evolution of innate immunity prof. jim kaufman · evolution of innate immunity prof. jim kaufman 2...

Evolution of Innate ImmunityProf. Jim Kaufman

1The screen versions of these slides have full details of copyright and acknowledgements

1

Evolution of Innate Immunity

Prof. Jim KaufmanUniversity of Cambridge

Department of Pathology

(and Department of Veterinary Medicine)

[email protected]

2

Pathogens range from the tiniest viruses...

Retroviruswith 3 genes

(xenotropic murine leukemia virus-related

virus, or XMLV)

http://www.nih.gov/researchmatters/june2011/06132011XMRV.htm

http://newscenter.berkeley.edu/2011/02/07/salmonella-anti-viral-gene-therapy/

Salmonella bacteriawith 4000 genes

(Salmonella typhimurium)Platyhelminthan (cestode)

parasite with >17,000 genes(Taenia saginata) tapeworm

to bacteria...

to eukaryote parasites, up to tapeworms twelve meters longhttp://en.wikipedia.org/wiki/File:Taenia_saginata_adult_5260_lores.jpg

3

Pathogens (generally)• High mutation rate

• Rapid generation time

• Huge effective populations

Pathogens and hosts are locked in an evolutionary arms race

Animal Hosts (relatively)• Low mutation rate

• Slow generation time

• Small effective populations



4

• Viruses (picorna to herpes and pox)

• Bacteria (Rickettsia, Salmonella, etc.)• Unicellular parasites (apicomplexins, schistosomes, etc.)

• Multicellular parasites (tapeworms, helminths)

• Tumours

• Transmissible Tumours (canine and Tasmanian devil)

Cells re-asserting their independence have always been a problem for multicellular organisms

What are the pathogens against which immunity must protect?

• Transmissible spongiform encephalopathies (TSEs, like “mad cow disease”)

Our immune systems aren’t really evolved to deal with all the pathogens we are exposed to...

5

Selection is

• constant

• very strong

• extremely variable

• over long time periods

Given how enormous and complex the challenge, it is perhaps surprising that any hosts survive at all...

The response to pathogens and tumours is complicated because it has arisen over many millions of years,

step-by-step in response to many different challenges,

utilising whatever molecular and cellular tools were available

Why is the immune response so complicated?

6

Random repertoire

of receptors, clonally

distributed

Antigen bindingto the receptors

with complementarityleads to activationand proliferation

• Specificity• Diversity • Memory• Autoimmunity

An adaptive immune systemBurnet’s model of Clonal Selection

Immunogen/Antigen(any molecular shape!)



7

What kinds of barriers, cells and molecules are usually included in innate immunity?

• Barriers • Tight junctions in epithelial cells• Mucins and mucus• Stomach acid• Nutrient sequestration (factors which strongly bind iron, biotin, etc.)

• Cells• Phagocytes and other blood cells (macrophages, granuloctyes)• Natural Killer cells• αβNKT and γδ T cells (invariant or semi-invariant T cell receptors)

• Molecules• Antimicrobial peptides (AMP, like defensins, magainins, cercropins)• Complement (C’, like thioester-containing proteins including C4, C3 and C5, TEPs)• Lectins and collectins (MBL)• Scavenger domain-containing proteins• Toll-like receptors (TLRs)• Nucleotide-binding oligomerisation domain (NOD)-like receptors (NLRs)• Cytokines and chemokines• Intracellular defences, including PKR and interferon, lectins, TRIMs, RNAi, etc.

Or are stopped at this stage!

Most pathogens that get past the barriers are stopped at this stage!

Most potential pathogens (that is, most microorganisms) don’t make it past the barriers!

