MMI 188A: Midterm #2

where do T-cell precursors develop

The thymus

Where are T cells generated

stem cells in the bone marrow

What type of T cells are found in the cortex of the thymus

Immature T cells

What type of T cells are found in the Medulla

Mature T cells

Notch 1

- Found on suface membrane of T cell

- Binds to ligand thymic epithilium

- Interaction between the two is cleaved as Notch1 intracellular domain is sent to Nucleus

- Turns on expression of :

Removal of repressive transcription factors

Recruiting coactivating factors

What happens to apopotic T cells in the thymus

- Macrophages in cortex ingest apopotic T cells that failed further development

- Most T cells die in thymic cortex

What lineage are most T cells

Alpha-Beta

Alpha-Beta T cell development vs Gamma-Delta T cell development

- Commited double negative T cell progenitor has 2 choices

Gamma and delta rearrangement or B chain rearrangement

( B chain rearrangement more common b/c only 1)

2nd chance for gamma delta rearrangement or alpha chain rearrangement

alpha chain gene rearangement produces double positive CD4 and CD8 alpha-beta T cells

pre-TCR

surrogate alpha chain

placeholder after successful Beta chain rearrangement

placeholder until successful Alpha chain rearrangement

pre-TCR development

Step 1: 2 heterodimers of a B chain and PT-alpha chain form the pre-TCR

Step 2: Heterodimers form superdimer together

Step 3: Interactions with CD3 complex and Theta chain (intracellular) form a functional pre-TCR - signal for alpha chain rearrangement to take place

Step 4: After functional alpha chain rearrangement, association with complex to form T-cell receptor

B chain rearrangement

- similar to Ig heavy chain

-contain all three segments V-D-J

- 2 genes per chromosome and only 1 needed meaning 4 chances for beta chain rearrangements

alpha chain rearrangement

- similiar to Ig light chain

- contain 2/3 segments V-J

- can undergo several rearrangements due to 61 J segments

- only need one successfull rearrangement

- delta chain is lost during alpha chain rearrangement

Positive selection

- T cells must be able to work effectively with own MHC molecules

- After a double positive T cell interacts with epithelial cells in thymus it will only express CD4 or CD8

- Mature into Single Positive T Cells

negative selection

T cell receptors must not recognize self antigens from healthy cells

Central Tolerance

Removal of T cells with autoreactive TCRs in Thymus

Peripheral Tolerance

Treg Cells suppress autoreactive T cells outside of Thymus

AIRE

Autoimmune Regulator

- Induces expression of tissue specfiic genes in thymus to allow for negative selection

- Mutations in AIRE lead to autoimmunity

- autoimmunity: autoreactive T cells wont be targeted for apoptosis/phagocytosis

FOX P3

- type of transcription factor

- Expressed by T reg cells that suppress auto reactive T cells

T reg cells

- secrete transcription factor FOX P3

- Secrete suppressor cytokines IL-10, and TGF-beta

- suppress auto reactive t cells outside of thymus (peripheral tolerance)

T cell vs B cell facts

- Very few T cells in the body are specific for any one microbe

- B cells differentiate into plasma cells and send antibodies to infection

- T cells must physically go to infection site

TCRs and antigen specificity

- Each T cell has only one TCR specificity

- Diversity is from gene rearrangement in the receptor binding site

- TCR antigen binding domain is made from V-alpha and V-beta regions

CD3

- All T cells express CD3

- Associates with TCRs

- facilitates transport to the cell surface

- Allows for signal transduction due to having transmembrane domains after TCR encounters antigen

gamma-delta receptors

- gamma delta t cells can recognize antigens without MHC

- only 1-5% of t cells

- fever V and J gene segments

CD8 T cells function and ligand

- kill virus infected cells

- more effective on intracellular pathogens

- Bind to MHC class I

CD4 T cells function and ligand

- Bind to macrophages, release cytokines, induce higher capacity to kill bacteria inside

- Bind to B cells, release cytokines, drive differentiation into plasma cells to release more antibodies

- more effective on extracellular pathogens

- Bind to MHC class II

TH1 cells provide help to ________ and are a _______ type of T cell

macrophages

CD8 T cell

TH2 cells provide help _______ and are a ______type of T cell

B cells

CD4 T cell

Human Leukocyte Antigen (HLA)

Same as Major Histocompatability Complx (MHC)

Interchangeable name of MHC

HLA - Human Leukocyte Antigens

MHC class I binding domain

- CD8 co receptor binds to Alpha 3 domain of MHC class 1 heavy chain

- peptides bind to domain via their ends

- usually 8-10 aa

MCH class II binding domain

- CD4 coreceptor binds to Beta 1 domain of MHC class II

- peptide is longer than peptide binding groove and is thus held by interactions along its length

