Signaling through C-type lectin receptors: shaping immune response

Teunis B. H. Geijtenbeek and Sonja I. Gringhuis

Nature Reviews: Immunology, Vol. 9, July 2009, p.465-479

report by: Yuri Yakushko

Talk overview

1. Dendritic cells2. Pathogen recognition3. C-type lectins classification4. CLR signaling in general5. CLRs that interact with TLRs (DC-SIGN)6. CLRs that interact with TLRs (BDCA2)7. CLRs that interact with TLRs (DCIR, MICL)8. TLR-independent signaling by CLRs (dectin 1)9. TLR-independent signaling by CLRs (dectin 2 and mincle)10. CLRs and T-cells differentiation11. Therapeutic potential12. Future directions. Sum up

DCs:

• located throughout the body

• capture and process antigens

• present antigens with:• MHC I for CD8+ cells• MHC II for CD4+ cells

Picture source: www.healthsystem.virginia.edu

Myeloid dendritic cells (mDCs)

Most similar to monocytes. mDC are made up of at least two subsets:(1) the more common mDC-1, which is a major stimulator of T cells(2) the extremely rare mDC-2, which may have a function in fighting wound infection

Plasmacytiod dendritic cells (pDCs)

Look like plasma cells, but have certain characteristics similar to myeloid dendritic cells.

Dendritic cells

Section of skin showing large numbers of dendritic (Langerhans) cells in the epidermis.CD1a staining

www.dermatologyoutlines.com/dermcdmarkers.html

Dendritic cells

CD4+ cell

IL-4, IL-5, IL-13 to induce humoral immune response

(e.g. against helminths)IL-17 to mobilize

phagocytes to clear extracellular fungi and

bacteria

IFNγ that activates macrophages to fight intracellular infections

Regulatory T cell

Suppress effector T-cellsSome subpopulations of

T-suppressors:• CD8+ cells

• CD4, CD25 and Foxp3 (CD4+CD25+ regulatory T cells are exclusively called "Tregs")

Pathogen recognition

PAMP – pathogen-associatedmolecular patterns: groups of pathogens

that share similar structures

PRR – PAMP Recognition Receptors:

1. Toll-like receptors (TLRs):• 1,2,4,5,6,10,11 on cell surface• 3,7,8,9 in cell compartments

2. non-TLRs:• retinoic acid-inducible gene I (RIG-I)• C-type lectin receptors (CLRs)

C-type lectins

Lectins are sugar-binding proteins which are highly specific for their sugar moieties

Ca2+-independent lectins Ca2+-dependent lectins

C-type lectin receptors (CLRs) bind particular sugars

in Ca-dependent manner

C-type lectin-like molecules bind either carbohydrates,

polypeptide ligands or both.

• have at least one carbohydrate-recognition domain• can be cytoplasmic or transmembrane• some of them function as PRR (pathogen-associated molecular patterns (PAMP) recognition receptors)

C-type lectin receptors

CLRs

asialoglycoprotein-receptor family (e.g. DC-SIGN, dectin 1)

mannose-receptor family (e.g. DEC 205)

Both families of receptors can recognize mannose, fucose, glucans etc.• mannose allows recognition of viruses, fungi and mycobacteria• fucose structures are usually expressed by bacteria and helminths• glucans are present on mycobacteria and fungi

Ag internalization, processing and presentation (on DC)

Some CLRs are expressed be several subsets of DCs:DC-SIGN and dectin 1 on subepithelial and some myeloid DCsOther CLRs are specific for particular DC subsets:• langerin (=CLEC4K=CD207) on Langerhans cells (e.g. epedermis), • BDCA2(=CLEC4C) on plasmacytoid DCs

C-type lectin receptors, pathogen recognition and signaling

C-type lectin receptors, pathogen recognition and signaling

Ag-processing via CLRs on dendritic cells

1

1

12 2

• CLRs are targeted in vaccine studies to increase Ag-specific immune responses: DEC205 was one of the first targets in these studies

• CLEC9A is expressed on mouse CD8+ cells and recognized necrotic cells. When bound to ligand, activates SYK (spleen tyrosine kinase) for cross-presentation of necrotic cell-associated Ag

