Adaptive immune system (What we and other vertebrates have in addition to an innate immune system)

1. Adaptive immune system(What we and other vertebrates have in addition to an innate immune system) • Makes specific immune response to a pathogen • (1790s): Jenner showed that exposure to cowpox protected against infection by a related virus: smallpox • Adaptive immune response has “memory” that can confer life-long immunity to infection • 1781: Swedish sailors bring measles virus to Faeroe Islands • 1846: Another ship with measles-infected sailors visits Faeroe Islands. People older than 64 did not get measles because they had life-long immunity to measles virus.

2. Comparison of innate and adaptive immunity

3. Two arms of the adaptive immune response Humoral: antibodies (proteins produced by B cells) that circulate in the blood. Antibodies can recognize virally-infected cells and free viruses. Cellular: Lymphocytes (T cells) that do not produce a soluble product. T cells can recognize virally-infected cells, but not free virus.

4. Adaptive immune system I: Antibodies • Antibodies normally work very well to get rid of pathogens. • Antibodies don’t work well to combat HIV. • Autoimmune disorders (e.g., lupus) can occur when antibodies are made against “self” proteins • Lupus patients make anti-nucleic acid antibodies

5. Why Bi1 is useful: Knowing about antibodies, pathogens, and viruses helps when watching late night TV

6. The power of the humoral immune response • Typical mammal (e.g., you, a mouse) can make >1016 different types of antibodies. • Antibodies can bind to many types of antigens. (protein, carbohydrate, nucleic acid, lipid, small molecules). • Mice can raise antibodies against synthetic compounds that don’t exist in nature (e.g., buckyballs). • Antibodies can be exquisitely specific: e.g., can distinguish between ortho-, meta-, and para-Aminobenzoic acid.

7. Vocabulary • Antibody (Ab) or Immunoglobulin (Ig) - a Y-shaped membrane-bound or soluble protein that binds specifically to an antigen (Ag). The hypervariable regions (HV) or complementarity determining regions (CDRs) of the antibody contact the antigen. • IgG (Immunoglobulin G) is the most abundant form of antibody in the blood. • Antigen (Ag) - The target of an antibody or a T cell (can be protein or non-protein). An antibody binds to a region of the antigen called the epitope. • Epitope - The region of an antigen that is recognized by an antibody or a T cell receptor.

8. Antibodies attack foreign invaders (antigens) Fantastic Voyage, 1966

9. Antibodies -- serum proteins that combat pathogens • Typical mammal (e.g., you, a mouse) has the capacity to make >1016 different types of antibodies. ~10 nm (100 Å) Immunoglobulin G (IgG)

10. Antibodies

11. Fab Fab Antibodies (Immunoglobulins) Note two identical Fabs, so two antigen binding sites Fab = Fragment antigen binding) • 2 heavy chains, 2 light chainsVariable (V) domains (VH and VL) Constant (C) domains (CH1 and CL) Fc Fc = Fragment “constant” Fc region (nearly constant in sequence) of Abs has effector functions (bind Fc receptors, complement, etc.).

12. Generating antibody diversity Clonal selection • Niels Jerne proposed the “Clonal Selection Theory” in 1954 -- see Bi1 website link to his 1984 Nobel Prize lecture. Each cycle of cell growth and proliferation takes ~12 hours, so takes ~1 week to make clone of ~20,000 identical B cells

13. Clonal selection theory accounts for: • Diversity -- information coding for all Abs is in DNA • Self/non-self discrimination -- eliminate cell clones bearing all self-reactive receptors on their surface or else inactivate them • Memory -- increase in number of cells the second time around

14. Clicker question We need ~108 different kinds of antibodies to ensure immunity against most/all pathogens. How can an individual make 108 different types of antibodies? 1) Encode each antibody gene in the genome. 2) Separate antibodies into different segments, then mix and match gene segments. 3) Start with one antibody gene, then mutate it to create different specificities. 4) Make antibodies that have no defined structure, then they can fold around an antigen.

15. How can we make enough different antibodies to protect us from all possible pathogens? Estimate: need 108 different antibodies --> 104 heavy chains and 104 light chains Human cells have ~30,000 genes If every heavy chain and every light chain encoded by a different gene, would use up half the cell’s genes just making antibodies!

16. Generating antibody diversity: Modular design of antibodies • Heavy chain variable region encoded in 3 gene segments (modules): V, D, and J • 100s of different V gene segments • Tens of different D gene segments • >5 different Js • Light chain variable region encoded in 2 gene segments: V and J • Hundreds of different V gene segments • >5 different Js

17. Generating antibody diversity • Permanent rearrangement of the DNA in a B cell • Same thing happens in T cells to generate T cell receptors • Mature B and T cells do NOT have the same DNA as other cells in the body

18. Generation of diversity (G.O.D.) • Multiple V, D, J gene segments • Junctional diversity -- random pairing of a V and a J gene segment (light chain) or a V, D, and a J gene segment (heavy chain). • Addition or deletion of bases during joining of V to D and/or V to J and/or D to J creates even more diversity within CDR3 • Combinatorial pairing of H and L chains • Somatic hypermutation -- high rate of mutation in antibody variable region genes during clonal expansion of a B cell. Result: some B cell descendents produce antibodies that bind more tightly to an antigen (these are stimulated to divide further); others produce antibodies that don’t bind as tightly (these are not stimulated to divide further).

19. B cell development Choice to become:Plasma cell -- produce secreted form of antibody; secrete ~2000 antibodies/sec; short lifetime (days) Memory cell -- have already somatic hypermutation. Can confer life-long immunity to infection. T lymphocytes can also become long-lived memory cells. CD4 memory T cells are an important reservoir for HIV.

20. Clicker question Differential RNA splicing creates antibody diversity. 1) True 2) False

21. Clicker question Differential RNA splicing creates antibody diversity. 1) True 2) False Antibody diversity is created by differential recombination at the DNA level, NOT the RNA level.

22. Clicker question Why is differential RNA splicing not a good method for the immune system to create a diverse set of antibodies? 1) Differentially-spliced RNA transcripts would not be inherited by clonal descendents of a stimulated B cell. 2) RNA splicing is inefficient, thus not enough of the correctly spliced antibody would be produced by each cell. 3) Differential RNA splicing would not provide an adequate level of potential diversity. 4) Differential RNA splicing only occurs in different cell types. A single cell type (e.g., a B cell) always splices RNAs in the same way.

23. Variability within antibody V domains clusters in three regionsWu and Kabat index of variability:# aa that occur at that position / frequency of most common aa at that position 3 hypervariable (HV) regions (CDRs) LightHeavy 24-34 31-35 50-56 50-65 89-97 95-102 Janeway et al. Immunobiology Figure 3.6

24. Hypervariable regions fall in loops of V domain structure Crystal structuresshow that loopscontact antigen Rename themCDRs (ComplementarityDetermining Regions) CDR1 CDR2 CDR3 Figure 3.7

25. Arrangement of CDRs in Ab combining sites: CDR3s always in center, CDR1 and CDR2 always on sides CDR2 (L) CDR3 (H) CDR1 (H) CDR2 (H) CDR3 (L) CDR1 (L) Branden and Tooze, Fig. 15.13

26. Antibody variable domains bound to an antigen Hypervariable loops (CDRs) Lysozyme Antibody variable domains Hypervariable loops (CDRs) Hypervariable loops removed Antibody variable Domains Antibody variable Domains (framework) Lysozyme Lysozyme