8

Cells all expressing a collection of the samePattern (or pathogen) Recognition Receptors (PRRs)

PAMP binding to PRR leads to immediate response as an effectorand/or induction of further responses

One kind of innate immune system

Pathogen-associated Molecular Pattern (PAMP)

(only molecular shapes that differ between pathogen and host)

Medzhitov and Janeway 2002 Science 296: 298-300

9

Cells expressing a receptor for “stress signals”

DAMP binding to “stress receptor” leads to immediate response as an effectorand/or induction of further responses

Another kind of innate immune system

Danger associated molecule pattern (DAMP)

(a self molecular shape that differsbetween a normal and a “stressed” cell)

Matzinger 2002 Science 296: 301-305



10

Cells (or molecules) recognise and spare cells with “self markers”and attack any cell lacking “self markers” (that is, “missing self”)

Cells with self molecules are safe

Yet another kind of innate immune system

Cells without self molecules are killed

Karre 2008 Nature Immunol 9: 477-480

11

Hosts respond to the bewildering complexity of disease and infectious pathogens with a enormous variety of responses

• Innate immunityBarriers like mucus and skinPattern Recognition Receptors (PRRs) like lectins and TLRsCells like macrophages and natural killer (NK) cells

• Adaptive immunityAntibodies (Ab) and T cell receptors (TcR)Variable lymphocyte receptors (VLR)

• Genetic ResistanceHemoglobin for malaria in humansNramp for bacteria in miceMx for influenza in mice


Very ancient(mammals, flies and even plants)

Ancient(all vertebrates)

More recent(species or evenpopulations)

A successful allele for genetic resistance gets “fixed” as a mechanism of innate immunity

12

A textbook view of immune responses(actually many waves of responses, which differ

for different pathogens and locations)

Response

TimeInnate• “Ready to go” (constitutive)• Recognises conserved

pathogen features (PAMPs)

Required for adaptive response

• Some diversity• Some specificity• “Evolutionary memory”

Adaptive• “Needs time” (inducible)• Recognises everything

(randomly generated)

Requires innate response

• Enormous diversity• Exquisite specificity• “Somatic memory”

Why it takes so long to get going

Vaccines

Adjuvants

Including self(autoimmunity)

Preformedinnateresponses

Induced innateresponses

Lymphocyteresponses Appearance of antibodyBarriers



13

So, what is the evolutionary history of immune responses,

And how is it determined?

14 Figure courtesy of Louis Du Pasquier, U of Basel, 2004

MHC, TcR, Ab (RAG)VLR (CDA)

A phylogenetic tree of the vertebrates

15

A phylogenetic tree of many of the multicellular animalsvertebratesinvertebrates

Metazoans(organised multicellular animals)

time

Porifera Cnidaria Platyhelminthes Nematoda Mollusca Annelida Arthropoda Echinodermata Chordata

humanmouseratopposumwallabyplatypuschickenclawed frogaxolotlsalmoncodnurse shark

lampreyhagfishlancettunicate



16

• Some mechanisms (or aspects) of innate immunity are already present in single-celled organisms

• Phagocytosis, intracellular PRRs and effectors

Lobosea

Mycetozoa

PelobiontaEntamoebae

Choanoflagellata

Ichthyosporea

Cristidiscoidea Fungi

Eubacteria Archaea

Green algaePlantae Cercozoa

EuglenozoaHeteroloboseaJakobida

OxymodadidaTrimastix

Malawimonas

DiplomonadidaCarpediemonasRetortamonadidaParabasalida

Excavata

RhizariaRhodophytaGlaucophyta

AlveolataStramenopila

Cryptophyta

HaptophytaRadiolaria

ChromalveolataArchaeplastida

Amoebozoa

Opisthokonta

Bacteria

MetazoaVertebrate andinvertebrate animals

Plants

Amoeba

A phylogenetic tree of living organismsThere are a large number of different protists (single cell eukaryotes) and colonial animals

time

17

DeuterostomesProtostomes

A phylogenetic tree of many of the multicellular animals

corals flukes whip worms clams leeches lobsters sea urchins humans sea anemones tapeworms hook worms snails earthworms spiders sand dollars lancets

sponges jelly fish flatworms round worms squids segmented worms insects sea stars tunicatesPorifera Cnidaria Platyhelminthes Nematoda Mollusca Annelida Arthropoda Echinodermata Chordata

time


18

• A pathogen that kills all of it’s hosts may face a rather bleak evolutionary future

• So, pathogens apparently adjust their virulence to the immune defences of their host(s)

• Pathogens of vertebrates are not better, just different!