- 13-25 aa in length

Antigen processing

The intracellular degradation of proteins into peptides

Antigen presentation

loading of peptides onto MHC molecules

Endogenous pathway

Antigen processing pathway

- 1. intracellular antigen cut up into peptides via proteasome

- 2. Peptides allowed to enter ER via TAP (transporter associated w/ antigen processing)

- 3. Peptides meet and bind to MHC class I in Endoplasmic Reticulum (ER)

- 4. Bound MHC class 1 leaves via Golgi Apparatus to present antigen peptide on cell surface

Exogenous Pathway

Antigen Processing Pathway

-1. Extracellular antigen enters and placed in vesicle

-2. Peptide production contained to phagolysosome

3. MHC class II leaves Golgi Apparatus in vesicle to go meet extracellular antigen peptides

4. fuse and bind together

5. MHC class II presents antigen peptide at cell surface

TAP

transporter associated with antigen processing

- found in endogenous pathway

- allows for peptides to pass from proteasome into ER

- peptides then meet MHC class I inside ER

ERAP

trims peptides bound to MHC class I to improve binding affinity

What would a mutation on TAP cause

Bare lymphocyte syndrome

- MHC class I proteins would not present any antigens

Tapasin

- binds to MHC class I causing a conformational change that allows low affinity peptides to leave

- If a high affinity peptide binds to MHC class I then tapasin unbinds and binding domain closes

peptide loading complex

MHC class I peptide loading complex includes chaperones :

- calreticulin

-ERp57

- tapasin

MHC class II endocytic vesicles

endosomes contain proteases that are only activated after pH is lowered

proteases then cut up antigens into peptides for MHC class II

MHC class II Assembly

- while in ER invariant chain blocks peptides from binding, also directs MHC class II into endocytic vesicles

- once in vesicles, invariant chain is cleaved and CLIP is left in MHC class II to block other peptides

- HLA-DM facilitates removal of CLIP to allow for peptide binding

Invariant chain

- prevents peptide loading while in ER

- Directs MHC class II into endocytic vesicles

HLA-DM

facilitates release of CLIP, allowing peptides to bind

cross-presentation (cross-priming)

ingested antigens (typically leading to MHC class II/endogenous pathway) are transported from phagolysosome, where peptides are degraded to proteasomes ( MHC class I) can then enter the ER to bind to MHC class I and present to CD8 T cells

How does Achieving MHC Binding Diversity occur

Polygenic - Multiple genes encode MHC class I, II, and Beta proteins

Polymorphic - Multiple alleles of each gene are found in any population

Advantage of being Heterozygous at the MHC

How are new MHC alleles generated?

Interallelic conversion/ gene conversion

recombination between alleles of the same gene, typically from different parents

result is random and thus can either be beneficial or negative

Naive T lymphocytes

- T cells that have not yet been stimulated by antigen

- Recirculate through peripheral lymphoid organs

- Express L-selectin that binds to ligand on high endothelial venules

- Express TCR, but do not engage in effector fxns (cytokine production, cytolysis)

Effector T lymphocytes

- T cells that have differentiated in response to an antigen

- Perform effector fxns (cytokine production, cytolysis [cytotoxic t cells])

Immature Dendritic Cells

present in tissues

Mature dendritic cells

found in lymph nodes

What is the exit of the lymph node

efferent lymphatic vessels

High endothelial venules

entry point for T and B cells into the lymph nodes

What are the 'Main players' in binding naive t cells through to the HEV

- L-selectin (on naive T cells)

- CD34 and GlyCAM-1 (vascular addressins ) on HEVs

how do circulation naive T cells enter lymph nodes

- naive T cell enters HEV

- L-selectin from N. T cell binds to GlyCAM-1 and CD34 on HEV endothelium

- LFA-1 activated by chemokine and binds to ICAM-1

- T cell enters lymph node

Functions of T cell Integrins like LFA-1

- Adhesion of T cells to APCS

- Binding of T cells to endothelium

- Inflammatory cytokines and microbes

- Affinity of integrins and ligand is increased by chemokines and TCR ligation

CD34

vascular addressins on HEVs

bind to L-selectin

GlyCAM-1

vascular addressins on HEVs

bind to L-selectin

LFA-1

- receptor on naive T cell surface

- binds to ICAM-1,-2,-3 on HEVS endothelium

-Promotes adhesion and cytoskeletal reorganization during T cell activation

- Promotes stable TCR:MHC interaction (Anchor)

What are the 3 signals for T cell Activation

1) Antigen

-determines specificity of response

- mediated by TCR:MHC-peptide interactions

2) Co-Stimulators

- surface molecules expressed by APCs and T cells

- provide amplification

-determine type of response

3) Cytokines

- growth and differentiation factors for T cells

- provide amplification

- determine type of response

CD28

- co-stimulator

- receptor on T cell

- binds to B7 on APC's

B7

co-stimulatory molecule on APCs

"accelerator" for T cell

CD28/B7 interaction

Induce activation of PI-3 kinase and NKkappaB signal transduction pathway

promotes cell survival through upregulation of Bcl-xL

Increases production of IL-2

Promotes IL-4 production and T cell differentiation

IL-2

-Secreted by all T cells

- production increased by CD28/B7 interaction

-Stimulates growth of helper, cytotoxic, and regulatory T cells, and NK cells.

