IMMUNOLOGY LEC 4,5 INNATE

what are the soluble proteins of the innate immune system?

what are the soluble proteins of the adaptive immune system?

compliment

antibodies

what does compliment do?

opsonize and lyse pathogens

who discovered compliment?

how many proteins is compliment composed of?

jules bordet

>30 proteins

where is compliment secreted? what is it secreted as?

the liver

secreted as inactive ZYMOGENS

how are zymogens activated?

by sensing a pathogen or antibodies bound to pathogen (and subsequent clevage/proteolysis)

compliment in inactive vs active form nomenclature

inactive: C1, C2, C3, Factor B, etc…

active: C1a, C1b, C2a, C2b, Ba, Bb, etc…

compliment proteins are part of what family of enzymes?

proteases

how does a zymogen that has sensed a pathogen or an antibody activate into compliment?

proteolytic clevage

what proteins trigger compliment pathwyas?

PRRs that detect pathogens

what are the 3 compliment pathways?

which was first discovered?

which is activated first?

classical, lectin, alternative

classical discovered first

alternative pathway activates first

where do all three pathways converge?

C3 convertase

what are the 3 end results (effector pathways) triggered by all pathways of compliment?

inflammation, MAC, phagocytosis

compliment can generally distinguish self vs non self and parts of compliment attach to the pathogen surface for recognition by phagocytic cells. what is this process called?

opsonization

what are the 9 C proteins in order of discovery?

C1, C4, C2, C3, C5, C6, C7, C8, C9

in general C proteins are cleaved into two: a and B. in general which is smaller and bigger?

what are the roles that each plays?

a -smaller- inflammatory anaphylotoxin

B- bigger- opsonin

what 2 exceptions are there to the rule that the B unit opsonin and the a unit inflammatory?

1. in C2 the C2a is larger and C2B is smaller

2. in C1 its subunits without cleavage are named C1q, C1r, C1s- they compose C1

proteins of alt pathway, ex. factor B -nomenclature when cleaved

Bb and Ba

7 of the 9 compliment proteins act in all 3 pathways (lectin, classical, alternative)

the other 3 do not, which are these and what pathways are they involved in?

C1- classical only

C2- classical and lectin

C4- classical and lectin

PAMPs are mostly made of what macromolecule?

carbohydrates

the lectin pathway is activated by soluble carbohydrate binding proteins: ________ and _______

mannose binding lectin (MBL)

ficolins

what protease triggers the clevage and activation of compliment in the lectin pathway?

MASPs (MBL associated serine protease)

the classical pathway is initiated when what compliment protein recognizes a microbe or binds to antibodies?

C1

the alternative pathway is activated by spontaneous hydrolysis and C3 activation, what is the important term for this process?

C3 tickover

what is the most central and important step that all 3 pathways converge on in compliment activation?

C3 convertase

which compliments recruit phagocytes and promote inflammation

C3a and C5a (and C4a mentioned later)

phagocytes with CR1 receptor bind ____ and engluf and destroy pathogen by phagocytosis

C3b

“all pathogens generate” a _____ convertase that leads to formation of MAC

C5 convertase

what is an anaphylotoxin and how does it work?

C3a and C5a are primary anaphylotoxins that trigger degrnulation of cells and encourage phagocytes to move in to site of infection and promote inflammation

opsonins attach to pathogen surface by _______ bonding

covalent

clevage of C3 exposes highly reactive_______ on C3b which is able to bind hydroxyl or amino groups on pathogen surface

thioester

what is a TED

thioester containing domain

what is the C3 convertase of

lectin

classical

alternative

fluid phase

lectin- C4b2a

classical- C4b2a

alternative- C3bBb

fluid phase- C3(H2O)Bb

C5 convertase of

lectin

classical

alternative

lectin- C4b2a3b

classical- C4b2a3b

alternative- C3b(2) Bb

in which order do compliment pathways act?

