UCF Immunology Exam 4 - Weigel

The process of positive and negative selection on the primary repertoire of T-cell receptors in the thymus serves what purpose?

Prevents autoimmunity by ensuring the TCR's don't recognize self-cells

Only 1-2% of the __:__ receptors get to leave the thymus and become mature naive T cells.

__:__ receptors don't go through positive and negative selection because they don't interact with MHCs.

Only 1-2% of the alpha:beta receptors get to leave the thymus and become mature naive T cells.

Gamma:delta receptors don't go through positive and negative selection because they don't interact with MHCs.

What is positive selection vs. negative selection?

Positive selection: leads to the death of immature T cells that don't react with any self-MHC I and II

Negative selection: induces death of immature T cells that do bind too strongly to self-MHC

This process ensures that TCRs can recognize self-cells, but do not bind them too strongly that they would cause autoimmunity.

What percentage of alpha:beta T-cells get to exit the thymus?

1-2%

Are other cell-surface proteins besides TCR expressed while the T-cell is developing in the thymus?

Yes; other proteins such as CD4, CD8, or the CD3 complex are present even while they are developing

What is the path through the body of a young T cell from creation to activation?

T cells are derived from bone marrow stem cells like B cells are. Then, they migrate to the thymus to develop. Once they have gone through positive and negative selection and are mature, they exit the thymus and enter secondary lymphoid organs to be activated.

In the thymus, what tissue surrounds and interacts with developing thymocytes?

The epithelial cells that line the thymus

How does a T cell move in the thymus as it develops?

The T cell moves downwards through the thymus in development. Development starts in the cortex. Majority of development occurs in the cortex. The medulla is the final step, which is negative selection, cells in the medulla are specialized for negative selection.

What cell types are found at the top/N end of the thymocyte's development? What about at the bottom/S end?

In the cortex at the beginning: immature thymocytes, cortical epithelial cells, and macrophages

In the medulla at the bottom: mature thymocytes, medullary epithelial cells, macrophages, and dendritic cells

What is the purpose of macrophages in the cortex and medulla of the thymus?

They remove thymocytes that fail to develop properly

What is DiGeorge's syndrome? What chromosome is it associated with?

DiGeorge's syndrome is a genetic condition caused by a deletion in a part of chromosome 22.

For the purpose of this class, this leads to complete failure of a thymus to develop. This means there is no T cell development. Meaning no adaptive immune system. This is a Severe combined immunodeficiency disease (SCID) like condition.

When in a human's life is the thymus fully developed?

It is fully developed before birth. It atrophies with age, which is called involution. T cell production reduces as we age as a result.

Are T cells long or short lived? Is this like or unlike B cells?

T cells are long lived and self-renewing. This is opposite of B cells.

How are thymocytes in different stages characterized? What makes a double-negative, double-positive, etc.?

Thymocytes are characterized by the coreceptor it has on its surface.

In the beginning a developing thymocyte is called double negative. In this case it has neither CD4 or CD8 on the surface.

It will then get a signal that has it put up CD4 and CD8. This will make a double positive (having both coreceptors on the surface).

The T cell will then go through positive selection and then pick one coreceptor to express, either CD4 or CD8. This is called a single positive thymocyte.

Which two coreceptors indicate an uncommitted progenitor cell? What about committed?

CD34 and CD44 tell you that it is an uncommitted cell, present when it first enters the thymus.

After it interacts with thymic stromal cells and becomes double negative and committed to the T cell lineage, CD2 and CD5 are expressed.

What does the Notch-1 receptor do?

Notch 1 exists on the thymocytes and binds to Notch ligand on the thymic epithelial cells. This releases Notch-1 to enter the nucleus and keep the cell transcribing genes for T cell development. We call this keeping Thymocytes on the T cell path.

Which TCR chain genes rearrange at about the same time and are considered to be in a "race"? Which combination typically wins?

The beta, gamma, and delta chain genes rearrange at about the same time.

