mammanan immunology

:

j,,,,,,,· .. ,,jf-u (recognition of an enormous variety of foreign

antigens); VDJ recombination and somatic

vs. negative

the periphery

Four fundamental themes II

II

positive selection

Some dichotomies of immunity:

III (agglutinins, complement, phagocytes, NK cells) vs. sm:~CU]IC

(antibodies, TCR)

specific immunity: cell produced antibodies) vs

UA ............ ,., ... (TCR, cytotoxic T lymphocytes)

the T (CTL, mtraceuuiar pathogens) vs,

UH"JAV!::;''''';U responses)

III

vs

III

Anatomy:

II Generative ("central") lymphoid organs: bone marrow

III peripheral ("secondary") lymphoid organs: nodes, '''In.'''',''LA,

patches (mucosal)

Basic morphology oflymphoid organs: structure is meant to optimize contact

interactions between cells (as well as antigen) to generate an response

II cortex, related to T development)

II collection, sampling, and response to antigen

consists of a cortex of primary follicles (naiveB cells) secondary .lI.V!J,"'''''''''''

or "germinal centers" (where response to antigen rnr"nT maturation, memory); 'U"'UY,AM"

plasma cells); T cells between follicles

enters subcapsular sinus, percolates

through the efferentlymphatic of node. B cells enter node

blood and traverse helper T cell zones en route to cooperation is critical for the immune response)

immunologically relevant area is white

periarteriolar lymphoid sheaths cell zone) germinal centers (B cell zone). These are surrounded by a (marginal B cells) peripheral lymphoid tissues (GI and respiratory: Peyer's patches, tonsils, appendix)

vs,

Innate Immunity: (invertebrates vertebrates) - self/non-self

protection by neutralization destruction (pathologic consequence = UU.H ...... uUJ'aUVH

Secretions and physical barriers: like mucus and skin, lysozyme (removes bacterial coats)

II Agglutinins: neutralize, opsonize, fix complement, non-

self; major examples are lectins to sugars on

binding protein) or receptors:

UU-Hl'.,", receptors: bacteria or bacterial 1111""",,,,,,,,1/",,,,

(2) Receptor: binds to f-Met

(3) CpG DNA receptor - recognizes unmethylated CpG

(4) Mannose Receptor - non-self sugars at surface

II Macrophages and other phagocytes (i:e.,

microbes either specific receptors ro host U;;:'!"'~UUULll'"

microbes

Complement: Especially alternative

complement system:

(3) mast cell activation, recruitment, and UUu.UJLu"euuvu,

clearance of immune complexes through NK cells: viruses can cause reduced expression NK cells mechanism ofCTL) can expression a non-specific manner.

cells: make natural antibodies of diversity; recognize common

polysaccharide and antigens of many bacteria LPS;

.... "'~nuJ'''u' ... A cavities.

yo T cells: recognize heat shock proteins, other common

bacterial antigens; are not restricted; some recognize

by CD 1; most commonly at epithelial

Specific Immunity: (vertebrates) self/non-self discrimination, 0"'''',,,,,,,,,,, specificity for destruction B T

II effector T cells (i.e., CTLs)

MHC molecules

and TCR) occurs as (2) the lymphocytes.

rearrangement of the rn"""" ... ,,,, "'UJ,uu.un,'VH of self-reactive

these naive cells see antigen

proliferate! acti vate

mature

Setting: Bone Marrow

1. Start hemopoetic stem cells (HSC).

2. Through various signals, send some HSCs to become erythrocytes, megakaryocytes, eosmopnus, granulocytes, monocytes.

3. Now, the leukocyte precursor (common lymphoid progenitor) is

Once a common progenitor commits to the B lineage IS """'OJU,,"'''' nril>_nri\_

5. recombination of the variable region of heavy

6. V joins to form heavy chain Transcribe.

Process RNA to splice out between VDJ and most proximal C segment

8. Translate to produce and express on surface (with surrogate

It is the product of positive selection I (as mediated by BTK, src

receptor heavy chain is properly formed. it is, signals for survival,

chain rearrangement take place.

9. If V joined out of frame (2/3 chance), or anything else went wrong on

2nd allele. alleles have been tried, enter cell death sequence.

Allelic exclusion (prohibits non-transcribed exists)

thought to mediate RAG-2 degradation to achieve

1 . Start chain rearrangement V recombines J, transcribe.

Splice out region between VJ and CK.

3. As step 9, processed RNA is non-productive, try are

chain alleles. for both K A enter

Translate and express complete molecule on surface. B

5. IgM receptor recognizes self antigen, it continues to express

undergoes K and/or A rearrangements. it exhausts possibilities, default is

16. If non self-reactive B cell is achieved (successful positive selection

allelic exclusion (prohibits generation chains).

