5a Increased expression of adhesion molecules Chemotaxis C3a and C5a Promotes migration of neutrophils, eosinophils, and macrophages toward site of complement activation A summary of outcomes – cont’d Outcome Leukocyte activation Opsonization Promotion of humoral responses Immune complex clearance Complement Action products C5a (C3a and Upregulation of adhesion molecules, phagocytic C4a) receptors, and antimicrobial effectors by neutrophils and monocytes C3b and iC3b Enhancement of particle phagocytosis by macrophages and neutrophils C3dg Enhanced B-cell activation, retention of antigen complexes in B-cell follicles C3b (and iC3b) Blocking of growth and facilitation of dissociation of immune complexes; immobilization and clearance of immune complexes through interaction with CR1 on erythrocytes