Week 4 Lecture 6 - Complement and Disease Flashcards
Complement System - Overview
Three distinct pathways activate complement: initiated by different molecules but converge to generate a C3 convertase and the same effector molecules.
1. Classical Pathway: initiated by binding of C1q to the pathogen surface
2. Mannose-binding Lectin (MB-lectin) pathway: initiated by binding of mannose- binding lectin to carbohydrates on bacteria or viruses
3 .Alternative pathway: initiated by binding of spontaneously activated C3 in plasma to pathogen surfaces
Complement - Classical Pathway
Role in innate and adaptive immunity
Activators of the classical pathway:
- antibody-antigen complexes, or the binding of antibody to antigen on a suitable target e.g. bacteria
Globular regions of C1q bind to C1q-specific receptors on the Fc regions of either one IgM or two closely spaced IgG molecules bound to antigen
- C1q can also bind directly to pathogens and other activators -> activation in the absence of antibody
- C1q can bind to CRP protein and pentraxin-3 bound to bateria, phosphatidylserine, double stranded DNA or annexins on dead and dying cells
Classical Pathway Steps
- First component of classical pathway is the C1 complex: C1q (6 subunits, globular heads and collagen-like tails) which bind 2 molecules of C1r & C1s
- circulates in plasma as Ca++ dependent complex - Binding causes a conformational change in complex -> enzymic activity in C1r
- Activated C1r cleaves C1s -> C1s esterase
- C1s esterase cleaves C4 -> C4a (off into fluid phase) & C4b which attaches to the pathogen surface
- Pathogen attached C4b binds C2 making it susceptible to cleavage by C1s esterase -> C2b (small bit) in fluid phase and C2a, an active protease associates with C4b to give C4b2a (the C3 convertase)
Mannose Binding-Lectin Pathway
Mannose-binding lectin binds specifically to mannose residues arranged in a pattern that allows binding
- common on bacteria, viruses and dying cells
MBL circulates associated with two serine proteases MASP-1 and MASP-2
- most MBL are associated with only one of the MASP-1 proteins
Upon target binding juxtaposition of MASP-2 & MASP-1 containing MBL complexes is required for MASP-1 to activate MASP-2
MASP-1 from one complex will activate MAST-2 from an adjacent complex
Activated MASP-2 cleaves C4 and C2 - in a manner like that of the classical pathway
Alternative Pathway
Alternative pathway is the dominant pathway during normal physiological conditions
It constantly monitors for pathogen invasion -> low constitutive activation: “Tick-over”
-> Spontaneous hydrolysis of a labile thioester bond - converting C3 to its bioactive form C3(H2O)
Alternative Pathway Steps
- Formation of C3(H2O) causes exposure of the binding site for Factor B(FB)
- C3(H2O)-bound FB is then cleaved by serine protease Factor D forming → fluid phase C3 convertase, C3(H2O)Bb
- C3(H2O)Bb cleaves C3 to give C3a & C3b -> small amounts of C3b bind to close surfaces containing a hydroxyl group, e.g. proteins & carbohydrates
- Binding of factor B to C3b induces a dynamic equilibrium between a closed (loading) and open (activation) form
- leads to a marked, reorientation of the catalytic serine protease (SP) domain of factor B - Factor D circulates in blood in an inactive conformation
- Factor D binds factor B specifically in the open C3bB state, and high affinity binding of factor D to C3bB activates factor D
- Factor D cleaves factor B releasing Ba fragment
- Proteolytic fragment Bb, stays bound to C3b -> active C3bBb protease complex
- C3bBb protease complex stabilised by properdin to cleave more C3 into C3a and C3b -> amplification
C5 Convertase
C3 cleavage is the central reaction
- C3a is a fluid phase anaphylatoxin
- C3b binds pathogen surface near site of complement activation
When C3b binds either C4b2a or C3bBb it makes C4b2a3b (classical C5 convertase) or C3bBb3b (alternative pathway C5 convertase)
Cleavage of C5
- C5a, a potent anaphylatoxin
- C5b which binds to the pathogen surface to form the nucleus for the terminal complement components
