Bloque 6: Cellular activation Flashcards
Signals needed for immune response activation
There are three signals needed in order to have lymphocyte activation and immune response.
The first signal is the complex formed by MHC and the ANTIGEN, which is placed on the surface of antigen presenting cells.
The second signal needed is the one caused by the presence of the CO-STIMULATORY MOLECULES.
Finally, CYTOKINES are responsible for the third signal.
Antigen recognition by lymphocytes
T cell recognition requires the presence of the MHC-antigen complex, co-stimulatory molecules and cytokines (3 SIGNALS).
Naïve T cells are activated by dendritic cells, which are the main type of antigen presenting cells. In contrast, mature effector T cells can recognise any antigen presenting cell such as macrophages or B cells.
B cells recognise whole native antigens, free or cell-associated. That means that APCs are not necessary for B cell activation.
Elements required in lymphocyte signal transduction
RECEPTOR: BCR or TCR
PROTEIN KINASE AND PHOSPHATASE that are able to activate amino acids that have an hydroxide group (serine, threonine and tyrosine). Activation is produced by phosphorylation whereas deactivation takes place once the amino acid is dephosphorylated.
ADAPTOR PROTEINS that will behave as linkers between different elements.
SECOND MESSENGERS that are molecules or ions produced as a consequence of previous events. They can either diffuse or be released so that they transmit signals to other parts of the cells and evoke changes.
Signal CASCADE amplified by enzymes.
Activation of T cells after antigen recognition (characteristics and steps)
Duration and affinity of TCR-Antigen interaction determines the fate after antigen recognition.
Antigen recognition is characterised by the fact that the affinity of TCR for antigens is usually low.
It is also true that the interaction is not stable by itself and antigen dissociation is quickly produced.
Finally, low number of MHC molecules contain the specific antigens for a TCR.
All these facts make it necessary fir T cells to have several and sequential signals so that a full functional activation is produced. These different steps are:
- Conformational change introduced to the cell after TCR interaction with Antigen-MHC complex
- Recruitment of kinases that phosphorylate ITAMS (immunoreceptor tyrosine-based activation motif)
- Linking between binding proteins and phosphorylated ITAMs
- Enzyme activation (triggered in minutes)
- Second messenger production by enzymes which may either activate other enzymes or cause and activate cascades in order to have active nuclear factors that produce new genes.
Immunologic synapse
Immunologic synapse is characterised by elements occurring outside the cell and interacting with the inner part. These elements are called LIPID RAFTS and they keep together TCR, CD3, tau proteins and others and are associated with MHC complex.
After cell recognises the antigen, a conformational change is produced. By this conformational change the SMAC (supramolecular activation cluster) is formed. SMAC is divided in two regions: pSMAC and cSMAC. The cSMAC is an inner circle in which those molecules responsible for antigen recognition and signal transduction (CD3, TCR, tau…) are placed.
Outside the cSMAC, in the pSMAC (periphery supramolecular activation cluster) will be placed those molecules like integrins responsible of maintaining the interaction between the antigen and the lymphocyte.
Early events in T cell activation
- Phosphorylation of ITAM (immunoreceptor tyrosine-based activation motif) region in tau protein by Lck enzyme
- Docking of ZAP-70, phosphorylation and autophosphorylation. ZAP-70 is part of the TCR.
- Phosphorylation of LAT (adaptor protein) by ZAP-70
- Docking of adaptor enzymes (LAT) and signal activation (divergence in activation pathways: Ras-MAP pathway activation)
Adaptor proteins
Adaptor proteins are proteins with a multidomain structure that link elements from one cascade to the elements of the next signalling cascade.
These proteins are able to bind to SH2 (oncoprotein), SH3 (viral protein), Phosphotyrosine binding domain or Ph domain (cytoskelleton protein).
In T cell activation, there’s an adaptor protein called LAT. LAT connects with other adaptor proteins like SLP-76 or Grb-2 present in immunological synapse or to other enzymes like PCLgamma1 present in the activation pathway.
Once these bounds are produced, there’s a divergence in signalling pathways.
Different signalling pathways in T cells
RAS-MAP kinase pathway:
The first step in this pathway is produced by the recruitment and activation of adaptor proteins like LAT. After that, there is in interchange between GTP and GDP in Ras protein.
RAS protein activates ERK or extracellular receptor-associated kinase-member of MAP superfamily and the MAP kinase cascade is produced, leading to a downstream activation of transcriptor factors which will end in the activation of AP-1 and others.
Calcium and protein kinase C mediated signalling pathway:
This pathway begins with the recruitment and activation of PCLgamma1 enzyme. This will lead to PIP2 hydrolysis and activation.
