mcb 150: molecular today’s game-plan...
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MCB 150: MolecularImmunology
Spring 2007Prof. Ellen Robey
Prof. Robert BeattyProf. Laurent Coscoy
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Today’s game-plan
• Structure of the course and administrative
issues
• Overview of the immune system
• Begin: Innate Immunity
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MCB 150 Spring 2007
Innate Immunity Humoral ImmunityB cells, antibody, complementPhagocytes, Toll-like receptors
Cellular ImmunityT cells, TCR, development, selection, tolerance, NK cells
Immune Responses
inflammation, allergy, autoimmunity, infection 4
Course InstructorsLecturers:
Ellen Robey471 LSA 2-8669Office hours: Tues 11-12
Laurent Coscoy451 LSA 3-4128Office hours: TBA
Robert Beatty176 LSA 2-0671Office hours: Tues 3-4
GSIs:
Dina WeilhammerDina@berkeley.eduOffice hours: TBA
Tim Nicetimnice@berkeley.eduOffice hours: TBA
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Course Components
• Lecture series• Discussion sections (not required, but highly
recommended)• Reading: Text (Kuby 6th addition), Journal Articles• Problem sets (posted on web, these are not graded, but
are to help reinforce what you learn in class)• Exams-- two mid-terms, plus final exam
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Grading
• Mid-terms: 25% each• Final 50%
Half of the material final is a cumulative reviewand half focuses on the last third of the class.
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Course Web Site
• www.mcb.berkeley.edu/courses/mcb150• All lecture notes, problem sets, answers will
be posted for viewing or download• Powerpoint presentations shown in lecture
will be posted for your review
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Getting Help!!!•Web site:www.mcb.berkeley.edu/courses/mcb150
•Discussion groups / GSIs
•Office Hours
•After lecture-- very brief questions only
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•No Discussion sections this week.
•Sign up sheets available in class today
•You may attend any or all of the 4discussion sections.
•Problem set/reading material fordiscussion will be posted on website
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Overview of the Immune system
Microbes: why they are formidable foes.
Gross anatomy of the immune system
Cells of the immune system
Effector mechanisms: how the immune system protects
Immune recognition of pathogens: innate vs adaptive immunity
Cytokines and the inflammatory response
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Microbes are ubiquitous in nature, extraordinarily diverse,rapidly evolve to exploit opportunities to infect hosts and to
evade their immune systems.
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Exponential Growth
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10
100
1000
10000
100000
1000000
10000000
100000000
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25
Time (minutes/10)
# of bacteria
8 hours = 280 trillion bacteria!!!!
Exponential growthN
umbe
r of b
acte
ria
Time (hrs)
Many pathogens can expand rapidly in the nutrient-richenvironment of the host.
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Some microbes hijack cellular machinery toreplicate and spread. Intracellular pathogens
include viruses (influenza, HIV) and intracellularbacteria (listeria) and intracellular parasites
(malaria, toxoplasma).
Listeria bacteria using the actin cytoskeleton ofthe host cell to spread from cell to cell.(Portnoy lab)
Toxoplasma parasites (red) and dendritic cells(green) within a mouse lymph node. (Robeylab) 14
Pathogens rapidly evolve to avoid the host immune response
Virally encoded decoy receptors
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Overview of the Immune system
Microbes: why they are formidable foes.
Gross anatomy of the immune system
Cells of the immune system
Effector mechanisms: how the immune system protects
Immune recognition of pathogens: innate vs adaptive immunity
Cytokines and the inflammatory response
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First some key definitions:
Pathogen: microbe that causes disease
Antigen: material (from a pathogen) that induces an immune response
Innate (natural) immunity: rapid, non specific immune response
Adaptive (acquired) immunity: slower, specific immune response
Leukocytes: blood cells
Lymphocytes: specialized blood cells that mediate adaptive immunity(e.g. T and B cells)
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The cells of the immune systemcirculate through the body via
lymph and blood. Pathogens andtheir antigens are transported from
tissues via lymphatic vessels tothe lymph nodes where they
encounter immune cells.