Bcl-xL

transmembrane molecule in mitochondria

anti-apoptotic protein

- activated via CD28/B7 interaction (naive T cells)

CTLA-4

Brake for T cells

CTLA4 when expressed inhibits both the activation and proliferation of T cells

CTLA4 in already active T cells, bind to B7 20x strongly than CD28

ITAM

Immunoreceptor tyrosine-based activation motif

the prongs from CD3 that go intracellularly into the T cell and send signals towards nucleus

Immunological synapse

region of contact between t cell and APC

How to TCRs and coreceptors initiate signaling within a T cell

- MHC and CD bind

- tyrosine kinase (Lck) phosphorylates ITAMS of CD3 and zeta chains from TCR

- Lck activates ZAP-70 which goes and transmits cell signal pathways

What 3 transcription factors are activated via ZAP-70

NFAT

NF kappa B

AP-1

these transcription factors activate cytokine IL-2

Why is IL-2 important?

- drives proliferation and differentiation of ACTIVATED T cells

- costimulation along with CD28 increases synthesis of IL-2 100x

Difference between Naive T cell and Activated T cell IL-2 Receptors

Naive T cells

- low affinity

- made of only gamma, beta chains

Activated T cells

- high affinity

- made of gamma, beta, and alpha chains

Antigen recognition without costimulation may result in :

CD40

- Costimulator

- expressed on APCs (B cells too)

CD40L

- Costimulator

-expressed on T cells

CD40/CD40L interaction

- expressed only on activated T cells

- Induces upregulation of B7 on APCs

- promotes IL-12 production (contributing to TH1 differentiation)

- CD40 signaling required for Ig class switching

Function of TH1 cells

activate macrophages

Cytokine of TH1 cells

IFN-gamma

Cytokine of TH2 cells

IL-4

Cytokine of TH17 cells

IL-17

Cytokine of TFH cells

IL-21

Cytokine of T reg cells

TGF-beta

TH2 cell function

activate cellular and antibody response to parasites

TH17 functions

enhance neutrophil response

TFH functions

Activate B cells to refine antibody response

Treg functions

Suppress autoreactive effector T cells via IL-10 and TGF-beta cytokines

Activation of CD8 T cells

2 methods

1. Naive CD8 T cell can be activated directly by a virus-infected dendritic cell

2. Virus infected dendritic cell induces insufficient costimulation, leads to needing to be helped by CD4 effector T cells (done via IL-2 secreted)

What key surface molecule is not expressed on activated T cells but is on naive ones

Naive T cells express L-selectin allowing them to enter lymph nodes through HEV

Activated T cells express VLA-4 enablling them to enter infected tissue

VLA-4

expressed on activated T cells

Binds to VCAM-1 on inflamed tissue in blood vessels

VCAM-1

expressed on endothelium of blood vessels in inflamed tissue

Recruits T cells from the blood by binding with VLA-4

Cytokines are

pleiotrophic - each one has multiple actions

redundant- different cytokines may have similar effects

Cytotoxins

secreted proteins used to kill target cells by apoptosis

how does cytokine receptor signaling work ?

1. Cytokine receptors bind their JAKS together

2. Cytokine binds to receptor phosphorylating JAKS

3. JAKS bind to and phosphorylate STATs

4. Phosphorylated STAT dimers enter the nucleus to start gene expression

What T cells enter circulation seeking sites of infection after activation in lymph nodes?

CD8+ CTLs and ,CD4+ TH1 cells enter the circulation seeking sites of infection

What T cells do not enter circulation after activation in lymph nodes

CD4+ T FH cells remain in secondary lymphoid tissues to help B cells become

plasma cells or memory cells

Do T cells always require costimulation ?

No

Naive T cells require costimulators (CD40/CD40L and BT/CD28 ) for activation

Activated T cells do not require costimulation MHC only is okay

CD4+ Helper T cells have what key functions

- Help B cells produce antibody and isotype switch

- Help activate CD8+ cytotoxic T cells

- Help activate APCs

- Enhance macrophage and neutrophil response

What is the lifespan of a naive T cell

up to 9 years

What is the lifespan of an activated T cell

Weeks