1 alternative splicing of original heavy chain transcript, is now expressed

joins the next C segment (Co).)

18. B cell leaves bone marrow travels to periphery

Now called B

19. If stimulated by "" .. "11- .. "",, "" ... 1I- .... ""n

cytokines 5), antigen is likely foreign (due to previous steps

stimulated B cell proliferates and the progeny begin making secreted antibodies.

allows for secreted vs. membrane form (tailpiece vs transmembrane cytoplasmic

20. The cells now undergo to make e heavy chains

region). is different from alternative splicing. class it chooses depends on

receives (by T cells other cells). Ex. yields secreted

switching regions 5' of each C segment.

IgG. Isotype switching relies on

region (so no to IgD) .

. The cells undergo ,..""."hllt,,,, .. ,,,

I"> ... A .. U •• ~. centers

the number of mutations stimulated B cell, and

and (or somatic nypermutauon

zone). These are mutations V region

increases with each generation of the progeny cells survive) have BCRs and make antibodies

3

affinity to new B cells use

center zone) to make sure is recognizable (and

Some of the cells become antibody-secreting plasma cells, while others become AU,,",UAV4J

23. Non-protein antigens (polysaccharides. nucleic acids. etc.) cannot reach T cells, so

somatic and memory B responses are not possible. Instead, B

stimulate is T-cell independent.

steps 19-23.

steps 5, 6, 11

RAG-I and RAG-2) specific to

enzymes.

F or recombination 3' of V segments, ,u .... An.JLU5 recombination by inversion

looping out and deletion, recombinases

segments, 5' segments. If the RSS is on

be necessary instead by

Each recognition sequence consists of a conserved heptamer

spacer of or 23 nonconserved base

heptamer is always closer to segment it is marking (so is immediately

segments and immediately 5' J segments).

a

RSSs

Cleavage and stranded break.

between heptamer and coding sequence,

hairpin at 3' end of ,",,",'-'AUI:'.

The

and recognize RSSs

coding sequence free displaces

protein kinase extranucleotides

GENERATION OF DIVERSITY: VDJNJ

nucs.

chain-light

combos, and somatic mutation.

Once antigen binds to BCR:

Signaling is NOT cytoplasmic of BCR """.."''''' .. "..

transduced by (components of the BCR complex).

contain immunoreceptor tyrosine-based activation motifs (IT AMs).

3 acids

cytoplasmic uomams,

4

1. BCR is cross-linked, the tyrosines IT AMS of Igu are IJUIJ;:)[I.J'HUJ

(lyn, ZYn).

Syk or Btk binds to these phosphorylated sites, and gets

3. activation can trigger a number of cascades DAG, PKC, Ras, c-Fos,

be a focus the second course.

Can trigger phagocytosis of the antigen on the BCR, B acting as an

antigen fragments to T cells (CD4) via development, yields

src

kinases

Btk deficiency (Xvlinked agammaglobunemia) - yields reduction cell maturation (positive selection signaling is impaired). Similar to syk knockouts

B

Other proteins on B signaling), CD45R CD40 (forT

surface: CRI (involved of lymphocyte subset),

comp, opsonization) and CR2 (positive

(feedback II IQ.UU~';;U

req. for somatic mutation,

2

Secreted antibodies produced as a IgA-dimer

- pentamer

are formed interaction tailpieces of

via disulfide bonds. is for joining).

of alternative splicing.

monomers

a

19B,· IgGs - monomers

chains

rll~nu" rpcnnn (VH and Vd composed of variable commemenrarnv determmmg regtons

3 is to VDJ recombination,

1. form). Recognition of antigen

initiates proliferation and secretion of antibody as described

least, Igu and to generate signal. and IgD are key players

2. toxic or infectious antigenic process. Secreted antibody

prevent it from cells.