AbTcR

MHC/VLR

Only innate immunity?

Adaptiveimmunity

How do invertebrates survive with just innate immunity?



time



19

Arthropods(fruit fly Drosophila melanogaster)Down syndrome cell adhesion molecule (DSCAM)Watson et al. 2005 Science 309: 1874Dong et al. 2006 PLoS Biology 4: e246

FREP DSCAM

In fact, some invertebrate have independently evolved adaptive immune systems of their own!

AbTcR

MHC/VLR

Innate immunity

Adaptiveimmunity



time

20

Echinoderms(purple sea urchinStrongylocentrotus purpuratus)toll-like receptors (TLRs)Rast et al. 2006 Science 314: 952Hibino et al. 2006 Devel Biol 300: 349

TLR,NLR

lectins

Other invertebrates have large multigene families of receptors that may or may not be independently evolved adaptive immune systems

FREP DSCAM

AbTcR

MHC/VLR

Innate immunity

Adaptiveimmunity



time

21

In this sense, the high allelic polymorphism of some single genes,

the large number of genes in some multi-gene families the huge variation in some genes between cells in a single individual

are all part of a continuum:

Variation due to • Multigene families• High allelic polymorphism• Existing in the germline (germ cells) • Selected in evolutionary time

(for instance, between generations)

Variation due to • Somatic mutation• Gene conversion, etc.• Generated in individual somatic cells• Selected within individuals

Adaptive immunity

Innate immunity

Given the relentless arms race between pathogen and host, the most important feature of the host responses is diversity,

no matter how it is generated and maintained



22

• There is a molecular arms race between host and pathogens which drives an enormous diversity of host responses, almost bewildering to consider and difficult to systematise

• All these many systems and variants need to work together within a single individual (although sometimes they don’t, which results in various kinds of autoimmune and other kinds of diseases)

• In this sense, all these host responses do form a system

A short summary, a conclusion and a caveat

Response

Time

Preformedinnateresponses

Induced innateresponses

Lymphocyteresponses Appearance of antibodyBarriers

23

There exists lots of variation between groups of animals!Some important systems of invertebrate innate immunity are not as important or even not present in vertebrates

• Peptidoglycan recognition proteins (PGRPs) found from flies to lancets (Amphioxus)

• Phenoloxidase-based melanisation found in arthropods

• Variable domain-containing chitin-binding proteins (VCBPs) in lancets (Amphioxus)

• Fusion/Fester system of protochordates like the colonial tunicate Botryllus

• Sp185/333 proteins in sea urchins

A short summary, a conclusion and a caveat

24

How can we determine the evolutionary history of innate immunity?


• Cells• *Phagocytes and other blood cells (macrophages, granuloctyes)• *Natural Killer cells• αβNKT and γδ T cells (invariant or semi-invariant T cell receptors)

• Molecules• *Antimicrobial peptides (AMP, like defensins, magainins, cercropins)• *Complement (C’, like thioester-containing proteins including C4, C3 and C5, TEPs)• Lectins, ficollins and collectins (MBL)• Scavenger domain-containing proteins• *Toll-like receptors (TLRs) • Nucleotide-binding oligomerisation domain (NOD)-like receptors (NLRs)• Cytokines and chemokines• Intracellular defences, including PKR and interferon, lectins, TRIMs, RNAi, etc.

