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Antigen Processing and Presentation

Aya Al-Nobani Ziad Al-Nasser Tuesday, 12/7/2011


Immunity Lecture #8 Tuesday 12/7/2011 Done by: Aya al Nobani

Review of the last lecture We have been talking about the antigen recognition molecules and the last one is the major histocompatibility antigen (MHC) and by now you should be able to know its structure, function, encoding genes which are present on the short arm of chromosome #6 consisting of 3 loci; class 1 (B,A,C), class 2 (DP,DQ,DR), and class 3 which is not involved with antigen recognition (it codes for complement, cytokines molecules and some enzymes needed for steroid metabolism). Class 1 MHC antigen: is present on all the nucleated cells in our body and they are involved in antigen processing and presentation to T-cytotoxic cells (the killing process requires class 1 MHC). Then we talked about the allelic forms of the encoding genes and these alleles are inherited in a co-dominant manner, so that in class 1 we have a maximum number of 6 different alleles and a minimum number of 3 (depends on having heterozygous or homozygous forms). Class 2: is present on the surface of antigen presenting cells (APC are nucleated >> they have both class 1 & 2). Their main function is to present antigens to the T-helper cells, and regarding the alleles it has a maximum number of 6 alleles and a minimum of 3. And we talked about the binding between the non-self antigen and the self MHC in order to bind with the TCR (restriction phenomena), and the additional binding (matching) between class 1 MHC with CD8 and class 2 with CD4 for the signalling process to take place. Each one of us has a haplotype and this haplotype is like a finger print and we use this haplotype in tissue transplantation (HLA antigens, transplantation antigen) so we have to match the haplotype of the donor
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with the recipient before the transplantation in order to minimize the chance of rejection, the higher the degree of matching the better the acceptance. In auto-graft (same person) or isograft (identical twins) the matching is 100%, and in allo-graft, which is more common, it depends on the degree of matching but still we have to supress the immune response of the recipient. Also the xeno-graft (retro-graft) we take the tissue from a pig and then transplant it into the patient, and here we must use immune-suppressants. In the clinical serology lab (immunology lab), we can do many tests in order to identify the tissue type (the tissue type means the haplotype) so we identify the allelic form of the antigen for the donor and the recipient. Therere two types of tests that can be used: 1. Micro-cytotoxicity: used to identify the allelic forms of class 1 (loci A,B,C). In order to do this test, you need to have antibodies against all the allelic forms of the antigens that are known (the source of these antibodies is a woman who has been pregnant a lot of times; so that she would be exposed to her husbands antigen, or people that have had blood transfusion more than once from different people; this person develops antibodies against these antigens but this is not a problem; only RBCs are of concern rather than WBCs) and we fix these antibodies in plates (each in a separate plate) then we take part of the buffy coat from the donors blood (to take the lymphocytes) and put it in the plates and we add a complement; now if the lymphocyte had the antigen specified for that antibody it will react and the complement will be fixed and the cell is going to be lysed otherwise it will not, and we have a die (that only stains dead cells) that we add that will give a blue colour if there was a reaction (reaction leads to cell lysis). Then you see which plates reacted (had blue dots). To know the allelic forms of the donor, you are supposed to have two of each kind (in case of heterozygote) or one (in case of homozygote), and then we do the same for the recipient. * The degree of colour gives an idea about the reaction. 2. Mixed lymphocyte culture: the idea of this test that if you have two lymphocytes that are incompatible in antigens of class 2 MHC, they react against each other and they multiply and increase in number. So we bring a lymphocyte with a known class 2 MHC and we fix that cell
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in a plate, then add a lymphocyte from the person that we want to know his haplotype; if these cells are alike, then no reaction is going to take place, and vice versa. And to know if a reaction took place, we add triated thymidine (I think what he means is thymine = one of the nitrogenous bases used in DNA duplication); so if the cells multiply, thymine is going to be used and you can measure the radioactivity, if no radioactivity can be detected, that means there was no reaction. *note: you can use the Micro-cytotoxicity test to detect class 2 but the Mixed lymphocyte culture is the assigned one!!

