Natural Killer Cells

Question 1

What distinguishes NK cells from other leukocytes?

Answer 1

They express specific markers such as CD56 and NKRP46.

Unlike B and T lymphocytes, they don’t have somatically rearranged receptors but instead a variety of germline-encoded receptors.

Generally they recognise transformed or infected cells and release lytic granules that kill virus infected cells.

So they are used for immunotherapy and treatment of leukemia; using activaed, tranformed NK cells into large granular lymphocytes and injected back into the blood.

Question 2

Describe the origin and activation of NK cells.

Answer 2

They are lymphoid cells originating from bone marrow.

They are activated with IL2 which triggers development into potent killer cells.

Question 3

Describe NK activity control

Answer 3

Activity is controlled by signals from activating and inhibitory receptors on long cytoplasmic tail.

Integration of all inhibitory AND activating signals determines whether the NK will/will not kill target cells.

Question 4

What secretion by NK cells is important in its effect on the immune response?

Answer 4

They secrete IFN-gamma.

Question 5

Describe activating receptors

Answer 5

Activating receptor recruit kinases through assiocated immunoreceptor tyrosine based activating motif (ITAM)

Question 6

Describe inhibitory receptors?

Answer 6

Inhibitory receptors recruit phosphatases through the immunoreceptor tyrosine bases inhibiting motifs (ITIM) located on long cytoplasmic tails

These interact with Major histocompatibility complex class I (MHC 1) ligands that normally present to CD8+ cytotoxic T cells. Therefore if cells escapes CD8+ T cells it can be targeted by NK cell induced cytolysis

Question 7

Describe the origin and activation of NK cells.

Answer 7

They are lymphoid cells originating from bone marrow.

They are classed as lymphoid, rather than myeloid as there is more properties in common with lymphocytes ans well as a common progenitor.

NK T cell has a T cell receptor; NK cells do not have a T cell receptor

Question 8

Describe NK activity control

Answer 8

They are activated with IL2 which triggers development into potent killer cells.

Activity is controlled by signals from activating and inhibitory receptors on long cytoplasmic tail.

Integration of all inhibitory AND activating signals determines whether the NK will/will not kill target cells.

This is a balancing mechanism that prevent over-activation of immune system.

Question 9

What is the Missing self hypothesis?

Answer 9

Where the MHC class 1 which binds antigenic peptides presented 
In absence of MHC1 is critical for allowing NK cells to become activated. Inhibitory receptors have a role in this pathway.

Question 10

Describe NK activity control

Answer 10

They are activated with IL2 which triggers development into potent killer cells.

Activity is controlled by signals from activating and inhibitory receptors on long cytoplasmic tail.

Integration of all inhibitory AND activating signals determines whether the NK will/will not kill target cells; both signals are operating at the same time

This is a balancing mechanism that prevent over-activation of immune system.

Question 11

What secretion by NK cells is important in its effect on the immune response?

Answer 11

They secrete IFN-gamma which has a role in immunoregulation.

Question 12

Describe activating receptors

Answer 12

Activating receptor recruit kinases through assiocated immunoreceptor tyrosine based activating motif (ITAM). If activated it kills cells.

Question 13

Describe inhibitory receptors?

Answer 13

Inhibitory receptors recruit phosphatases through the immunoreceptor tyrosine bases inhibiting motifs (ITIM) located on long cytoplasmic tails.

Phosphatase removes kinase Pi and blocks ability for NK to be switched on.

MHC1 ligands for inhibitory receptors; MHC1 is expressed on every cell of body except redblood cells.

These interact with Major histocompatibility complex class I (MHC 1) ligands that normally present antigen to CD8+ cytotoxic T cells. Therefore if cells escapes CD8+ T cells it can be targeted by NK cell induced cytolysis; NK CELLS second step of defense.

Question 14

What is the Missing self hypothesis?

Answer 14

Where the MHC class 1 which presents antigenic peptides identifies cells as self.

In absence of MHC1 (self) is critical for allowing NK cells to become activated. Inhibitory receptors have a role in this pathway.

Inhibition of MHC1 is a common immunoevasion mechanism of viruses (HIV) and cancer cells. Cancer mutations are selected for and variants that lose MHC1 evade immune system and are selected for.

Question 15

What are the dominant resident lymphocyte in liver?

Answer 15

NK cells.

Question 16

What two contexts are the NK cell important for the immune system?

Answer 16

1) recognition of transformed cancer cell

2) recognition of pathogens

Question 17

What method can NK cells use to kill?