Typically, the beta-chain gene productively rearranges before the gamma delta, which sends a signal to stop rearrangement of the gamma delta chains.

Conversely, if the gamma delta rearranges first, it signals the beta chain to stop rearranging.

Describe how the pre-Ta surrogate alpha chain works and what it does.

T cells have a surrogate chain, similar to how B cells have a surrogate light chain during development.

Pre-Ta is a surrogate alpha chain used to test the beta chain. It provides 4 signals.

1: Allelic exclusion happens (turn of rag1 and rag2, tag Rag 2 for destruction, and close of the DNA)

2: T cell proliferates

3: Then it will have those cells express BOTH CD4 and CD8 (This will make the thymocyte be called double positive)

4: Alpha chain rearranges (AFTER double positive cells stop dividing)

What is important about the location of the delta gene?

The delta gene locus is within the alpha chain locus. So, if the alpha chain rearranges, the delta locus is deleted.

This is part of why T cells are mostly alphabeta.

What gene segments do the alpha and beta chains contain? Which is like the Ig's light chain? The Ig's heavy chain?

The alpha chain contains V and J segments, like an Ig's light chain.

The beta chain contains V, D, and J segments, like an Ig's heavy chain.

How does the pre-Ta surrogate chain give off the 4 signals described before?

Two pre-Ta's must come together to form a superdimer

Beta chains have the change to "try out" up to __ gene rearrangements per cell. Thus, __% of T cells can make a successful beta chain rearrangement.

Beta chains have the change to "try out" up to 4 gene rearrangements per cell. Thus, 80% of T cells can make a successful beta chain rearrangement.

Ig's heavy chain cannot do this, so their success rate is only around 55%

The beta chain has a minimum of __ and a maximum of __ attempts to productively rearrange

The beta chain has a minimum of 2 and a maximum of 4 attempts to productively rearrange

What are the two checkpoints in TCR development?

1 - Checking to see if there's a functional beta chain with the pre TCR. We have our 4 signals changing the double negative thymocyte to double positive.

2 - Tests the alpha chain for a productive rearrangement. Remember it doesn't become single positive until positive selection

Which happens first, positive or negative selection? Where does each occur?

Positive happens before negative selection.

While negative selection CAN happen in the cortex, it mostly happens in the Medulla, as the medulla is specialized for negative selection

Which cell surface proteins are expressed at what times during T cell development?

RAG1/2 go on/off

CD3 complex is always on at every stage

TdT Is turned on, then turned off before it leaves

pTalpha turned off after proliferation

How long does a thymocyte have to be positively selected? What happens after it is?

A thymocyte has about 3-4 days to be positively selected.

If they do not find an MHC to recognize in about 2-3 days, it will rearrange the alpha chain. It will put up both TCRs with different alpha chains on the surface and attempt to find an MHC.

So while it technically breaks allelic exclusion, since it is so minimal with the amount that gets positively selected, we do not functionally break it.

What is central tolerance?

Tolerance to self gained in the primary lymphoid tissue

What cell type mediates negative selection?

Dendritic cells and macrophages

What is the purpose of AIRE/autoimmune regulators?

AIRE/autoimmune regulators are used by medullary thymic epithelial cells to create self peptides normally only found in the periphery.

This is another layer of negative selection that again ensures that when mature T cells are released into the blood, they won't cause autoimmunity.

What is an important transcription factor specific to Tregs?

FoxP3 is a transcription factor that is specific for Tregs. Think of Tregs as suppressing the immune system. It is a type of CD4 T cell.

What are the levels of CD4 and CD8 T cells in a patient who has AIDS?

An AIDS diagnosis is

What are the 6 kinds of effector T cells?

Cytotoxic CD8 T cells - Kill infected cells

CD4 T cells - secrete cytokines > activate other immune cells

- TH1

- TH2

- TH17

- TFH

- Treg

Does the immune system initiate the adaptive immune response at the site of infection?

No; the pathogen is captured and transported to secondary lymphoid tissues. This is mediated by dendritic cells.

