Complement System (CS) Part II - Ruhr University Bochum · 3 Most basic facts about the complement...

Complement System (CS)

Part II

Lukas Menges

24.5.2016

Molecular Immunology

2

CONTENTS

▪ Most basic facts about the CS (repetition)

▪ Control of recognition

▪ Modulation of increasing

▪ Complement receptors (CRs)

▪ Small fragments C5a and C3a

▪ Regulation of membrane attacking

24.5.2017 – LUKAS MENGES – MOLECULAR IMMUNOLOGY

3

Most basic facts about the

complement system (repetition)

▪ Antibodies only detect antigens. The complement system is one system which

complements the antibody function

▪ 3 modes of action:

- complement fixation (binding and lysis of the cell membrane)

- opsonisation (stimulation of phagocytosis)

- enticement of phagocytes by triggering of inflammatory reaction

▪ CS consists of an cascade of approx. 20 zymogens (mostly serine proteases)

activating each other


4

Prevention of autoimmunity

▪ Complement activation should take

place on pathogens, not on normal

„harmless“ cells. How it is ensured?

▪ All activated zymogens bind nearby,

otherwise they are rapidly inactivated

by hydrolysis

▪ Complement regulatory proteins

protect endogenous cells from

complement activation on their

surfaces


5 24.5.2017 – LUKAS MENGES – MOLECULAR IMMUNOLOGY

▪ Complement activation is regulated by

proteins that serve to protect cells from

accidental damage

▪ These proteins act on different stages of

the complement cascade, dissociating

complexes or catalyzing the enzymatic

degradation of covalently bound

complement proteins

Prevention of autoimmunity

Regulation of

recognition

6

Recognition unit

▪ The recognition unit C1 consits of C1q, C1r and

C1s and binds to the Fc terms of IgM and IgG.

Binding is only possible if IgM and IgG have

bound to antigens.

▪ C1q is composed of 18 polypeptide chains

(A6B6C6) forming 6 triple bundles (“bunch of

tulips”)

▪ At least two C1q subunits must be linked to

immunoglobulins to activate C1 („lower limit“)

Prevention that one unspecific antibody

activates CS

▪ Two IgGs or one IgM activate(s) C1. Why?!


EM-p

icture

7

Stage at which complement activity is regulated


C1q binding to antibodies: C1r

and C1s are activated

C1 inhibitor (C1NH)

dissociates C1r and C1s from

the active C1 complex

C1r and C1s cannot propagate the complement cascade

8

Regulation of increasing


9

Important role of C3

▪ Formation of C3 convertase: All pathways of complement

activation converge.

▪ C3 most abundant complement protein in plasma (1.2 mg/ml)

Approx. 100 proteins in plasma, abundance: 60-80 mg/ml

▪ C3 is cleaved into C3b and C3a. C3b binds to molecules on the

pathogen surface

▪ C3b forms a covalently bonded coat that can signal the ultimate

destruction of the pathogen by phagocytes (macrophages,…).


10

Complement receptors (CRs)

▪ CRs bind pathogens opsonized with

complement components

▪ C3b is an opsonin

▪ C3b receptor is called CR1, specific to C3b

▪ CR1 promotes C3b decay. Stimulates

phagocytosis (requires C5a)


+ C5a

Receptor Specificity Functions Cell types

CR1 (CD35) C3b, iC4b Promotes C3b andC4b decay. Stimulates

phagocytosis(requires C5a)

Erythrocytetransport of

immune complexes

Erythrocytes, macrophages,

monocytes, polymorphonuclearleucocytes, B cells, FDC (Dendritic cell)

CR2 (CD21) C3d, iC3b, Epstein-Barr virus

Part of B-cell co-receptor

B cells, FDC

CR3 (Mac-1) iC3b Stimulatesphagocytosis

Macrophages, monocytes,

polymorphonuclearleucocytes, FDC

CR4 iC3b Stimulatesphagocytosis

Macrophages, monocytes,

polymorphonuclearleucocytes,

dendritic cells

CRIg C3b, iC3b Phagocytosis ofcirculatingpathogens

Tissue-resident macrophages

Hepatitic sinusoidmacrophages

C5a C5a Binding of C5a activates G protein

Endothelial cells,Mast cells,phagocytes

C3a C3a Binding of C3a activates G protein

Endothelial cells,Mast cells,phagocytes

11



C4b2a is the active C3

convertase, cleaving C3 into

C3a and C3b

DAF, C4BP and CR1 displace

C2a from the C4b2a complex,

C4b bound by C4BP, MCP or

CR1 is cleaved by a soluble

protease I to inactive forms

C4d and C4c

Destruction of the C3 convertase

12

C3b is another target of regulation

▪ 1. Competition. Decay-accelerating factor DAF,

membrane-attached protein, competes with Bb for

binding to C3b on the cell surface - preventing

formation of the C3bBb C3 convertase

▪ 2. Cleaving. The plasma protease Factor I

cleaves C3b to inactive iC3b. Needs cofactors

(membrane cofactor of proteolysis = MCP or

CD46)