1

2

3

1

23

Cross-presentation means presenting Ag with MHC I to CD8+ cells

CLRs signaling

CLRs

Protein kinases or phosphatases that direclty or indirectly interact with their cytoplasmic domains

ITAM/ITIM-containing adaptor molecules

(e.g.Fc-receptor γ-chain (FcRγ) and DAP12)

signaling through

• mincle• dectin 2• BDCA2

• dectin 1• DC-SIGN• DCIR• MICL

ITAM – immunoreceptor tyrosin-based activation motifITIM – immunoreceptor tyrosin-based inhibition motif

CLRs signaling

CLRs

ITAM / ITIM-containing adaptor molecules orpKs/phosphatases

NF-kB(nuclear transcription factor)

??

inducible gene expression in the immune system

CLRs signaling and TLRs pathways

1. BDCA2, DC-SIGN, DCIR, MICL can modulate TLR-induced gene expression (but only when they are activated by their specific PRR). All these molecules do not effect gene expression without signals, induced by other PRRs.

2. Dectin 1, dectin 2, mincle do not interact with TLRs signaling. But they can induce gene expression without other PRRs.

DC-SIGN activation

Mannose-containing pathogens(e.g. M. tuberculosis, HIV-1,C. albicans, measles virus)

NF-kB

TLR4

TLR3TLR5

Ras (GTPase)

RAF1-GTP

RAF1 (Ser/Thr kinase)

Src and PAKskinases

RAF1-P

1

2

+3

?

5

4

Dendritic cells-sprecific ICAM3-grabbing non-integrin (DC-SIGN) signaling

?

1

2 3

4

5

Nf-kB

CLRs that interacts with TLRs pathways

NF-kB: subunit p65+P

CREB(cAMP response element binding)

DNA regions called CRE (cAMP response elements)

CBP (CREB-binding protein)

Histone acetyltransferase: increases DNA binding activity of NF-kB p65 subunit

CBP + CREB + NF-kB + DNA complex

2

+

1

Transcriptional activation of il8 and il10 promoters

• IL8• IL10

3

4

Dendritic cells-sprecific ICAM3-grabbing non-integrin (DC-SIGN) signaling

RAF1-P?

1

34

2

DC-SIGN signaling. Sum up

DC-SIGN

Ras + RAF1

Nf-kB + CREB-CBP + DNA

il8, il10 genes expression

?

MEK-MAPK/ERK cascade

Ixodes scapularis protein (“Salp15”):• activates RAF1, but downstream cascade does not involve p65 (of Nf-kB). • MEK is activated, but not the following ERK.

gp120 of HIV-1 is linked to ERK activation, but DC-SIGN role not proved

RAF1-MEK-MAPK/ERK-cascade launched by some DC-SIGN ligands (peanut allergen Ara h1or Schistosoma mansoni egg Ag)

RAF1-MEK-MAPK/ERK-cascade not launched by DC-SIGN (but by anti-DC-SIGN antibody)

Ras – RAF1 - MEK

Borrelia burgdorferi

TLR

il6 and Tnf genes

mRNA

2

1

1

1

3

1

22

mRNA decay

3

Ixodes scapularisDC-SIGN signaling plasticity

I.scapularis spreads B.burgdorferi that causes Lyme disease (borreliosis)

So DC-SIGN signaling is of high plasticityand poorly understood. So guys, work more:)

?

http://www.genome.jp/kegg/pathway/hsa/hsa04010.htmlor http://www.kegg.jp

MAPKpathways

BDCA2 signaling through ITAM-containing FcRγ

BDCA2 – Blood DC antigen 2 proteinITAM – immunoreceptor tyrosin-based activation motif

Fc-receptor γ-chain, bound with ITAM 1

BDCA2

+ 2P

FcRγ-ITAM-PP

+

SYK (spleen tyrosine kinase)

BTK (Bruton’s tyrosine kinase)BLNK (B-cell linker)

PLCγ2 phospholipase

comple

x

2

3

Ca2+ TLR pathway

?