Why are dendritic cells highly specialized and effective at triggering the T-cell response?

They are migratory, and have the highest concentration of B7, so they can carry a load of antigen from the site of infection to the secondary lymphoid tissue.

What is the difference between immature and mature dendritic cells? What are immature dendritic cells called?

Immature dendritic cells a.k.a. Langerhans' cells - In the periphery, are phagocytic so they can take up antigens

Mature dendritic cells - In the secondary lymphoid tissue, can interact with T cells via finger-like extensions. They are considered "activated" and are no longer phagocytic/destructive

What receptors do immature versus mature dendritic cells use?

Immature dendritic cells surface marker is DEC205, involved in receptor-mediated endocytosis.

The surface marker for mature dendritic cells is DC-SIGN.

Generally, how does a naive T cell become activated?

1. Naive T cells enter the draining lymph node via capillaries (blood) or afferent lymphatics

2. Go through HEV's into the cortex of the lymph node

3. Encounter dendritic cells and use their TCR to examine the peptide:MHC complexes on its surface

4. When the T cell discovers peptide:MHC complex it can bind, that T cell stays in the lymph, is activated, and proliferates into many effector T cells

If the T cell does not encounter their specific antigen, they leave through the HEVs, out via efferent lymphatics, and recirculate in the blood.

Which chemokines draw naive T cells into the lymph node by binding to what surface receptors on the T cell?

Receptor on the T cell - CCR7

Chemokines produced in lymphoid tissue - CCL21 and CCL19

What are the two pairs of adhesion molecules to know? What are examples of each?

Selectins (L-selectin) bind Vascular Addressins (GlyCAM-1, CD34, MadCAM-1)

Integrins (LFA-1) bind Proteins in Ig superfamily (ICAMs)

Describe how a naive T cell binds in HEVs and enters the lymph node?

1. Lymphocytes bind to the endothelium: L-selectin on T cell surface binds two vascular addressins on HEV venules' surface (CD34 and GlyCAM-1)

2. Chemokines activate LFA-1 on the lymphocyte's surface, so it can bind ICAM-1 on the endothelial cell

3. Extravasation: Tight binding allows the lymphocyte to squeeze between two epithelial cells to enter the lymph node

4. The lymphocyte follows high conc.'s of CCL19 + CCL21 within the lymph node

What Ig superfamily cell-surface molecule on a mature dendritic cell initiates lymphocyte adhesion?

The mature dendritic cell's DC-SIGN receptor binds to the T-cell's ICAM-3 receptor

What is S1P and what is its purpose?

S1P is a chemoATTRACTANT that draws out naive T and B cells that have not been activated through the efferent lymphatics.

It is suppressed if a T cell binds an antigen so that it can stay and proliferate, and not be pushed out of the lymph node.

Is the signal generated from TCR:peptide:MHC binding alone enough to activate a naive T cell?

No; a second co-stimulatory signal from B7 on the antigen presenting cell must also be present

What does B7 on the APC/dendritic cell bind to on the naive T cell?

B7 binds to CD28

What is the difference between B7 binding to CD28 vs. CTLA-4 on the T cell?

B7:CD28 is the gas pedal for proliferation

B7:CTLA-4 is the brake

Since B7 binding CD28 increases proliferation, we need a way to slow it down (If we don't, we get cancer).

So as the T cell proliferates, it will increase the expression of CTLA-4. This will bind to B7 at a higher affinity that CD28. This means that with less CD28-B7 interaction, we will slow down proliferation to a halt.

What differentiates PAPC's from other antigen-presenting cells?

PAPC's (Dendritic cells, macrophages, and B cells) have B7 co-stimulatory molecules on their surface

What chemokine do activated mature dendritic cells secrete to attract naive T cells?

CCL18

What receptor molecule is on the surface of immature dendritic cells and what is an example of an immature dendritic cell? What about for mature?

Immature dendritic cells express DEC205 on their surface, an example is Langerhans' cells.