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The cells of the immune systemspend much of their time in
lymphoid organs. They develop(arise) in primary lymphoid
organs, and they interact withantigens in secondary lymphoid
organs.
Thymus: primary lymphoid organfor T cell development
Bone marrow: primary lymphoidorgan for B cell development
Lymph nodes: collect antigensfrom tissues
Spleen: collects antigens fromblood stream
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Overview of the Immune system
Microbes: why they are formidable foes.
Gross anatomy of the immune system
Cells of the immune system
Effector mechanisms: how the immune system protects
Immune recognition of pathogens: innate vs adaptive immunity
Cytokines and the inflammatory response
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Many different types of blood cells participate inthe immune response to microbes:
Innate immune cells: “phagocytes”macrophage, neutrophils, dendritic cells
Adaptive immune cells: “lymphocytes”T cells, B cells
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Most blood cellsact to fightinfection.
AdaptiveimmunityInnate
immunity
Blood cells lineages.
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Lymphocytes of the adaptive immune system
T helper cells: regulate other immune cellsT cytotoxic (killer) cells: kill infected cellsB cells: produce antibodies (immunoglobulin)
Dendritic cells and macrophage: directly kill microbes by phagocytosis and othermechanisms. They also help to activate T cells (connection between innate and adaptiveimmunity)NK cells are lymphocytes that have characteristics of innate and adaptive immunity.
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Overview of the Immune system
Microbes: why they are formidable foes.
Gross anatomy of the immune system
Cells of the immune system
Effector mechanisms: how the immune system protects
Immune recognition: innate and adaptive
Cytokines and inflammation
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Immune Effector Mechanisms:
Cell-mediated immunity:Phagocytosis (cellular eating)cytotoxcity (cellular killing)
Humoral immunity:complement: group of serum proteins that can directly kill
pathogens.antibodies: (also called immunoglobulin) proteins secreted
by B cells that bind directly and specifically to pathogens.Antibodies target pathogens by marking them for destruction byother components of the immune system.
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Macrophage fight microbes by engulfing anddigesting them (phagocytosis or cellullar eating).
A macrophage engulfing yeast
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Bacteria fight back againstphagocytosis
A macrophage attempting toengulf a bacterium Streptococcuspneumoniae, that is covered with
a slimy coat.
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Target cell killing by a cytotoxic (killer) lymphocyte
A cytotoxic T lymphocyte (CTL) killing a cellthat has been infected by a virus. NK cells use a
similar mechanism to eliminate tumor cells.28
Overview of the Immune system
Microbes: why they are formidable foes.
Gross anatomy of the immune system
Cells of the immune system
Effector mechanisms: how the immune system protects
innate and adaptive immunity
Cytokines and inflammation
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Found in:
Substancesthat trigger:
Receptors:
all multicellullar organisms
A limited number ofpathogen-associatedmolecular patterns (PAMPs)
A limited number of PatternRecognition Receptors(PRRs) expressed on manycells types
Vertebrates only
Virtually any component ofpathogens
Highly variable receptors of2 types: antibody made by Bcells and TCR made by Tcells 30
Comparison of innate and adaptive immune recognition
Receptors that mediateinnate immune recognition: Toll-like receptors (TLR)
Receptors that mediate adaptive immune recognition:Antibody and the T cell receptor (TCR)
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The genes encoding the T cell antigen receptor (TCR) are assembledby DNA rearrangement as T cells develop in the thymus
TCR ! locus:structure in germline
TCR ! locus:structure in T cells
DNA rearrangement(rag1, rag2)
transcriptionRNA splicingtranslation
T cell
!"
V segments D J C
The genes encoding the antigen receptors of T and B cells areassembled by DNA rearrangement as these cells develop. As aresult of V(D)J recombination, every B and T cells expresses aunique version of the antigen receptor.