▪ Further C3b binding proteins: Factor H, CR1


13

Factor I deficiency

▪ Genetic disorder

▪ Uncontrolled complement activation

▪ Complement proteins rapidly become depleted

because they are mostly active on “wrong” cells

▪ Symptoms: Repeated bacterial infections


14

A closer look at the pathway…increase


15

A closer look at the pathway

▪ Generation of the C5 convertases

▪ Classical pathway: C4b2a3b

▪ Alternative pathway: C3b2Bb

▪ From different pathways, but same function

▪ C5 is cleaved into C5a and C5b. What is the small

fragment?

▪ C5b and C5a propagate the complement cascade


16



The C5 convertases cleave C5

into C5a and C5b

CR1 and H displace C3b from

the convertase. CR1 and H act

as cofactors in the cleavage of

C3b by I

Destruction of the C5 convertase

17

A closer look at the pathway…C3a and C5a


Small fragmentswith a big effect

18

The small complement fragments C5a and C3a

▪ C5a and C3a produce local inflammatory

responses by acting directly on local blood

vessels, stimulating an increase in blood flow,

increased vascular permeability and increased

binding of phagocytes to endothelial cells.

▪ C5a also activates mast cells to release mediators,

such as histamine that contributes to the

inflammatory response


enticement of phagocytes by triggering of

inflammatory reaction

Higher permeabilityof bloodvessels

Infiltrates get intothe tissue

C5a is more activethan C3a, which ismore active than C4a

http://www.linguee.de/englisch-deutsch/uebersetzung/permeability.html

19

Regulation of membrane-attacking


20

Membrane attack complex (MAC)

▪ C5b is the cleavage product from the C5-

convertase and binds to C6 and C7

▪ The C5b/C6/C7-complex (amphiphilic) binds to

C8 and to (up to 18) C9-molecules forming a

tube in the membrane

▪ The MAC is a pore with a diameter of 3-10 nm

▪ Cell loses electrolytes but no macromolecules

(interior of the cell is hypertonic). Thereby the

cell takes water and bursts („osmotic lysis“ or

„colloidal osmotic lysis“)

▪ One single MAC is sufficient for killing a cell


EM-p

icture

21



The terminal components of

complements form a

membrane pore: The

membrane attack complex

CD59 prevents final assembly

of the membrane-attack

complex at the C8 to C9 stage

The terminal complement proteins

cannot polymerize to form a pore

in membranes that kill pathogens

22

Knowledge about CS helps us to develop immunotherapies

▪ The initiating Cells of the diffuse large B Cell Lymphoma (DLBCL) arewhite blood cells (B-lymphocytes)

▪ The B-lymphocyte-antigen CD20 is a glycosylated phosphoprotein, whichis expressed on the membranesurface of B-cells.

▪ CD20 is a target of monoclonalantibodies due to cure B-cell-lymphoma and leukemia with theaid of CS


https://www.youtube.com/watch?v=sINGPr6buvw

23

Take home messages

▪ Complement activation must be regulated to

protect cells from accidental damage

▪ Regulation by “lower limitation”, regulating

proteins and rapid inactivation

▪ C3b receptor called: CR1 (effects phagocytosis)

▪ The small complement fragments C5a and C3a

produce local inflammatory responses

▪ The membrane attacking complex (MAC) forms

a pore in the membrane that kill pathogens


24

References

▪ Janeway, 2009, pp. 59-71

▪ Lecture: Biochemistry III, part 15, WS2015, Michael Hollmann

▪ Voeth & Voeth, 1995, figs 34.41

▪ Alberts et al., 1994, fig. 23.27, 23.28, 23.29

▪ https://www.youtube.com/watch?v=sINGPr6buvw, access 20.5.2017


https://www.youtube.com/watch?v=sINGPr6buvw

Thank you for attention!

Any questions left?

Complement System (CS) Part II - Ruhr University Bochum · 3 Most basic facts about the complement...

Documents

Transcript of Complement System (CS) Part II - Ruhr University Bochum · 3 Most basic facts about the complement...