Mature dendritic cells express DC-SIGN on their surface, an example is interdigitating reticular cells

What is the difference between c-SMAC and p-SMAC? What are important examples of each?

c-SMAC - Central portion - clustering of receptors and signaling molecules

Ex: TCR, CD4, CD8, CD28

p-SMAC - The area around c-SMAC - comprised of adhesion molecules. Allows the cells to be kept closer together so receptors in the c-SMAC can bind.

Ex: LFA-1, ICAM-1, Talin

What is the signaling cascade of a naive T cell?

1. TCR recognizes MHC and peptide, and also interacts with the coreceptor. Coreceptor brings Lck closer to the TCR leading to ITAMs getting phosphorylated

2. ZAP70 binds, and will also get phosphorylated by the same Lck

3. ZAP-70 cascade activates PLC-gamma, which cleaves PIP2 into the "second messengers" DAG and IP3

4. 3 different signaling cascades from DAG and IP3 lead to transcription factors NF-kB, NFAT, and AP-1 being expressed

5. These transcription factors lead to cell division, proliferation, and differentiation of effector T cells

What cytokine drives T cell clonal expansion upon T cell activation?

IL-2, causes a burst in cell division and acquisition of effector function.

It is autocrine (meaning the T cell makes it, excretes it, and then it further activates itself). The principal effect of co-stimulation increases IL-2 synthesis by 100 fold.

Do naive T cells have a high affinity receptor for IL-2?

Naive T cells have a low affinity receptor, while activated T cells have a high affinity receptor

We don't want a naive T cell that has not been activated by a dendritic cell to proliferate, since IL-2 is a growth factor!

What happens if B7 is not present but a TCR recognizes MHC:peptide on an APC?

It will become anergic, so in the future even if B7 is present, it cannot become activated. This is creating peripheral tolerance.

Can immunization of a protein Ag alone induce an immune response?

Not often; typically, adjuvants (products of microbial breakdown) are needed to induce co-stimulatory activity in PAPCs.

For TH1 cells, what are the

1. Cytokines that induce differentiation

2. Defining transcription factor

3. Characteristic cytokines, and

4. Function?

1. Cytokines that induce differentiation - IL-12 + IFN-gamma

2. Defining transcription factor - T-bet

3. Characteristic cytokines - IFN-gamma

4. Function - Activate macrophages, increase inflammation at infection site

For TH2 cells, what are the

1. Cytokines that induce differentiation

2. Defining transcription factor

3. Characteristic cytokines, and

4. Function?

1. Cytokines that induce differentiation - IL-12

2. Defining transcription factor - GATA3

3. Characteristic cytokines - IL-4 + IL-5

4. Function - Activate cellular and antibody response to parasites. DOES NOT lead to inflammation.

For TH17 cells, what are the

2. Defining transcription factor

3. Characteristic cytokines, and

4. Function?

2. Defining transcription factor - RORgammaT

3. Characteristic cytokines - IL-17

4. Function - Enhance neutrophil response

For TFH cells, what are the

2. Defining transcription factor, and

4. Function?

2. Defining transcription factor - Bcl-6

4. Function - Activate B cells to refine the Ab response

For Treg cells, what are the

1. Cytokines that induce differentiation

2. Defining transcription factor

3. Characteristic cytokines, and

4. Function?

1. Cytokines that induce differentiation - TGF-beta

2. Defining transcription factor - Fox3P

3. Characteristic cytokines - TGF-beta + IL-10 (down regulate immune response)

4. Function - Suppress other effector T cells

How and why do CD4 T cells help CD8 T cells activate?

Naive CD8 T cells need stronger co-stimulation than CD4 T cells.

In suboptimal conditions, CD4 T cells can help naive CD8 T cells activate. Both must be bound to the same APC, then CD4 T cells will secrete enough IL-2 so that the high-affinity IL-2 receptor on the CD8 T cell can activate.

Which effector T cells go to the site of infection, and which stay in the secondary lymphoid tissue?

CD8s, CD4 TH1 + TH17 + Treg go to the site of infection. CD8s induce apoptosis in infected host cells, and CD4s secrete cytokines.