Now lets start with chapter 10 Antigen processing and presentation Antigen processing and presentation is the sole function of the MHC, whether the antigen is going to bind with class 1 or class 2 MHC depends on the way the antigen is going to be trafficked inside the antigen presenting cell (APC). *APC for class 2 MHC: macrophages, dendritic cells, B cells (the doctor mentioned another name but it was unclear and I couldnt find it) and its used for the extracellular antigens so that the t-helper cells (CD4) will recruit the B cells (by producing cytokines) to form the antibodies in order to neutralize the antigens (e.g. toxins and bacteria) or that cytokines will induce the phagocytic cells to kill the bacteria. *APC for class 1 MHC: cells that are infected with a virus or a tumor cell (the tumor antigen is a modified antigen intracellularly) and its used for the intracellular antigens (cytosolic derived antigens) so that the cytotoxic cells (CD 8) will be activated to kill the infected cell or the tumor cells. The doctor said that the viral antigen is formed within the cell.

Remember the MHC restriction: it means that the TCR does not
recognise the antigen unless it was presented by MHC.

*Endocytic(extracellular) pathway
Extracellular antigen will enter a vesicle by phagocytosis to be in the soluble form, 95-99% of theses antigen will be killed by the phagocytosis and the rest will require the formation of antibodies (adaptive immune response), these antigen are the ones that go in the whole processing and presentation process (to be presented with MHC class 2 to activate CD4). The antigen (e.g. toxin or bacteria) will enter the APC either by binding to the surface or if antibodies will facilitate it by opsonisation to form the phagosome (where the pH is low) then the lysosome will bind to form the phagolysosome and then the toxins or the bacteria will be degraded into simple polypeptides (by enzymes like endosomal proteases) and here comes the role of class 2 MHC which is formed in the endoplasmic reticulum and processed through the Golgi apparatus then it will be covered with a protective chain which is called the invariant chain, this chain will protect the class 2 MHC when its not in use, but when needed a proteolytic enzyme will lyse this chain so that the MHC will be free to bind with the antigen, MHC will go into the vesicle and bind to the antigen so that when the vesicle comes in contact with the cytoplasmic membrane they will be presented on the surface. T-helper cells (CD4) will bind to the antigen MHC complex through the TCR leading to the activation of t-helper cells which are going to produce cytokines needed to continue with the immune response. The nature of the antigen is important because its going to determine the antibody which will neutralize the toxin or the cytokines that will activate the macrophages which will kill the bacteria inside like mycobacterium tuberculosis. If there was a deficiency in one of the enzymes (proteases) needed to degrade the antigen, processing will not take place and this will make the person immune-compromised.

*Cytosolic pathway
Suppose a person is infected with a virus, we know that viruses multiply inside cells and utilize the protein synthetic apparatus and make new viral proteins the cytosolic pathway will be activated. **If the viruses were killed or inactivated so that they cant multiply inside the cell they will trigger the extra-cellular pathway When the viral proteins are formed within the cell, they are going to be processed and degraded into small amino acids and then they enter the endoplasmic reticulum through the transporter antigen presentation where they will bind to MHC antigen class 1 and then they presented on the surface of the cell so that the t- cytotoxic cells will recognize and bind to them to be activated. This is one factor that is required to activate the t-cytotoxic cells the other one is the cytokines from the t-helper cells. Proteins (TAP 1 & 2) >> these proteins come from the MHC antigen class 2. TAP1 & TAP2 are transporter proteins of viral antigens (proteins) into the endoplasmic reticulum where the class 1 MHC is going to be formed. If TAP 1 &2 were missing (genetic mutation) , the antigen presentation of viruses will stop >> and the cell-mediated immunity is going to be defected (while the humeral one wont be affected). Certain viruses like Herpes virus are evasive; they can escape the cellmediated immunity by producing enzymes that will inactivate the TAP 1 & TAP2 so that the viruses cannot be transported to the area where class 1 MHC are made. The viruses then will survive because they cannot be detected by the cytotoxic cells. What really determines which pathway the antigen is going to take (the cytosolic or the extracellular) is how the antigen is going to be trafficked inside the cell not the antigen type.