Isotype specific functions (remember isotypes come isotype

at

IS

to constant domains of Igfi l, IgG3 or

classical complement pathway ~

that are part complexes or on

Secreted IgG coats free <'uU.''''-'"'U.

receptors that recognize the Fe ofIgGl or IgG3~ FcyR. phagocytes

easily and ingest (Also C3b, a product of the complement pathway can also opsonize

C3b receptors on phagocytes.)

surface. The antibody to destroys Ex. NK cells have

Mast cells basophils have

high affinity, IgE need not be bound to an antigen (this is ADCC).

cluster these activated Fe receptors on the cell, which causes to release

synthesize leukotrienes, prostaglandins, cytokine, of which causes an Ull. ...... ULU"" .. V .• s

ex. IgG can

to

receptors on B

Using antibodies FACS staining. It

immunofluorescence histology, unmunoprecipnauon, be good to understand the basics of how these

Key cytokines:

stimulates IgG production;

stimulates

Thymocyte precursors develop bone marrow migrate to

rearrangement takes place after arriving thymus. Positive selection occurs

the Importance stromal cells thymus: ) express

mature T cells; (2) secrete thymic hormones and cytokines survival. Steps development:

HSC diffentiates oligopotent pregenitor (CD44+, CD25-); progenitor travels to

2. Progenitor commites to T lineage (CD44+, CD25+); double negative

3. recombination of f3 chain gene (requires RAG, Ku80, DNA-PK,

receptor with surrogate a (pTa)

If this recombination is productive, pre- TCR provide

6

5. negative now converts to positive (CD4+/CD8+)

6. VJ Recombine a. chain (requires RAG, etc); express membrane TCR is presented self-MlfC + self peptide from is triggered strongly (by selfMHC + self antigen)

TCR sees nothing that even closely resembles

c. recognized the MHC/peptide complex,

8. Positive selection yields a single positive cell (CD4+ it saw

selected

""-==-,,-,,,-===,_> leads to

from cell death. CD4 CDS also required Due to

presented by Leads to self-Ml-lC

TCRs recognize self-MlK' are selected.

Ne~ative selection> leads to tolerance not

cells that self-MHC are by an apoptotic mechanism-

avidity/affinity interaction between thymocyte and peptide-MHC complexes on

T

Fate of thymocyte depends on: ofTCR molecules it expresses; (2)

molecules on thymic stromal cells it contacts; (3) affinity ofTCR for peptide-MHC ""V'U~J'''''''''~'''

or CDS on thymocytes can strength of signals generated by TCR.

Basics of ITAMs on CD3 (esp. ~) are phosphoylated

recognized by syk (zap70) family kinases. Downstream yields: (

) allelic exlusion at TCR f3 locus; (3) of TCR a. rearrangement;

the above description, it is clear that T cell development and TCR rearrangement are ""U'lAUU

development and BCR (Ig) rearrangment. TCR rearrangement B

respects> no somatic (2) no maturation; (3) use constant

isotype switching as no functional association C region; (5) fewer V genes

genes - greater diversity.

Physiologic relevance: 1. Determines response to foreign

cells bearing MHC-antigen T cell reaction soluble antigen); 3. Determines

antigens stimulate. Expression of class I II largely transcriptionally

Class II differential expression on different cell types eg mononuclear phagocytes UAY .... "'~' ....

cytokines whereas B cells dendritic cells express class

Class

= B, C; Mouse = H2K,

by CD8+ T cells

• structure: a. chain + microglobulin chain

• cytosol generated ('endogenous') proteins [cytosolic space']

7

• generation of peptide-class I MHC complex is ,,",VAHU.lUV

distinguish between foreign self antigens

not

Class

=

DQ; Mouse = 1

recognition by CD4+ T

III a +(:3

• endosome/lysosome generated

(exogenous') antigens

The most of genes species! Most alleleic

Class 1 heterozygous = 6 different alleles (3 each narent

can express 0-20 different class products

of more

Each MHC can a

conformation changes'P)

of different peptides (interaction not very Sm~CIJnc:

only specific T

Other genes MHC region: genes for biosynthesis

antigen processing - for (2) genes for subunits

generating peptides from cytosolic proteins incorporated class I

class II

Forms of antigens recognized by T cells: T cells recognize mostly

proteins, nucleic acids, polysaccharides, haptens. T """'Jl.- •. U"' ....... "' ....

antigen.

Uses basic foreign antigens displayed

mechanisms

Extracellular proteins (MHC

III specialized APC' s (macrophages, B III recognition by T cells

cells)

Cytosolic proteins(MHC .

• proteins synthesized inside cells eg viral, cancer cells,

III recognition by CD8+ T cells (CTLs)

I

microglobulin synthesized on rough ER > as separate

chains. a. chain associated with chaperones and to prevent degradation

2. binds partially folded a. chain and chaperones dissociate -

associate portion (MHC I specific) for transport to ER

3. Meanwhile, cytosolic proteins are targeted

8

shaped 7 subunits, 3 active subunits) degrades

special catalytic subunits and are synthesized

when equipped these subunits, is converted a general nousekeemnz

peptides of proper residues for MHC I UUJ,UUl,~.

(transporter for antigen processing) then transports peptides from cytosol to ER,

associated with MHC I the peptide can readily associate I.