CD4 TFH + TH2 stay in the secondary lymphoid tissue. They secrete cytokines.

How are activated effector T cells different from resting naive T cells?

1. Type of surface molecules expressed - Effector express 2-4x more adhesion molecules (CD2 + LFA-1) than naive cells. Resting naive cells have L-selectin but low levels of CD2 + LFA-1 until activated.

2. Number of surface molecules expressed

What integrin is expressed on T cell surface only after activation and is a homing receptor for inflammation?

VLA-4 binds to VCAM-1 on activated endothelial cells that occurs during inflammation. Basically this is a homing technique for inflammation. Once we have binding, there will be the process of extravasation.

What is the difference between a cytokine and a cytotoxin?

Cytokines are basically cellular messengers.

Cytotoxins are not cytokines, they don't carry a message. Cytokines kill target cells. CD8 T cells use cytotoxins to kill their target cells.

What cytokines do TH1 produce? What do they do?

TH1 produces: IFN-gamma, TNF-a, and GM-CSF (a growth factor that goes to the bone marrow and tells the bone marrow to make more neutrophils and macrophages)

Their purpose is to help macrophages to suppress intracellular infections.

What cytokines do TH2 produce? What do they do?

TH2 produces: IL-4 + IL-5

Their purpose is to help basophils, mast cells, eosinophils, and B cells respond to parasite infections.

What cytokines do TH17 produce? What do they do?

TH17 produces: IL-17

Their purpose is to enhance the neutrophil response to fungal and extracellular bacterial infections.

What does TFH do?

Their purpose is to help B cells become activated, isotype switch, and increase Ab affinity

What cytokines do Tregs produce? What do they do?

Treg produces: TGFbeta and IL-10

Their purpose is to suppress the activities of other effector T cells.

What cytokines and cytotoxins do CD8s produce? What do they do?

Cytokines: IFN-gamma

Cytotoxins: Serglycin, perforin, granzymes, and granulysin

Their purpose is to kill virus-infected cells.

What do each of the CD8 cytotoxins do?

Serglycin - acts as a scaffold protein keeping them all together.

Perforin - makes a hole into the cell for granzymes and granulysin to enter the cell.

Granzymes - major proteins causing apoptosis, with granulysin assisting.

What pathway do cytokine receptors often signal through? How does it generally work?

The JAK-STATs.

Upon cytokine signaling, JAKs phosphorylate STATs, which dimerize and then move to the nucleus to activate specific genes.

Note that TCR signaling is Lck > ITAMs > ZAP70 > PLCgamma > PIP2 > IP3 + DAG > NFkappaB, NFAT, AP-1 > IL2... Does not use JAKSTAT. JAKSTAT is for cytokine signaling only!

What are cytotoxic cells considered and how do they carry out their function?

They're considered the serial killers.

They have lytic granules with modified lysosomes and cytotoxins. They are able to release these granules extremely specifically just on the infected cell, but not on the surrounding ones.

This is done by MTOCs (microtubule organizing centers) that rearrange so that the cytotoxic granules only shoot toward that specific direction.

CD8 cells are not only serial killers, but also release cytokines such as IFN-gamma. What are some functions of IFN-gamma?

- Inhibits replication of viruses in infected cells

- Increases processing and presentation of viral antigen by MHC I molecules

- Activates macrophages in the vicinity (to get rid of dying infected cells, to help repair)

What is TH1's role with intravesicular pathogens?

Intravesicular pathogens such as tuberculosis and leprosy may hide in macrophages' phagosomes and prevent combining with lysosomes.

TH1 can activate macrophages to become more phagocytic and produce more cytotoxins, so that they can overcome intravesicular pathogens.

What is a cognate interaction?

Cognate interactions mean that the cells must recognize the same antigen. Meaning that if a B cell and an T cell are to interact, they must both recognize the same peptide.

How does TFH aid B cell recognition?

CD40L on the T cell binds to CD40 on a macrophage and B cell