*Five examples



25

and also found among Cnidaria, the basal group Anthozoa (sea anemone and corals)

but missing from Hydrozoa (hydras and Portuguese Man o’ War), presumably due to secondary gene loss

(Miller et al. 2007 Genome Biol 8: R59)

time




TLR TLR

TLRTLR

TLR TLR TLR TLR TLR

Medzhitov et al 1997 Nature 388: 394-397

26

Leucine-rich repeat (LRR) domains

Toll-interleukin 1 receptor (TIR) domains

And genes (or portions of genes) similar to TLR (Toll or TLR-like genes) and some portions of certain signalling pathways

are reported to be present in distantly related organisms• TIR domains in amoeba

(Sillo et al. 2008 BMC Genomics9: 291)

• TIR domains suggested for bacteria (Beutler & Rehli 2002 Curr Top Microbiol Immunol 270:1)

• LRR-TIR genes along with Pelle homologs as resistance genes in plants (Tang et al. 1999 Plant Cell 11:15)

Rock et al. 1998 PNAS 95: 588


27

Lobosea

Mycetozoa


Metazoa

ChoanoflagellataIchthyosporea


Eubacteria Archaea

Green algaePlantae

RhodophytaGlaucophyta

AlveolataStramenopilaCryptophyta

Haptophyta

Radiolaria

Cercozoa



Malawimonas


Excavata

Rhizaria


Amoebozoa

Opisthokonta

Bacteria

Vertebrate andInvertebrate animals

Plants

Amoeba

TLRs have a very complicated evolutionary history, starting with portions of TLRs which are united into TLR genes and then used in different ways

TLR

“TLR”

“TLR”

“TLR”?




28




• Molecules• *Antimicrobial peptides (AMP, like defensins, magainins, cercropins)• *Complement (C’, like thioester-containing proteins including C4, C3 and C5, TEPs)• Lectins, ficollins and collectins (MBL)• Scavenger domain-containing proteins• *Toll-like receptors (TLRs)• Nucleotide-binding oligomerisation domain (NOD)-like receptors (NLRs)• Cytokines and chemokines• Intracellular defences, including PKR and interferon, lectins, TRIMs, RNAi, etc.

29

The complement system• Composed of around 40 proteins in mammals

many soluble in blood and some membrane bound on cells

• Important as PRR to detect non-self, PAMPs and DAMPs

• Important as effector system in innate and adaptive immunity

• Influences adaptive immune response (particularly B cells and Ab)

• Affects many other biological systems as part of homeostasisRicklin et al. 2010 Nat Rev Immunol 11: 785

Components of complement found widely in metazoans• Particularly important for human resistance to bacteria

Serruto et al. 2010 Nat Rev Microbiol 8: 393-399

• Important for mosquito resistance to malaria parasitesBlandin et al. 2009 Science 326: 147-150

Complement is an ancient and important molecular system

30

Thioester-containing proteins (TEPs including C3 and C4)

Molecular glue!

GlutamineO

CH2

CH2 CH2

CysteineC S

TEP

CH2

CH2 CH2

SHO

COH

TEP

Any protein (or sugar)

Also found in the protease inhibitor a2-macroglobulin

Complement is an ancient and important molecular system



31

Complement tickover

self DAF

non-self

selfDAF

non-s e l f

C3

Complement can detect “missing self”

Decay accelerating factor (DAF or CD55)

Why isn’t this enough?

32

C4C1r/C1s/C1q/Ab

C4b

C4MASP1/MASP2/MBL

C4b

C3fBb/C3(H20)

C3b

Complement cascadeCollagen-tulip family molecules

a2-macroglobulin family moleculesSerine protease family molecules

Classical pathway (adaptive immunity)

Lectin pathway (innate immunity!)

Alternative pathway (innate immunity/amplification loop)

C3

C2a/C4b

C3b

C3

C2a/C4b/C3b

C3b

C5C2a/C4b/C3b

C5b

C5fBb/C3b/C3b

C5b

fBb due to fD

Evolution by duplication of little modules...