12

3

4

4

1. Tonic Ca2+ signaling also inhibits TLR pathway via activating calcineurin (phosphatase). Calcineurin then inhibits MYD88 (myeloid differentiation primary response protein 88).

2. Thus BDCA2, perhaps, also inhibits MYD88 (and downstream TLR-induced pathway) through calcineurin.

3. In macrophages Ca2+ signaling is induced by TREM2 (triggering-receptor expressed on myeloid cells 2), that also binds to ITAM-proteins and down-regulates TLR-signaling.

BDCA2 signaling through ITAM-containing FcRγ

CLR signaling through ITIMs

ITIM – immunoreceptor tyrosin-based inhibitory motif

DCIR and MICL are the only known CLRs that contain ITIMin their cytoplasmic tails

Both molecules are not shown to induce immune responses on their own, but can modulate signaling pathways induced by other PRRs

DCIR signaling

1

DCIR (with ITIM tail)

?

DCIR – DC immunoreceptor

SHP1 or SHP2

SH2-domain containing protein tyrosin phosphatases

TLR8-mediated IL-12 and TNF production

by myeloid DCs

TLR9-induced IFNα and TNF production

by plasmacytiod DCs

1 2

2 3

3

33

MICL signaling

MICL – myeloid C-type lectin like receptorERK – extracellular signal-regulated kinase

1

MICL (with ITIM tail)

SHP1 or SHP2

SH2-domain containing protein tyrosin phosphatases

ERK?

21

2

TLR-induced IL-12 expression

3 ?

Well, you already know, right? WORK MORE

ITIM-bearing molecules sum up

1. ITIM-bearing molecules seem to suppress cytokine responses induced by other PRR through the recruitment of SHPs.

2. SHPs decrease the production of TLR-induced pro- inflammatory cytokines

3. but SHPs increase production of type 1 IFNs induced by TLRs

ITIM – immunoreceptor tyrosin-based inhibitory motif

DCIR – DC immunoreceptor

MICL – myeloid C-type lectin like receptor

SHP – SH2-domain containing protein tyrosin phosphatases

DCIR and MICL pathways plasticity and interaction with TLR signaling are not clear.

TLR-independent signaling by CLRs. Dectin 1

1. Dectin 1 induces gene expression independently of other PRRs.

2. Works through recognition of β-1,3-glucan PAMPs that are expressed by many pathogens (C.albicans, Aspergillus fumigatus, Pneumocystis carinii)

Dectin 1 – DC-associated C-type lectin 1

TLR-independent signaling by CLRs

Dectin 1

1

2 x Dectin 1 (YxxL motifs, where x – any aa)

“canonical NF-kB pathway”

SYK (spleen tyrosine kinase)

NF-kB (p65 and REL subunits)

CARD9BCL-10MALT1

comple

x

? ?? TRAF2-TRAF6 (TNF receptor-associated factors)

2

3

4

1

2

3

4

2 x Dectin 1 (YxxL motifs, where x – any aa)

SYK (spleen tyrosine kinase)

NIK (NF-kB-inducing kinase)

IKKα (IkB kinase-α)

Dectin 1

Non-canonic NF-kB pathway

NF-kB

?

1

3

2

1

2

3NF

-kB Never shown to SYK before.Fast kinetics comparing

with other inductorssuggests that dectin 1

launches unique pathway

Dectin 1. Some more

1. SYK-CARD9-dependent pathway activated in response to C.albicans leads to activation of NLR family, pyrin domain containing 3 (NLRP3=NALP3) inflammasome. The pathway leads to processing pro-IL-1β to active form IL-1β by caspase 1 through the generation of reactive oxygen species. However, details are unclear and need clarification

2. SYK also converge with RAF1 pathway, as SYK induces phosphorylation of p65 subunit of NF-kB by RAF1, which, finally, results in induction of il6, il10, il12a and il12b genes transcription.

3. SYK- and RAF1-pathways fine-tune NF-kB-induced cytokine responses:p65-P subunits form dimers that cannot bind to DNA. This leads to reduced production of IL-1β, IL-12 and IL-23 – major cytokines for Th differentiation.