Answer 17

1) release of toxic granules containing protein inducing death
2) DEATH receptors on cell surface that ligate to other ligands on target cell and induce apoptosis.

Question 18

Give clinical use

Answer 18

they are used for immunotherapy and treatment of leukemia; using activated, tranformed NK cells into large granular lymphocytes and injected back into the blood.

Question 19

What is the Missing self hypothesis?

Answer 19

Where the MHC class 1 which presents antigenic peptides identifies cells as self.

Inhibition of MHC1 is a common immunoevasion mechanism of viruses (HIV, herpes) and cancer cells. Cancer mutations are selected for and variants that lose MHC1 evade immune system and are selected for. However these cells are susceptible to NK cells.

In absence of MHC1 (self) is critical for allowing NK cells to become activated. Inhibitory receptors have a role in this pathway.

Question 20

What impact does missing self have on NK cells?

Answer 20

Loss of MHC1 removes inhibitory signals provided by MHC1 inhibitory receptors on Antigen presenting cell (KIR-killer inhibitory receptor)

NK now detect and eliminate infected cells with ligands for activating receptors on NK cell.

Question 21

Describe an experiment involving loss of self

Answer 21

1) Tumour cells with antigen presented, cell with no MHC1 variant, and cell with MHC1 variant transfected with MHC gene.
2) Measure tumour growth in comaprison to mice with no T cell (nude mouse): where antigen is diplayed on MHC tumour mass decreases in relation to nude mice. Where there is no MHC and no antigen presented, normal mouse with T cell tumour increases but nude mouse with NK cells, the tumour decreases. Where cells are transfected with MHC gene, CD8+ and tumour decrease in normal cells is restored
3) Measure killing by CD8+ and NK cells; where no antigen presented, percentage of cells killed by NK increases.

Question 22

What impact does missing self have on NK cells?

Answer 22

Loss of MHC1 removes inhibitory signals provided by MHC1 inhibitory receptors on NK (KIR-killer inhibitory receptor)

However, it still needs to be activated.

NK now detect and eliminate infected cells with ligands on pathogen activating receptors on NK cell.

Activated NK cell releases granule content, inducing apoptosis in target; this is called a cytotoxic signal.

Question 23

What method can NK cells use to kill?

Answer 23

1) release of toxic granules containing PERFORIN and GRANZYMES inducing death.

Perofin makes holes in membrane causing cell lysis through osmosis. Granzymes are protease in granule that enter target cells via the perforin pores made triggering apoptosis via other enzymes like caspases.

2) DEATH receptors such as FasL on NKcell surface that ligate to other ligands on target cell and induce apoptosis.

One NK cell can kill multiple targets.

Question 24

What stains target cell?

Answer 24

Propidium iodide (red) binds DNA once cell is dead.

Question 25

Describe the sequence of events in Virus infected cells and the benefits of IFN in target cell.

Answer 25

Virus enters cell, triggers type 1 interferon response (IFN-alpha, beta, TNF-alpha, IL12) these increase ability of NK to kill early on. Later T cell kills too, once NK killing has decreased.

IFN results

i) induce resistance to viral replication in all cells
ii) increase expression of ligands for receptors on NK cells
iii) activate NK cell to kill

Question 26

How do NK receptors show convergent evolution.

Answer 26

mice have evolved to have more Lectin receptors and humans immunoglobuin receptors. They both have same function of mediating activation

Two

Question 27

How do NK receptors show convergent evolution.

Answer 27

mice have evolved to have more Lectin receptors and humans immunoglobuin receptors. They both have same function of mediating activation or inactivation but recognise through different mechanisms.

Two different protein types have come to recognise MHC1.

Question 28

Give some examples of inhibitory receptor

Answer 28

In total, around 10 of these receptor types involved in binding MHC; where 2D= two domains, L = long motifs needed. 
These bind to different MHC 1 alleles.
 -CD94
-KIR2DL1/2/3
-KIR3DL1/2
-LILRB1

Question 29

CD94:NKG2a

Answer 29

CD94:NKG2a binds to MHC class1 that normally doesnt mediate T cell recognition.

Question 30

Describe the significance of CD94:NKG2a inhibitory receptor

Answer 30

CD94:NKG2a binds to MHC class1 that normally doesn’t mediate T cell recognition.

Polypeptide produces containing a leader peptide to direct protein to ER and secretion. The leader peptide must be cleaved off as it inserts into membrane.