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Overview of the Immune system
Microbes: why they are formidable foes.
Gross anatomy of the immune system
Cells of the immune system
Effector mechanisms: how the immune system protects
Immune recognition: innate and adaptive
Cytokines and inflammation
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Cytokines: (also called interleukins)
Small secreted peptides that used forintracellular communication between cellsof the immune system
Can turn on or off immune responses
Mediate inflammation
Chemokines: subset of cytokines thatspecialize in regulating cell motility
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Inflammation: a complex series ofevents induced by tissue damage(described in medical literature in 1500s)
• Redness (rubor)
• Pain (dolor)
• Swelling (tumor)
• Heat (calor)
35Infection can induce inflammation, but even sterile injuries can be
sufficient to induce inflammation.
Inflammation occurs when injured tissues release mediators that promotevasodialation (increased blood flow) and chemotaxis (directed migration)
of leukocytes.
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Inflammation causesblood cells to move
from blood stream tosite of injury
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Blood cells (leukocytes) travel from the blood stream intotissues by a process known as extravasation
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Blood cells can also be attracted to sites of infection byproducts produced by pathogens, as well as by
chemoattractants made by host (chemokines, inflammatorymeditators).
Neutrophils (a type of white blood cell) are attracted to bacterialproducts. Here they are moving toward a gradient of the
bacterial peptide fMLP.
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Why study the immune system?Importance of the immune system in human health
Provides model systems for studies of:gene regulationmolecular recognitionsignal transductionetc, etc
Provides powerful techniques for use in medicine and science
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Diseases associated with immunesystem dysfunction
• Autoimmunity (SLE, Arthritis,Myasthenia gravis, Graves disease)
• Immunodeficiency (inherited, acquired)• Allergy (environment, drugs)• Cancer (Leukemia, Lymphoma)
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Myasthenia gravis
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Asthma & Allergy
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Therapeutics
• Vaccination• Adoptive immunotherapy• Transplantation
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Vaccination: THE major successCases per
100,000population
Cases per100,000
population
Cases per100,000
population
Diphtheria
Polio
Measles
year46
Treating cancer
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Tissue Transplantation
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Powerful methods for detecting andquantitating proteins and cells based onthe highly specific binding of antibodies
(immunoglobulins)
• ELISA: enzyme-linked immunosorbant assay(measures small amounts of hormones, drugs,microbes in body fluids)
• Western blot (detects disease associated proteins)• Flow cytometry (quantifies various cell types in
mixed population-- HIV patients)
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Western Blot
•Separate proteins by size using PAGE gel
•Transfer gel to blotting membrane•Probe membrane with antibody
specific for protein of interest•Detect bound antibody by
chemiluminesence
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ImmunofluoresenceImmunofluoresence
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Flow Cytometry: QuantitativeSingle-cell Immunofluoresence
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Innate (natural) immunity
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Found in:
Substancesthat trigger:
Receptors:
all multicellular organisms
A limited number ofpathogen-associatedmolecular patterns (PAMPs)
A limited number of PatternRecognition Receptors(PRRs) expressed on manycells types
Vertebrates only
Virtually any component ofpathogens
Highly variable receptors of2 types: antibody made by Bcells and TCR made by Tcells 54
Innate ImmunityInnate immune effector mechanisms
Physical and biochemical barriers (defensins)Phagocytosis and reactive oxygenCell autonomous defenses
ApoptosisInterferons and PKR
Innate immune recognitiondiscovery of the Toll-like receptorsmammalian TLRs and their ligandsnon-TLR recognition of PAMPs
Connections between adaptive and innate immunity
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Physical & Biochemical barriers
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Defensins
• Originally isolated from frog skin based on their ability tokill bacteria
• Small polypeptides (<10kDa) secreted at mucosal surfaces• Direct bacteriocidal properties• Insertion into biological membranes leading to target cell
lysis• Inhibited by cholesterol (specificity) 58
Hanging out with Tolstoy 1909
In 1882, the Russian scientist Ilya Mechnikov was working in Messina, Italy, studying thelarvae of the sea star. When he inserted a thorn into a larva, something weird happened.Mechnikov noticed strange cells gathering at the point of insertion. The cells surrounded thethorn, eating any foreign substances that entered through the ruptured skin. Mechnikov wasthrilled. He decided to name these new cells phagocytes from the Greek words meaning"devouring cells."