Terminal Pathway
Terminal components are common to all pathways
Union of the terminal components forms the membrane attack complex (MAC) -> cell lysis
- first, C6 first binds C5b on cell surface, followed by C7 and C8 -> complex penetrates into the cell membrane. C8 is first component to penetrate the membrane
C5b-8 complex acts as receptor for C9 -> many C9 molecules interact with the complex -> polymerisation of 10-16 C9 molecules into a pore -> MAC
Complement - Regulation of Activity
Complement pathways are regulated to prevent:
1. The destruction of host cells and tissues, &
2. Uncontrolled complement activation rapidly depleting complement components leaving the host unable to remove infectious bacteria
Series of control proteins regulate the cascade at 3 points:
1. At pathway initiation
2. At C3 convertase
3. At formation of MAC
Regulation of Classical and MBL - Pathway Inhibition
A plasma serine protease inhibitor, C1 inhibitor (C1 INH) controls the activation of C1 complex
C1 INH binds C1r:C1s causing them to dissociate from C1q, limiting the time active C1s has to cleave C4 & C2
C1 INH regulates the MBL pathway by inhibiting MASP-1 & MASP-2
MBL can also bind MASP-3 – but MASP-3 cannot activate MASPs or cleave C4 or C2
Regulation of Classical Pathway - C3/C5 Convertase
C4b2a C3 convertase on a host cell will be rapidly dissociated by DAF (decay accelerating factor/CD55) &/or CR1 (cell surface complement receptor-1/CD35) depending on the cell type, releasing C2a
Bound C4b will be inactivated by Factor I (FI) in the presence of co-factors CR1 and/or MCP (membrane cofactor protein/CD46)
C4d remains bound to the membrane but C4c is released
C4 binding protein (acts in fluid phase) can interact with several C4b molecules at once - dissociating the C2a
Regulation of Alternative Pathway - C3 and C5 Convertase
MCP, DAF (CD55), Factor H (FH)and CR1 can dissociate Bb from C3bBb
C3b with co-factors: MCP, CR1 or FH, is rapidly cleaved to iC3b by FI
With Co factor CR1, FI cleaves iC3b to give C3c (released into fluid) and cell bound C3dg, which becomes C3d by cell proteases or plasmin
Serum protein Factor H (FH) competes with Factor B for binding to C3b in the fluid phase and on cell surfaces
- factor B binds C3b -> alternative pathway progresses
- factor H binds C3b -> alternative pathway does not progress
Surface of mammalian cells (presence of sialic acid) favours Factor H binding, whereas surface of bacteria favours Factor B binding
In the fluid phase Factor H binds to C3b and C3(H2O), but NOT C3
Regulation of Terminal Pathway and MAC
Binding of C5b to cell membranes is relatively non-specific - but generally occurs adjacent to site of complement activation
Clusterin and vitronectin bind C8 and prevents insertion into the membrane
CD59 (protectin) is widely distributed in mammalian cell membranes, and it inhibits the binding of C9 to the C5b678 complex -> blocks MAC formation
C5b-9 can be removed by endocytosis or exocytosis
Biological Activity of Complement - Opsonisation
Main opsonins are C3b, C4b and iC3b
Phagocytic cells express complement receptors CR1, CR3 and CR4, which have broad specificity for these opsonins
Binding of C3b to CR1 leads to phagocytosis when other immune mediators (e.g. C5a) have activated the macrophages
Cells dying by necrosis can activate complement - causing deposition of C4b or C3b on their surface
C4b or C3b bind CR1 or CR3 on phagocytic cells causing clearance of the necrotic cell
Biological Activities of Complement - Removal of Immune Complexes
Deposition of C3b on immune complexes breaks them up into pieces that are cleared by macrophages
C3b deposition also allows binding to RBC via CR1 & RBC clear the complexes by transporting them to the liver & spleen where they are stripped off the RCB and phagocytosed by macrophages
C3dg enhances B cell responses
C3dg binds Ag bound to Ig on B cell surfaces &
simultaneously binds CR2 on the B Cell
- signalling through both membrane Ig & CR2 increase activation of B cells
- lower threshold for B cell activation (1000x)