4. Dectin 1 also induces CCL17 and CCL22 production (CC-chemokine ligand) which are involved in the recruitment of other leukocytes.

Dectin 1 in the only CLR known to induce non-canonic NF-kB pathway!

Dectin 2 and mincleBoth molecules signal through ITAM-containing FcRγ (like BDCA2)

ITAM – immunoreceptor tyrosin-based activation motif

Dectin 2 / mincle

SYK (spleen tyrosine kinase)1

?

NF-kB

CARD9-BCL10-MALT1 complex?2

2

1

• CARD9 – caspase recruitment domain family, member 9• BCL10 – B-cell lymphoma 10• MALT1 – Mucosa associated lymphoid tissue lymphoma translocation gene 1

Dectin 2 • C.albicans, Trichophyton rubrum, Microsporum audouinii

• house dust mite allergens

Dectin 2 and mincle

+

Pro-inflam. cytokines: TNF, IL-10

Mincle in macrophages

+ dead cells

CXCL2 neutrophils migrations

CXCL2

+ pathogenic fungus Malassezia spp.

TNF

CLRs and T cell differentiation. C.albicans

DC-SIGN Dectin 1+

β-glucan structures on C.albicans+

NF-kB

Th1 Th17

differentiation

+

1

3

2

1

2

3

1

CLRs and T cell differentiation. M.tuberculosis

DC-SIGN Dectin 1+

mannose on M.tuberculosis+

NF-kB

Th1 Th2

differentiation

+

1

3

2

Th17+

1

2

3

M.tuberculosis activation mechanisms are not fully understood

Therapeutic potential of CLRs signaling1. CLEC5A inhibition during dengue virus prevents virus-induced plasma

leakage and reduces mortality in mice.

2. Patients with SLE have reduced number of pDCs expressing BDCA2 that results in excessive production of type I IFNs, which is major pathophysiological factor in SLE.

3. DCIR deficiency in mice leads to the development of autoimmune diseases, such as rheumatoid arthritis.

4. Dectin 2 triggering by dust mite (Milben) allergens leads to cysteinyl leukotriene production that causes inflammation.

5. Peanut allergen Ara h1 interacts with DC-SIGN and induces Th2-cells responses.

• CLR agonists might activate pathways to prevent autoimmune disorders• CLR antagonists could attenuate or modulate inflammation

Vaccination strategies1. Specific delivery of antigens to DCs (e.g. on Ag-covered particles). This

approach was proved to work, but remains expensive and difficult.

2. Targeting DEC205, DC-SIGN or mannose receptor by antibodies induces CD4+ and CD8+ cells responses. This can be used to induce tolerance to pancreatic β-cells, thereby preventing type I diabetes (autoimmune type mediated by T-cell destruction of β-cells). Therapy against β-cell antigens expressed on DEC205 DCs reduces autoreactive CD8+ cells.

3. Mice immunization with zymosan or curdlan (dectin 1 ligands) induces specific CD4+ Th1 and Th17, as well as CD8+ responses through SYK-CRAD9 pathway. So could be used as adjuvants.

4. RAF1 inhibition is also studied, but RAF1 is involved in too many vital mechanisms to be a proper immunomodulatory target.

5. Carbohydrate-expressing ligands lack specificity as are recognized by several CLRs.

Vaccination strategies

DC-SIGN Dectin 1+

carbohydrate-coated particle with antigens+

NF-kB

Th1 Th2

differentiation

+

1

3

2

Th17+

Selective inhibition of RAF1 and/or SYK allows the modulation of the immune responses

4

1

2

3

4 4

Future directions. Summary

1. CLRs are not just antigen uptakers, but modulators or even initiators of immune responses.

2. Some of them can induce different signaling pathways by themselves.

3. Cross-talks between CLRs and TLRs signaling remains to be fully understood (well, as everything in biology)

4. Many CLRs act either through SYK-CARD9 or RAF1 pathways and activate NF-kB.

5. Role of other CLR-inducible transcription factors in immune system is still unclear

Thank you for you attention