When HLA MHC1 molecule is transferred into ER its leader peptide is cleaved and it binds to another MHC1 molecule HLE. This is means HLE can be presented at cell surface with leader peptide from HLA.

The CD94 recognises this complex and acts as an indirect sensor for HLA.

NK inhibitory receptor expression, in bone marrow the precursor matures into NK it expresses KIR and CD94. Cells expression of CD94 is turned off if MHC1 HLE is absent is turned off; other MHC1 molecules used in cells and CD94 is not needed.

The receptor means despite the different combinations of receptor on individual NK cells, there is at least one inhibitory receptor that can recognise MHC class1 indirectly or directly.

Question 31

Give an important activating receptor expressed on NK cells

Answer 31

NKG2D:

Question 32

Describe gene clusters and receptors

Answer 32

Both inhibitory and activating receptors are present in gene clusers; in humans, gene clusters on chromosome 12 (NK complex) and 19 (Leukocyte receptor complex) that contain different families of NK recptors (lectin, Ig)

Question 33

Give an important activating receptor expressed on NK cells

Answer 33

NKG2D

Its ligands are structured like MHC but have a different function. They are present on cells that are stressed:

• MIC-A,
• MIC-B,
• RAET-1 family of proteins.

Question 34

Describe gene clusters and receptors

Answer 34

Both inhibitory and activating receptors are present in gene clusers; in humans, gene clusters on chromosome 12 (NK complex) and 19 (Leukocyte receptor complex) that contain different families of NK receptors (lectin, Ig)

Question 35

Describe experiment of NK activating receptor

Answer 35

NKG2D activating receptor ligand Rae1 is on cancer cells which makes cancer cells susceptible to NK killing via NKG2D receptor.

Cells under stress (ionization, UV ect) upregulate Rae1.

Question 36

List the NK activating receptors

Answer 36

C type lectin: NKG2D with adaptor protein DAP10 triggering activation

Ig family: NKp30, NKp44, NKp46.

Common adaptor proteins with ITAM motif that form dimers with receptor. Activation triggered through the ITAM motif.

Question 37

Which activating receptor triggers NK activation upon recognition of Influenza haemagglutinin on flu cells?

Answer 37

NKp46

Question 38

List the NK activating receptors

Answer 38

C type lectin: NKG2D with adaptor protein DAP10 triggering activation

Ig family: NKp30, NKp44, NKp46.

Common adaptor proteins with ITAM motif that form dimers with receptor. Activation triggered through the ITAM motif.

Activation signals from
-Fc-gammaR3/4 which asscioates with ITAM gamma adaptors.

Mast cells, Basophils have
-FcERI which binds to IgE (produced in an allergic reaction) and cross links cause mast cells to degranuate and lead to asthma.

Question 39

Describe NK activity control

Answer 39

They are activated with IL2 which triggers development into potent killer cells.

Killing Activity is controlled by signals from activating and inhibitory receptors on long cytoplasmic tail.

Integration of all inhibitory AND activating signals determines whether the NK will/will not kill target cells; both signals are operating at the same time

This is a balancing mechanism that prevent over-activation of immune system.

Question 40

How can NK killer cells interact with antibodies/

Answer 40

arget cell covered in antibodies. Antibodies ligate with Fc-gammaR3/CD16, leading to ITAM activation signalling.
Releases granules and cell death.

Question 41

Describe a therapeutic antibody treatment used for arthritis

Answer 41

Rituximab: a antibody for a protein found on B cells called CD20. Aim to kill B cell and stop over production of antibody (autoimmune.) Antibody for CD20 on B cell ligates to NK cell Fc-gammaR3 receptor, activates and granules kill B cell. Antibodies used to target B cell.

Question 42

Describe activating receptors

Answer 42

Activating receptor recruit kinases through assiocated immunoreceptor tyrosine based activating motif (ITAM) which triggers activation signalling (in absence of inhibitory signal from MHC1 binding of inhibitory receptor) If activated it kills cells.

These can bind to pathogen associated molecules and self ligands .

Question 43

What two contexts are the NK cell important for the immune system?

Answer 43

1) recognition of transformed cancer cell

2) recognition of pathogens or viruses

Question 44

Describe a therapeutic antibody treatment used for arthritis

Answer 44

Rituximab: a antibody for a protein found on B cells called CD20. Aim to kill B cell and stop over production of antibody (autoimmune.) Antibody for CD20 on B cell ligates to NK cell Fc-gammaRIII receptor, activates and granules kill B cell. Antibodies used to target B cell.