6. Peptide binds and enhances stability causing release of TAP; move Golgi

modification; transported to surface exocytic vesicles - recognition by CD8+ T

is also

Note

1. ex and f3 chains coordinately synthesized and associate with each other

associate with (folding chaperone) and

2. The invariant binds exf3 complex, releases calnexin and complex moves out

or nascent proteins ER associating newly formed class II

formed MHC molecules to specialized endosomal/lysosomal organelles where

proteolytically cleaved - thes compartments are called (class II compartment/vesicle)

3. Meanwhile, proteins have been endocytosed and digested by proteases work

environment of endosomes/lysosomes. These vesicles fuse with

the proteases digest a part of invariant chain, only

the peptide cleft of MHC

CLIP is removed by MIIC compartment; after

class II can form - peptide binding to class II MHC

.v .. ~~", .... peptide-Ml+C complexes exocytic vesicles> displayed for

1. Granulocytes, Lymphocytes, Monocytes express

(and also Psselectin) response to inflammation (TNF,

Lymphocytes to along endothelium of venules

2. Chemoattractants type of cell.)

3. Leukocytes express cells. Lymphocytes LeukotrieneB4).

whose ligand is .... '""''',.-, due to this

Emigration out of vessel

inflammed tissue.

t.eukocvte Adhesion Defects: LAD -- CD 18 (integrin) defect -- no sticking

LAD II - fucose metabolism defect -- required for - no

group antigens)

no

9

cells express TCA -4 is made by

(ligand Lselectin) so leukocytes

which activates CCR 7 only on Naive T

it

Naive T cells sticks via LFA-lIICAM-I antigens.

emigrates

to

activation. pathway;

It TNF,

• microbes (pneurnocystis, leischmaniasis);

It macrophage and neutrophil activation;

• complement and opsonizing antibodies;

activation pathway: o

• production; activates ,",v,nu'''pAUA';;)!

• eosinophil activation;

• mast degranulation; (allergies)

• suppression of macrophage activation; (antagonize

• IgG antibodies

3 phases:

Effector cell is activated has phagocytosed microbe, eg

proteins--tissue ('hypersensitivity')

response> antigens

node> T cells recognize _ .... ""_ ..

> differentiation of narve T cells

• Migration phase - T cells to site of infection> previously activated T

restimulated by . antigens > antigen-stimulated T cells stay tissues

• Effector phase - cells secrete cytokines activates macrophages to

microbes> > lead to local (Also, v ... u ...... , ....... VUWl

and macrophages: and CD28; ...... '-'· .... H/

It Sensitization phase -

naive T & APCs ,,,,11",,,,,,,,,,"11"

>

The reaction is a variant of this, where foreign protein, from

skin test), T cells activate macrophages to induce inflammation

'delay' is to effector T home to site of

Role of macrophages CMI: CD40L and T cells signal to macrophage> AU",,,""'''' expression

IIB7-2, Fe receptors, class II MHC, IL-12, Nitric oxide

• Antigen presentation and positive feedback to T cell following class II and expression

Ell Changes local tissue environment (chronic D'THc-fibrosis: PDGF, FGF)

relevant for intracellular infections, allograft rejection, tumor rejection.

to mature for lysis

Microbes (eg viruses) infect non-phagocytic cells cannot eliminated

eliminated by CD8+ cytolytic T cells (although antibodies play role

A~U,Jl.UE:> (1) antigen specific; (2) requires targets .

contact; (3)

are 'serial

- not themselves

. Induction of CTLs:

/I Pre-CTLs (prior to activation) not active cytolysis

/I 2 signals for activation:

cytosolic peptide association with MHC

2. costimulators/cytokines produced by other simultaneously activated T

Mechanisms of CTL lysis:

/I Antigen recognition and conjugate formation: TCR-peptideIMHC; CD2ILF A-3, • Activation of CTL

/I Lethal osmotic hole membrane; enter > proteases cleave

ICE >ICE-proteolytic cascade leads to activation of DNA cleaving enzymes> anootosrs

/I Release of CTL

Natural Killer Cells:

Non B/non T cells perf orin and granzymes, V.,.,'nn1'n,p

NK cells - part of innate immune system against mtracenutar microbes:

/I Secrete phagocytosed microbes

/I Lyse to

cells express receptors = tyrosine phosphatases)

Recognize self class I Replacement of self peptides cleft

reduction class I expression) release this and lead to

known second activating signal required (can be an integrin interaction).

response to secretion of cytokines: NK cells express

converts Activated (LAK)

receptors.

dose

Role ofNK: lysis

macrophages by secreting

by

system

activate

12

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