1. alt

2. lectin

3. classical

C reactive protein (CRP) is an acute pahse protein of the classical pathway that is made in liver during inflammation- essentially flags stuff for compliment to see. what molecular patterns on microbes and dammaged cells does CRP recognize?

phosphocholine

a healthy host has ____ on carbohydrates that help lectin pathway understand self vs non self

sialic acid

in the lectin pathway MBL and ficolins form complexes with MASPs.

MBL binds reptitive structures such as _________ carbs

ficolin fibrogen-like domains bind __________ carbs

mannose, fucose, GlcNAc

acetylated sugar

MBL in plasma binds to 5 proteins

3 MLB assoctiated serine proteases:

2 nonenzymatic proteins:

MASP1, MASP2, MASP3

MAp19, MAp44

1 C3 convertase can cleave 1000s of C3 (T/F)

T

MASP 2 associated with MBL or ficolin cleaves ____

C4→ C4a, C4b

C2→ C2a, C2b

what is formed by C4b2a?

C3 convertase

C4b2a does what?

cleaves C3→ C3a, C3b

once C3→ C3a, C3b what happens to C3b?

binds to C3 convertase of to the surface of pathogen where C3 convertase keeps C3→ C3a, C3b (same as all other pathways)

key enzyme for initiating lectin compliment pathway

MASP-2

what is the first protein activated in the classical pathway?

C1

what domain of C1 binds to pathogen surface or indirectly to antibody? what domain does it activate?

C1q binds, activates C1r

what does C1r do?

cleave and activate C1s which then cleaves

C4→C4a+C4b

C2→C2b+C2b

the classical and lectin pathways both are the same starting at C4 cleavage→ C2 cleavage→ C3 clevage

the alternative pathway joins in at ___

C3

the lectin pathway and classical pathway are the same from the clevage of C4. how do they start differently?

lectin: MBL, ficolin and MASPs

classical: C1, C reactive protein and phosphocholine

the lectin and classical pathways are attached to cell surface from beginning (T/F)

T

how is the alternative pathway activated?

C3 is not stable in plasma and can undergo spontaneous hydrolysis - TICKOVER

alternative pathway is activated when pathogen detected (T/F)

F- alternative pathway is spontaneously activated by tickover

how is iC3 aka C3(H2O) produced?

C3+ H2O→ iC3

how does iC3 in the alternative pathway become active C3 convertase?

factor B hydrolyses iC3

factor D proteolytically cleaves factor B

iC3Bb (aka C3(H2O)Bb) version of active C3 convertase produced

iC3Bb (aka C3(H2O)Bb) functions to

cleave C3 (C3b binds pathogen surface and C3a anaphylotoxin)

the C3b that binds pathogen in alternative pathway can be made into a C3 convertase how?

C3b binding factor B

factor B is cleaved by factor D

leaves C3bBb which can cleave more C3

C3 convertase in lectin and classical pathway vs C3 convertase of the alternative pathway:

C3 convertase of lectin/ classical: C4b2a

C3 convertase of alternative pathway: C3bBb

what is the fluid phase C3 convertase vs surface bound alternative pathway C3 convertase?

fluid pahse: iC3Bb

surface bound alt pathway: C3bBb

which of the compliment pathway(s) begin in blood/ plasma and which of the pathway(s) begin on the pathogen?

alternative- start in blood/plasma

classical and lectin- start on pathogen

___________ plasma protein stabilizes the alternative convertase C3bBb to help in compliment activation and stablization

properdin (factor P)

what is properdin (factor P) made by?

neutrophils

why can properdin stabalize C3bBb on pathogen but not host?

host expresses compliment regulatory proteins and bacteria does not allowing for properdin to bind

how does the alternative pathway amplify the classical and lectin pathways?

it can form an alternative C3 convertase to help deposit more C3b molecules in addition to the ones the regular convertase deposits.

if iC3Bb doesnt land on a pathogen surface what happens?

the alternative pathway activation loop is not completed (no C3bBb made) and it rather ends with iC3Bb which becomes inactivated

what 2 membrane proteins on host cells ensure that C3b fixation does not result in host cell destruction?