Activated PIP2 can activate two different proteins: DAG and IP3. In one hand, DAG will activate PCK, which in turn, activate other signalling pathways. And on the other hand, IP3 increases intracellular calcium levels which leads to a cellular response
Transcription factors in T cell activation
NFAT: Necessary for cytokines expression. It can be inhibited by CsA and FK-506, which binds to and inhibit calcineurin activity (chaperon in endogenous pathway).
AP-1: It is activated by TCR-derived signals (Ras-MAP signalling pathway). It is a dimmer connected by a leucin zipper between Fos and Jun and associated to NFAT.
NFKB: It is activated by TCR-derived signals and it is essential for cytokines synthesis.
Pharmacology of transplants
In transplants the immune system must be suppressed. In order to do that, there are different drugs:
- BASILIXIMAB suppresses IL-2 receptor
- ANTIMETABOLITES are used to suppress purine synthesis, so that DNA can’t be transcribed and there’s not effector function
- CYCLOSPORINE will inhibit calcineurin, which is responsible of phosphatase activity activating T-cells
- TACROLIMUS inhibit inflammatory mediators
Biochemistry of co-stimulation and co-inhibition
CD40-L in T cell bind to CD40 on APCs and the maturation of T cells is induced.
CD28 is the principal co-stimulatory receptor for delivery of second signals for T cell activation. It binds to CD80 or CD86 (B7) molecules present in antigen presenting cells and it is involved in increasing immune response by helping in the activation of Ras-Map pathway, in the activation of NFKB transcription factor (essential for cytokine synthesis) and blocking inhibitory pathways.
The main inhibitors of immune response are CTLA-4, which inhibits PI3k-AKT and RAS-MAP pathways, and PD-1 which dephosphorylates tau chain, ZAP70 and fyn.
B-cell activation
Scr tyrosine kinase induce IG alpha and beta phosphorylation which recruits Syk and SLP-56 proteins. This will lead to the activation of two different pathways: PCL gamma 2 and Ras-MAP.
B-cell co-receptors are molecules that complement BCR. All antigens can be recognised by BCR, but depending on the structure of some proteins, they can be recognised by C3d protein of the complement system. Complement system acts as a second signal for B-cell activation by amplifying antigens that enhance immunogenicity.
Differentiation of naïve T cells into effector T cells
After antigen stimulation, activated T cells differentiate into effector cells producing the pathogen clearance and the activation of other cells.
According to their cytokine profile, we can differentiate T cells into CD4+ T cells, that interact with APCs and CD8+ T cells that will lyse target cells.
In order to produce their functions, these cells express different effector molecules: cytokines, lytic molecules, Fas, CD40L (it induces cell maturation)…
T cell proliferation
Proliferation is important as it increases the number of cells able to destroy specific pathogens. It is crucial in the case of T cells, as there are few MHC-Antigen complex.
The process of proliferation is mediated by an autocrine growth factor called cytokine.
There are many different cytokines with a wide variety of functions. However, in CD4+ T cell proliferation, the main cytokine is IL-2, whereas in CD8+ T cell proliferation, the main one is IL-15.
As a consequence of this process, few naïve T cells are able to develop into a high number of effector and specific cells for the antigen and also memory cells.
Differentiation of naïve T cells into memory cells
A proportion of stimulated T cells will become memory cells thanks to different homeostatic signals caused by interleukins.
T memory cells can be divided into two main populations: effector memory cells and central memory cells. Both of them are different in several aspects.
The first one refers to the cell markers expressed on their surface. Central memory T cells express CD44, CD62L (selectin that allows them to reach lymphoid organs) and CCR7 (chemokine that allows cells homing in lymphoid organs), whereas effector memory T cells express the same CD44, but CD62L expression is variable and there is no CCR7 expression.
Another fact distinguishing these cells is their process of activation, which is longer in central T memory cells and shorter (meaning that they are able to get rid of the antigen after detecting it) in effector T memory cells.
Finally, the last fact that differentiate both populations is their location. Effector T memory cells are circulating while central T memory cells are waiting in lymphoid organs to be needed.
Nevertheless, there are other memory T-cell populations depending on their location.
First of all, there are tissue resident memory T cells (Trm) that offer rapid protection from tissue invading pathogens and may be shorter lived compared to other memory populations.
Another population is the one formed by the least differentiated, multi-potent and self-renewing T memory cells, which is called T stem memory cells (Tscm).
The last group of T memory cells is follicular helper memory T cells (Tfh). These cells may be found near B follicles or in the blood and require antigen to maintain their phenotype. They provide rapid help for B cell germinal centre responses.
