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Leukocytes Polarize in Gradients of Chemoattractants, then Crawl to the Source. Lamellipod. Uropod. Crawling Leukocyte. Resting Leukocyte. CHEMOATTRACTANT (
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Leukocytes Polarize in Gradients of Chemoattractants, then Crawl to the Source
RestingLeukocyte
CHEMOATTRACTANT
(<1%)
Lamellipod
Uropod
Crawling Leukocyte
CX3C: ……………CXXXC…………………… C…………………C……………… CX3CL1 non-ELR CXC: ……………CX__C………………………C…………………C……………… CXCL# ELR CXC: …ELR…CX__C………………………C…………………C……………… CXCL# 4C CC: ……………C___C………………………C…………………C……………… CCL# 6C CC: ……………C___C…………C…………C…………………C……C……… CCL# C: ………………………C……………………………………………C……………… XCL#
Class NamesStructural Signature
Chemokines: Defined by Structure, Classified and Named by Patterns of Conserved Cysteines
…but unequally distributed among the 4 classes
CX3C (n=1)
(n~16)
(n~28)
C
CXC CC
(n=2)
The Chemokine Fold
G.M.Clore & A.M.GronenbornPDB ID: 1IL8
C
N
Chemokine receptors are defined by function, not structure: they bind chemokines and transduce signals.
To date all known chemokine receptors are members of the 7TM GPCR superfamily, coupling to Gi.
Chemokine Receptors
Leukocyte Chemoattractants Act Through a Family of 7-trans-membrane domain G Protein-coupled Receptors
7TM Receptor
G protein
Leukocyte Trafficking
Chemoattractant
Chemokine Binding Protein Classification
CXCR1 CXC CXCR2 CXCR3 CXCR4 CXCR5 CXCR6 CCR1 CC CCR2 CCR3 CCR4 CCR5 CCR6 CCR7 CCR8 CCR9 CCR10 XCR1 C CX3CR1 CX3C D6 CC Duffy CC & CXC CCX/CKR CC
Chemokine Subclass Specificity
Chemokine Binding Proteins (7TM architecture)
7TM G Protein-coupled Receptors
Neurotransmitter Receptors
TSH R
LHCG R
b1 AR b2 AR
HT 2 HT 1c A1 P1
A2 P1 a2a AR a2b AR a2c AR
HT 1a M 1
M 5M 3
M 2M 4
RhodopsinOpsin BlueOpsin GreenOpsin Red
SKR SPR IL8RAIL8RB
MIP-1a/RANTES R
FPRL1 R
FPRC5a RPAF R
Leukocyte Chemoattractant Receptors
Polypeptide Hormone Receptors
Light Receptors
Prokaryotic opsins
PTH, calcitonin, secretin receptors
metabotropic glutamate receptors
?
Neuropeptide Receptors
cyclic AMP chemoattractant receptors of Dictyostelium
The GPCR Superfamily
out
+
NH2
COOH
-
P P
P
PS S CC
S/T}
}
}C C S S
VR
LF
F
C
R
i2GDP
21
2 3 4
56
7
N--
1
2C
Chemokine Receptor Models
Hemmerich et al. Biochemistry, 38:13013, (1999)
Model of MCP-1/CCR2 Interaction
Neutrophil
IL-8
IL8RB
IL8RA
GRO
GRO GRO
NAP-2
ENA-78
CXC CC
Eosinophil
Macrophage
CK
R2b
CKR2a
CKR1
CK
R5
CKR4
MIP-1
MIP-1
MCP-3
M
CKR3
CKR1
Eotaxin
Basophil
CKR4
RANTES MCP-1
Chemokine Receptors Can Bind Many Chemokines, but are Class-restricted
i2GDP
i2GDP
21
2 3 4
56
7
N--
21
2 4
56
7
N --++
IL8
C
21
2 3 4
56
77
N--++
3
i2GTP
1
2
PI-PLC 2
?1
2
1
2
IL8
CIL8
C
Extracellular Milieu
Chemokine Receptor Activation Mechanism
Regulation
• Gene expression (cytokines/chemokines, microbes, microbial products, etc.)
• GAG association• Post-translational processing (CD26, cathepsin
G, TACE)• Endogenous antagonistic chemokines• Decoy receptors (Duffy, D6)• Receptor desensitization (phos/intern.)