33time

Complement components are very ancient Cerenius et al. 2010 Trends Biochem Sci 35: 575 Nonaka and Kimura 2006 Immunogenetics 58: 701-713 Dishaw et al. 2005 Immunogenetics 57: 535-548

MASPfBC3

fBC3

fBC3

fBC3

fBC3

fBC3

MASPfBC3

fBC3






34





35

Natural Killer (NK) cells • Innate lymphocytes, each with many receptors

• Crucial roles in infectious disease, cancer, autoimmunity, transplantation and reproduction

Parham 2005 Nat Rev Immunol 5: 201

NK receptors• Ligand recognition by lectin or immunoglobulin domains

• Signalling either activating or inhibitory

• Often highly diverse between individuals

NK cells are an important arm of the innate immunity

36

NK cells detect “missing self”

selfI

?

I

I

NK

+--

-

virus

selfI

?

I

I

NK

+--

-

NK

+

-

virus

self?

I

Inhibition wins:no kill

Viral infection:Down-regulation of class Ifor immune evasion of CTLs

Activation wins:Kill!

KIR orLy49

MHC class I molecules

Ljunggren and Karre 1990 Immunol Today 11:237



37

NK cells detect stress

NK

+--

-

NK

+--

-

Viral infection:Down-regulation of ligandsfor “stress receptors”

NKR-P1

NKR-P1

self?

LL

LLLT1/clr

virus

self?

LL

LLLT1/clr

virus

self?

I

NK

+

- LNKR-P1 LLT1/clr Voigt et al. 2007 Immunity26: 617

Inhibition wins:no kill

Activation wins:Kill!

38

No stress:no kill

Viral infection:Stress marker is expressedon the cell surface

Stress:Kill!

A really simple system...perhaps how it all started

NK

+

NKG2D

virus

selfNK+

M

virus

selfNK

+ M

selfM MIC-A

Bauer et al. 1999 Science285: 727

NK cells detect stress

39 Spits and Di Santo 2011 Nature Immunol 12: 21

A new concept of “innate lymphocytes”

Involved in Th1 responses




Involved in lymph node formation

Each with its own function

Each with its own transcription factor

Figure 2. Transcription factors regulate the differentiation of distinct ILC subsets. Different transcription factors are essential for the development of different ILC subsets, including bone marrow NK precursors (bmNKP), thymic NK precursors (thyNKP), ILC2 precursors (ILC2P), LTi precursors (LTiP), ILC17 precursors (ILC17P) and ILC22 precursors (ILC22P), from Id2-expressing ILC precursors (ILCP)



40

Hosts respond to the bewildering complexity of disease and infectious pathogens with a enormous variety of responses

• Innate immunityBarriers like mucus and skinPRRs like lectins and TLRsCells like macrophages and NK cells

• Adaptive immunityAntibodies (Ab) and T cell receptors (TcR)Variable lymphocyte receptors (VLR)

• Genetic ResistanceHemoglobinNrampMx

• Are innate lymphocytes the evolutionary precursors of adaptive lymphocytes

• Or are innate lymphocytes descended from adaptive lymphocytes by secondary evolution

• Or was there a mixture of the two processes?


41

NK receptors evolve quickly under selection by pathogens!

KIRKIRKIRKIRKIRKIR

NKR-P1

Clr

NKG2Ly49Ly9Ly49

NILTNILTNILTNILTNILTNILT

NKR-P1

LLT1

NKG2lectinlectinlectin NKG2/CD94

Fish Mouse Human

NK receptors show different molecules doing the same job

Yoder and Litman 2011 Immunogenetics63: 123

42time

Porifera Cnidaria Platyhelminthes Nematoda Mollusca Annelida Arthropoda Echinodermata Chordata


NK?