Phagocytosis was discovered by Ilya Mechnikov in 1882
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Phagocytosis “cellular eating”
3. Phagosome fuses withlysosome.
1. Bacterium attaches tomembrane
2. Bacterium is ingested, formingphagosome,
4. Lysosomal enzymesdigest the bacteria.
5. Digested material isreleased from cell.
Phagocytes: macrophage,neutrophils, dendritic cells 60
Phagocytes use a variety of methods to destroy ingested microbes.
(household bleach)
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Macrophage fight microbes by phagocytosis andproduction of toxic molecules
A macrophage produces reactive oxygenspecies to aid in destruction of the microbe
Reactive oxygen is revealed by the blue dye,NBT 62
Innate ImmunityInnate immune effector mechanisms
Physical and biochemical barriers (defensins)Phagocytosis and reactive oxygen speciesCell autonomous defenses
ApoptosisInterferons and PKR
Innate immune recognitiondiscovery of the Toll-like receptorsmammalian TLRs and their ligandsnon-TLR recognition of PAMPs
Connections between adaptive and innate immunity
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Some microbes hijack cellular machinery to replicateand spread. Intracellular pathogens include viruses(influenza, HIV) and intracellular bacteria (listeria)and intracellular parasites (malaria, toxoplasma).
cell-autonomous defense:cell produces an immuneresponse that acts on itself
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Apoptosis: Cellular Suicide
•Nuclear fragmentation•Proteolysis•Blebbing•Death
Remnantsundergophagocytosis
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Apoptosis versus Necrosis•Tidy: contents of cellsdegraded from within,producing small cellular“blebs”
•Programmed from inside thecell
•Messy: contents of cellreleased.
•Induced by external insult
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Cell death by necrosis is more likely to produce inflammation.
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Interferons are cytokines that areproduced in response to viral infection.
Produce an “anti-viral state” in targetcells.
Acts on cell that produces it, as well asneighboring cells.
Together with dsRNA, act to triggeringthe Protein Kinase R(PKR) pathway.
Shuts down protein synthesis machineryof cells, thus preventing viral replication.
Cells can avoid being hijacked by viruses by activating the ProteinKinase R (PKR) pathway. PKR is triggered by dsRNA and interferon.
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Protein Kinase R:Interfering with Infection
RNA-bindingdomain
Kinasedomain
dsRNAor
interferon
PKR
eIF2a
PhosphorylatedPKR
(active)
PhosphorylatedeIF2a
(inactive)
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Innate ImmunityInnate immune effector mechanisms
Physical and biochemical barriers (defensins)Phagocytosis and reactive oxygenCell autonomous defenses
ApoptosisInterferons and PKR
Innate immune recognitiondiscovery of the Toll-like receptorsTLRs and their ligandsnon-TLR recognition of PAMPs
Connections between adaptive and innate immunity70
Comparison of the adaptive and innate immune responses
innate adaptive
Response time hours days
Response to identical to primary stronger response uponrepeat infection second exposure
Receptors that pattern recognition receptors antibodies and T cell antigen receptors (TCR)Mediate pathogen Toll-like receptors (TLR)Recognition limited diversity, unlimited diversity
fixed in germline generated by V(D)J recombination
Ligands Pathogen associated molecular virtually any component of pathogenpatterns (PAMPs)
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