Chemokines Act as Tethered Ligands
CKR
CK
Tethering mechanism: Genetic or GAGs
*Expressed on the surface of neurons, activated vascular endothelium, and in atherosclerotic lesions
*Chemoattracts monocytes, NK cells, CD8+ T
*Free and membrane bound forms exist
*Membrane-bound form mediates strong integrin-independent adhesion
*Both forms act at CX3CR1
Fractalkine (CX3CL1): an exceptional chemokine
Adapted from Rossi, et al. Genomics 1998
a b c d e f
Endothelium
WBC
aRbR
cR
Area Code forLeukocyteTrafficking
ChemoattractantsSelectins
Integrins
Stage
Flow
Rolling
Firm Adhesion
Transendothelial Migration
Activator
none
IL-1, TNF, lps
Chemoattractant
Chemoattractant
Adhesins
none
Selectins
Integrins
PECAM
Disease
none
LAD-2
LAD-1
LAD-1, LAD-2
Extravascular Space
Intravascular Space
Degranulation Chemoattractant none SGD
Multistep Paradigm for Leukocyte Trafficking
•Inflammatory
•Innate (inducible L -- constitutive R)•CXCR1 & CXCR2 Neutrophils•CCR2, CCR5 Monocytes•CCR3 Eosinophils
•Adaptive (inducible L – inducible R)•CXCR3 •CCR2 •CCR5 •CCR3•CCR9-gut homing•CCR4, CCR10-skin homing
•Homeostatic (constitutive L – constitutive R)
•CXCR4 Bone marrow•CXCR5 B cell homing to LN•CCR7 mDC & T cell homing to LN
Classification of the Chemokine System by Immunologic Function
Homeostatic
CXCL12 Bone Marrow. BCXCL13 LN-BCCL19 LN-TCCL21 LN-TCCL25 gut homingCCL27 skin homingCCL28 mucosal homing
Inflammatory
CXCL1-8 PMNCXCL9-11 Th1CCL2-5, 11 M, Eos, T, NKCX3CL1 M, Th1, NK
Classification of Chemokines by Immune Function
Lumbar Nodes
Axillary nodes
Tonsils
AdenoidsCervical Nodes
Thoracic Duct
L. Subclavian Vein
L. Lymph DuctL. Subclavian Vein
Peyer’sPatches
Appendix
Thymus Bronchial Nodes
Bone Marrow
Intercostal Nodes
Peyer’s Patches
Spleen
Iliac Nodes
Inguinal Nodes
Chemokine Receptors Regulate Effector T Cell Balance
CXCR3CX3CR1CCR1CCR2CCR5
CCR1CCR3CCR4CCR8
Th1
Th2
Host defenseAngiogenesis
chemokines
Autoimmune Disease
Hematopoiesis
InfectionAtherogenesis
Inflammation
Neoplasia
LHD
CHEMOKINES: A DOUBLE-EDGED SWORD
Chemokines/Receptors with Roles in Human Disease
CCR5 HIV/AIDS, WNV Duffy P. vivax malaria CXCR4 WHIM syndrome CXCL4 Heparin-induced thrombocytopenia(CXCR4) HIV/AIDS(CX3CR1) Atherosclerosis
Phenotypes Associated with Chemokine or Chemokine Receptor Deficiency
Abnormal Development: SDF-1/CXCR4
CXCR2CCR7/SLCCXCR5/BLC
Modulation of Inflammation
EotaxinMIP-1CCR1, 2, 3, 4, 5, 8CXCR2, 3CX3CR1
Altered susceptibility to infection
CCR1, 2, 3, 5CXCR2Duffy
Duffy and Malaria• 1950s: serologically defined as a minor blood group antigen
• 1975: pathologically defined as a virulence factor for P. vivax. Duffy is absent from the red cells of most black Africans who are thereby resistant to vivax malaria.
• 1993: molecularly defined as a 7TMD protein, and a promiscuous chemokine binding protein, binding several but not all members of both the CC and CXC groups. The Duffy mutation is in a GATA site in the promoter.
• expressed in Purkinje cells of cerebellum, post-capillary venule endothelial cells and CD45RO+ T cells
• biochemical and physiological roles undefined
Angiogenesis is Regulated by the Balance of Angiogenic and Angiostatic CXC Chemokines
ELR Non-ELR
Angiogenesis
+ -
Wound Healing Chronic Inflammation
Cancer
IFN- +-
Classification of Viral Chemokine Mimics•Structure
-chemokine-7TMD protein-unique
•Function-anti-chemokine-chemoattractant-gene expression-growth factor-membrane fusion factor (cell entry)
Receptor antagonistLigand scavenger
Host ChemokineReceptors (7TM)
Host ChemokineBinding Proteins (7TM)
Viral Chemokine Binding Proteins (non-7TM)
Viral Chemokine Receptors (7TM)
Human & Viral Orphan Receptors
Host Chemokines
(-)
?
(-)
(+)
(+)
Non-chemokineViral Ligands for
Chemokine Receptors
Viral Chemokines
Non-chemokine Host Ligands for
Chemokine Receptors
(+/-)
(+/-)
(+/-)
The Extended Chemokine Family
Host Chemokine Receptors
HIV-1 Infection
HIV-1 Progression
HIV and Chemokine Mimicry
TCL PBMC M
CD4 CD4 CD4
T
HIV-1 Cytotropism and Pathogenesis
AIDSInitial Infection
M
M-tropicT-tropic
HIV-1
EnvHIV-1
Target Cell
gp120 gp120
gp
41 g
p4
1
Target Cell
CD4
Chemokine Receptor
gp
12
0
Genome
gp41
Exploitation of Chemokine Receptors by HIV
Target Cell
CXCR4 CCR5
CD4
CD4
CXCL12
CCL3CCL4CCL5
X4
HIV
R5
12
34
5CCR532
CCR5 & CXCR4 and HIV Pathogenesis
RBDEU HIV+
Fre
qu
ency
of
CC
R5
-/-
(%)
Comparison Groups
5
4
3
2
1
CCR5 is Required for Efficient HIV-1 Transmission: Evidence from Population Studies of the Inactive CCR5 Allele, CCR5 delta 32
(~ZERO)
p<0.001
n= 111 387 614
Chemokines & Cancer Metastasis
Breast Blood Target Organ
Murphy (2001) NEJM; 345:833-835