A phylogenetic tree of many of the multicellular animals

Follow evolution of NK cells by transcription factors and signalling cascades?

humanmouseratopposumwallabyplatypuschickenclawed frogaxolotlsalmoncodnurse shark

lampreyhagfishlancettunicate

NK



43





44

Anti-microbial peptides (AMPs) are important for innate immunity

Wiesner and Vilcinskas 2010 Virulence 1: 440

AMPs are small peptides (12-50 amino acids) composed of hydrophobic and positively charged residues, classified by structure, amino acid composition and number of disulfide bonds

Brogden 2005 Nat Rev Microbiol 3: 238Wimley & Hristova 2011 J Membr Biol 239: 27

That disrupt pathogen membranes And/or interfere with intracellular functionsAnd also stimulate inflammatory reactions by host cells

45

• In humans, there are families ofDefensins (secondary structure of β-strands)

– α-defensins from macrophages, neutrophils and Paneth cells of the small intestine

– β-defensins from epithelial cellsCathelicidins (secondary structure α-helices) mainly from neutrophils

Circular (theta) AMPs, most recently described

• AMPs kill gram-positive and gram negative bacteria, mycobacteria, enveloped viruses, fungi, parasites and even tumour cells

• AMPs from across the living organisms have been described, including Magainins from frog skin

Cecropins from insects and nematodes

Purothionins from plants

Bacteriocins and lantibiotics (e.g. gramacidins) from bacteria

Anti-microbial peptides (AMPs) are widespread in living organisms



46

Lobosea

Mycetozoa


Metazoa



Eubacteria Archaea

Green algaePlantae




Cercozoa



Malawimonas


Excavata

Rhizaria


Amoebozoa

Opisthokonta

Bacteria


Plants

Amoeba

A phylogenetic tree of living organisms

AMP

AMP

AMP

AMP

47





48 Pluddemann et al. 2011 Immunol Rev 240: 11

Macrophages (and other cells like granulocytes) are key cells for innate immunity in humans and other vertebrates Also, critical in delivery of effector functions,

including phagocytosis, reactive oxygen species (ROS) and nitric oxide species (NOS), as well as antigen presentation to lymphocytes

Critical in detection of pathogens, by a host of PRR and other receptors

And have a range of other functions, including development, tissue homeostasis and wound repair



49

• Macrophage-like cellsAre found in all metazoan animals examined, including sponges

Carry out a range of biological functions, including pathogen recognitionEngulf bacteria and destroy them by ROSAre extremely similar to amoeboid protists, such as Acanthamoeba

• Cells with some similarities to granulocytes are found in some other metazoans

In humans, there are neutrophils, basophils (mast cells) and eosinophilsIn chickens, there are a range of heterophilsIn fruit flies, there are four kinds of hemocytes (one like macrophages)In some other metazoans, there are “coelomocytes”

• So, these other cells are less well-characterised but apparently more heterogeneous

• Functions may be traded around:In chickens, thrombocytes (analogous to platelets) are phagocytic antigen presenting cells

Macrophages are found throughout the metazoans, and cells like granulocytes

are also widely distributed

50

Macrophages

Macrophages?

Lobosea

Mycetozoa


Metazoa



Eubacteria Archaea

Green algaePlantae




Cercozoa



Malawimonas


Excavata

Rhizaria


Amoebozoa

Opisthokonta

Bacteria


Plants

Amoeba

A phylogenetic tree of living organisms

51


• Barriers• Tight junctions in epithelial cells• Mucins and mucus• Stomach acid• Nutrient sequestration (factors which strongly bind iron, biotin, etc.)





52

Conclusions • Immunity as a whole is very complex, but has to work together within an individual

• The enormous complexity of immunity is primarily due to the ancient and on-going arms race with pathogens

• Adaptive immunity and innate immunity differ the way in which diversity is created, but the important principle is to have enough diversity to cope with existing pathogens

• Innate immunity is composed of many very different systems, based on several different principles

Some systems started at the time of single cells

Other systems appeared with the onset of organised multicellular organisms

And still other systems appeared later yet

Some systems may still be appearing, recruited from “genetic resistance” genes

• After being established, innate immune systems have complex evolutionary histories, becoming more or less important in particular groups of animals

• It is generally believed that adaptive immunity grows out of innate immune systems

Thank you for your attention.

53

evolution of innate immunity prof. jim kaufman · evolution of innate immunity prof. jim kaufman 2...

Documents