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Restriction-modification systems in bacteria

Restriction-modification systems in bacteria. Sites on bacterial chromosome protected by proteins/methylation. G A A T T C C T T A A G. EcoRI. Sites on phage DNA/RNA unprotected and cut. Penicillin and other antibiotics.

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Restriction-modification systems in bacteria

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  1. Restriction-modification systems in bacteria Sites on bacterial chromosome protected by proteins/methylation G A A T T C C T T A A G EcoRI Sites on phage DNA/RNA unprotected and cut

  2. Penicillin and other antibiotics • Discovered by Fleming in 1929 and turned into a useful treatment by Florey and Chain (1940s) • Prevents the formation of the bacterial cell wall (by interfering with peptidoglycan cross-linking) • New cells subject to osmotic lysis • Only acts against Gram-positive bacteria • Other antibiotics • Streptomycin from bacterium Streptomyces acts on protein synthesis • Tetracycline, also from Streptomyces, also acts on protein synthesis on a wide range of bacteria

  3. The immune system of Drosophila • Body fluid of insects contains haemolin • a member of the immunoglobulin superfamily, which binds to microbes • Also several short peptides that play an unknown role in antibacterial action • Defensins – like scorpion venom taxins • Cecropins and attacins (disrupt cell membrane) • Drosomycin (antifungal) – similar to plant substances • Rapid evolution of some these proteins indicates an ongoing evolutionary arms race • Cellular immunity • Haemocytes secrete antimicrobial peptides and are involved in coagulation, phagocytosis and encapsulation

  4. Immune systems in vertebrates - overview • Mechanical barriers • Collagen/skin , mucous membranes, secretions • Non-specific defence (innate immunity) • Phagocytosis by neutrophils (70% WBC) • Macrophages (5% WBC) • The inflammatory response (histamine, chemokines, fever) • Antimicrobial proteins (complement, interferons) • Specific immune defence (adaptive immunity) • Humoral (antibodies, B cells) • Cell-mediated (Cytotoxic T cells, Helper T cells)

  5. Antigen (1st exposure) Engulfed by Free antigens directly activate Antigens displayed by infected cells activate Macrophage Stimulates B cell Helper T cell Cytotoxic T cell Stimulates Stimulates Gives rise to Gives rise to Memory helper T cell Stimulates Stimulates Antigen (2nd exposure) Memory B cells Stimulates Plasma cells Memory T cells Active cytotoxic T cells Antibodies

  6. B cells (made in bone marrow) • B cells secrete antibodies which recognise intact foreign particles outside cells and block their action/target them for destruction Infected cell B cell Antibody Antigenic determinant Each antigen can present many different antigenic determinants and be recognised by many different Antibodies Antigen

  7. The structure of antibodies

  8. The diversity of antibodies • Collectively referred to as immunoglobulins • IgM (pentamer) first to appear in response to infection • IgG (monomer) most abundant circulating form, confers maternal immunity • IgA (dimer) produced by mucous membranes • IgD (monomer) Differentiation of B cells • IgE (monomer) stimulate mast cells and histamine • Each has own heavy chain (a, d, e, g, m) • There are 2 types of light chain (k and l) • Both light and heavy chains are encoded by libraries of subunits which are shuffled to generate diversity

  9. Light-chain diversity Variable (40 units) Joining (5 units) Constant Somatic recombination with imprecise joining TGGCCAG TGGCTGGCAG TGGAG TGGCTGGAGCAGCC x ‘Fine-tuning’ by somatic hypermutation (error-prone DNA repair process) Possible diversity – (prior to mutation) about 1012

  10. Cell memory and clonal selection • Acquired immunity ensures that self-antigens are not attacked through • Clonal selection • Clonal deletion • Repeated editing • Absence of Helper T cells ensures that immune response is not mounted • Initially, B cells produce membrane bound antibodies • Cells stimulated by an antigenic response proliferate and differentiate into effector cells that secrete antibody • A fraction of the simulated cells differentiate instead into memory cells that are rapidly induced in subsequent antigenic challenges

  11. T cells (made in thymus) • Only respond when foreign antigen displayed on surface of self-cells in periperal lymphoid organs • Recognise fragments of foreign particles displayed by MHC proteins • Cytotoxic T cells recognise and destroy infected cells • Helper T cells stimulate other cells • Macrophages • B cells • Cytotoxic T cells • T cell receptors are very like antibodies, but are membrane bound ONLY and are less variable

  12. Cytotoxic killing • Cytotoxic cells release perforin (related to C9) which makes the infected cell permeable to secreted serine proteases • Activate apoptosis (cell death) through capsase family

  13. MHC proteins • Major histocompatibility complex (chromosome 6 in humans) • Class I MHC present to Cytotoxic T cells • Class II MHC present to Helper T cells • The most polymorphic loci in humans (some have >200 alleles) 6p21.3 4 Mbp c. 127 genes DP DQ DR C4 C2 TNFa,b HLA-B HLA-C HLA-A HLA-D 18 genes Class II Class III Class I

  14. Rapid evolution of MHC genes In both humans and house mice, the antigen-binding site (ABS) of class I and II MHC molecules (light blue) have a high rate of nonsynonymous versus synonymous nucleotide substitutions, which is the opposite pattern for genes under purifying selection, such as nonantigen-binding sites of MHC molecules (dark blue). Adapted from Potts WK and Wakeland EK (1990) Evolution of diversity at the major histocompatibility complex. Trends in Ecology and Evolution5: 181–187.

  15. Helper T cell activation • Two signals are required • MHC bound peptide • Costimulatory signal • Different costimulatory receptors occur in different cell types • CD3 complex on all T cells • CD4 on helper T cells • CD8 on cytotoxic T cells • Cytokines secreted by T cells (and B cells) provide communication between elements of the immune system inducing and suppressing reactions • Interleukins • Interferons • TNF-a

  16. The immunoglobulin superfamily

  17. How to evade the immune system • Once in, replicate really fast • Bacteria, viruses • Replicate within cells that are trying to destroy you • Mycobacterium tuberculosis replicates in macrophages • Trick normal cells into taking you in • Yersinia pseudotuberculosis expresses a protein that binds to E-cadherin and stimulates cells into forming a cell junction, through which the bacterium can enter • Present very diverse surface proteins which immune systems are unlikely to have experienced before • Trypanosomes somatic switching

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