DAF (decay accelerating factor)

MCP (membrane cofactor protein)

how does DAF work to protect host cell?

breakdown of the alternative C3 converatse

C3bBb→ C3b + Bb

how does MCP work to protect host cell?

binding C3b and enhance cleavage to iC3b by factor I

factor I makes: C3b into iC3b

what is the difference between iC3 and iC3b

iC3 is the precursor to iC3Bb- the active convertase in liquid phase of alternative pathway

iC3b is a deactivated version of C3b (can still function as opsonin but stops further rxn)

factor H is a (host made/ pathogen made) plasma protein

host made

what does Factor H do?

enhances factor I in MCP action as a host protective compliment regulatory system

C3b+ Factor H + Factor I→ iC3b

what does Factor H bind on cell membranes?

sialic acids (carbohydrate found on host cells)

how can pathogens mimic host to avoid being attacked by compliment system?

displaying sialic acid (thus being able to recruit factor H)

how does compliment help in phagocytosis

phagocytic cell surface receptors bind to compliment on a pathogen (opsonized pathogen) allowing recognition as a pathogen for phagocytosis

macrophages express _____ which binds C3b to recognize and phagocytosis

CR1

all compliment receptors are opsonin receptors (T/F)

F- not all compliment acts as an opsonin

CR1, the receptor for C3b does act as an opsonin receptor becuase C3b acts as an opsonin

the cleavage products of C3b (cleaved by factor I) are also recognized by compliment receptors

ex. iC3b is a ligand for ________

ex. C3dg is a ligand for ________

CR3 and CR4

CR1

which compliment protein initiates the MAC?

C5b

after C5b, what compliment protiens follow in MAC creation

C5b, C6, C7, C8, C9

what is the alternative C5 convertase?

C3b2Bb (2 units of C3b and 1 unit of Bb)

how does host cell block MAC formation?

soluble plasma proteins block insertion and assembly

surface proteins block final pore forming step

what plasma proteins block compliment insertion and assembly? at what step is this blocked?

S protein, clusterin, Factor J

at the step of C5b+C6+C7

what human cell surface proteins block pore forming? at which step is this blocked?

CD59 (also HRF)

at the step of C5b78 +C9

anaphylotoxins C3a and C5a act on blood vessels to increase vascular permeability and cell adhesion. what does this allow for?

increased fluid leakage from blood vessels- inflammation

increased migration and activity of macrophages neutrophils, lymphocytes as well as increased microbicidal activity of macrophsges

what happens when the compliment system malfunctions

• abnormal bacterial or fungal clearance

• autoimmune disorders (lupus) due to improper clearance of soluble immune complexes

• increased infection rate due to malfunctioning MAC

people with dermatitis or exema make less FAs and their skin is more prone to what commensal that can become an opporotunistic pathogen?

staph aureus (MRSA)

which compliment pathway connects the innate and adaptive immune system?

classical (recognizes antibody/ CRP by C1)

lectin pathway recognizes ______ on a pathogen

carbs

in presence of a pathogen what occurs in alternative pathway?

it is amplified

C5 convertase of the classical and lectin vs alternative pathway

classical/ lectin: C4b2a3b

alternative: C3b(2) Bb

C3b can be cleaved into many other fragments by factor I , H and MCP.

what fragments is C3b cleaved to?

C3b→ iC3b + C3f

iC3b further cleaved → C3dg, C3c

what compliment receptor binds C3dg?

CR2

what receptor is iC3b a liagnd for?

what receptor is C3dg a ligand for?

CR3, CR4

CR2

factor I is a part of what enzyme family?

serine proteases

iC3b is a functional convertase (T/F)

F

when a phagocyte PRR binds an extracellular PAMP on a macrophage what happens?

pathogen endocytosed in phagosome and then mixed with lysosome to become phagolysosome

macrophage then begins to secret cytokines to bring more inflammation and attract neutrophils

when a macrophage phagolysozome has chopped up a bacteeria, what does it do next?

kickstarts inflammation to bring more cells to infection site

compliment receptors are PRRs (T/F)

T- some compliment receptors are a part of the PRR opsonin receptor group.

C3a, C5a and C4a act on _______ with receptors C3aR, C4aR, C5aR

blood vessels (endothelial cells) to promote inflammation