Super-antigen/ T-cell activation independent of processing We talked about it before , it doesnt need processing or presentation it binds to the variable B-chain of the TCR and MHC class 2 (not in the groove) and it
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cause toxic shock syndrome because it activates massive number of lymphocytes. This no specificity is an exception rather than the rule. *B-cells have a dual function; it can act as an antigen presenting cell (they have class 2 MHC) and it can develop into plasma cells to produce antibodies. *T-helper cell activation leads to the production of cytokines and there are many kinds of cytokines, one of them is the one that activates the macrophages in order to enhance their ability of killing inside the macrophage or help in the production of antibodies. *t-cytotoxic cells: when activated, produce substances called perforin and cytolysin (their function is to kill the cell by a biochemical pathway that will convince the cell to commit suicide; apoptosis).

This table in the book lists the differences between the cytosolic, phagocytic, and endocytic pathways: (the doctor read the whole table)

** Some bacteria multiply intracellualrly like chlamydia and rickettsia; these bacteria require the activation of T-cytotoxic cells so it utilizes class1 MHC. *Some of the bacteria can evade the immune response; these are very successful pathogens. For example, mycobacterium tuberculosis that causes TB disease. The cell wall of these organism is composed of a special type of lipid called mycolic aids and this kind of lipid is extremely resistant to
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phagocytosis, so they can escape the phagolysosome into the cytosol and stay for a longer period of time, in fact the phagosome can take these bacteria to other places in our body, so how can we get rid of them? We require the macrophages to be activated which is done by the mechanism we have been talking about (cytokines from the T-helper cells). Another example is Adenoviruses (naked viruses) which interfere with the transcription of class 1 MHC, so they can stay in our body for a longer period of time. In order for the body to kill those evasive bacteria or viruses, its going to activate other mechanisms including the natural killer NK where its way of killing is completely different from that of the cytotoxic cells. Vaccination If I can get a virus and inactivate it (e.g. measles) and then inject it in our body, since they are killed, they are going to be trafficked through the extracellular pathway (phagocytic or endocytic pathways) and antibodies are going to develop. But if I want to have memory cells against future infections with the same virus, I require the activation of cytotoxic cells, so the antigen must be trafficked through the cytosolic pathway. In order for this to happen, I can use live-attenuated vaccines (e.g. measles, mumps, polio and rubella). These attenuated vaccines are very successful; the memory cells that they make are from the T-cytotoxic cells. However, its not always possible to attenuate viruses or to use it on human. Another way to make memory cells is to use DNA vaccines ; if you can get a DNA of a virus (if the virus has RNA instead of DNA, you can make a complimentary copy) and then through a mechanism like retro-viruses you can transfer this DNA into the cell then transcription and translation into viral protein will occur. So the trafficking here simulates the cytosolic pathway. And as we know this will activate the T-cytotoxic cells and memory cells develop. The idea of these vaccines is to change the way the virus is trafficked in order to form memory cells. And the Doctor MUST give an introduction for the next lecture

And in the next lecture we will talk more about the molecular changes that happens in order for the lymphoid cells to be activated; this is important in order for me to know what level I am going to affect in order to activate or suppress the immune response. In general, first you need a stimulus to activate the lymphocyte which is either an infection or an extracellular antigen; when it enters the body, it has to bind to receptors as we explained earlier, and then it sends signals through transmembrane cell surface receptors (many types) and these have to be occupied or crossed over in order to send the signal inside after that transduction of these signals takes place. When we talked about the CD3 and we said they have protein tyrosine kinases that get activated by the signals. After the signal passes through, it has to amplified intracellularly through biochemical pathways (secondary pathways) and hours after that youll have a response and the outcome of this response is gene transcription and then translation into proteins > cytokine production > cell division > clonal expansion > release of defensive molecules. The most important cytokine is interleukin 2 which plays a major role in the growth and activation of cells. Other interleukins play roles in isotype switching (IgM > IgG, IgM > IgE ). Also, interleukins can recruit macrophages or activate the bone marrow to produce more progenitor cells and call them into the area and so on. And we will talk about this in details in the next lecture El 7amdella 3ala el salameh :D hon nezlat ra7met rabna o 8arar el doctor ywa2ef :D Thanx to all my friends who disturbed me while I was writing the lecture :) Thanx to you my friends for being such a wonderful people who bring joy into my life

*love all , trust few , harm none *

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