Fc Receptor Signaling to the Nucleus CARLOS ROSALES LEDEZMA - - PowerPoint PPT Presentation

Fc Receptor Signaling to the Nucleus

Instituto de Investigaciones Biomédicas Universidad Nacional Autónoma de México

CARLOS ROSALES LEDEZMA

Receptors for Fc part of IgG

Berken & Benacerraf

• J. Exp. Med.1966; 123:119

Macrophage Ab

Fc Receptor Functions

Mast cell IgE FcεR Macrophage IgG FcγR Phagocytosis ADCC Respiratory Burst Degranulation Inflammatory Mediators Gene Activation Cytokine Production

Human IgG Fc Receptors

FcγRI FcγRIIA FcγRIIB FcγRIIIA FcγRIIIB

CD64 CD32 CD32 CD16 CD16 γ2 α α α α α α-GPI γ2 γ2 β

Macrophage Neutrophil Eosinophil Dendritic Cell Macrophage Neutrophil Mast Cell Eosinophil Platelet Dendritic Cell Macrophage Neutrophil Mast Cell Eosinophil Dendritic Cell B Cell Macrophage Mast Cell Basophils NK Cell Dendritic Cell Mast Cell Basophil Neutrophil ITAM ITIM

• Annu. Rev. Immunol. 1991; 9:457
• Annu. Rev. Immunol. 1997; 15:203

FcR signaling (~1996)

Nucleus DNA IL-1, IL-8 TNF

?

Y-kinase Fc receptor Membrane

NF-κB NF-κB IκB

Ras signaling

RTK Membrane GAP Grb2 Sos Ras GDP Ras GTP Raf

SRE

Nucleus DNA Gene expression

MEK ERK Elk-1

Assay for NF-κB activation

Luciferase mRNA luc Gene Fc receptor THP-1 Membrane

NF-κB IκB NF-κB

Dominant negative Ras does not inhibit NF-κB activation

1 2 3 4 5 pZIP Ras N17

Luciferase Activity

(Fold Increase)

Medium

IIC

Dominant negative Raf does not inhibit NF-κB activation

1 2 3 4 5 6

pCGN Raf 23-284

Medium

IIC

Luciferase Activity

(Fold Increase)

MEK is required for immune complex-mediated NF-κB activation

1 2 3 4

Medium IIC IIC + PD98059

Luciferase Activity

(Fold Increase)

FcR signaling

Nucleus DNA

?

Fc receptor THP-1

NF-κB NF-κB IκB Syk MEK ERK

• J. Biol. Chem. 1998; 273: 27610

Gene Expression

ERK activation requires PI 3-K

• phospho-ERK

MEK activation depends on PI 3-K

PI 3-K is required for NF-κB activation

0.5 1 1.5 2 2.5 3 3.5 4

Luciferase Activity

Medium IIC IIC + Wor IIC + LY

FcR signaling

Nucleus DNA Fc receptor THP-1

NF-κB NF-κB IκB Syk MEK ERK

• J. Leukoc. Biol. 2001; 70: 649

Gene Expression PI3K

Neutrophil Fc Receptors

FcγRIIA FcγRIIIB

CD32 CD16 α α-GPI

• J. Leukoc. Biol. 2005; 15: 203

Phagocytosis Calcium Rise Degranulation Respiratory Burst Calcium Rise Respiratory Burst Actin assembly Integrin activation

• J. Biol. Chem. 1992; 267: 5265
• Biochem. Biophys. Acta 1999; 1452: 46

Are FcRIIA and FcRIIIB Signaling Different?

FcγRIIIB

?

PMN

FcγRIIA

Nuclear Factor Activation

PMN

FcγR

mAb NF-κB IκB

FcγR

NF-κB

• J. Immunol. Meth. 2007; 320: 104

Nuclear Factor Activation

FcγR

NF-κB NF-κB NF-κB FITC FACS

• J. Immunol. Meth. 2007; 320: 104

NF-κB Activation

0.5 1 1.5 2 2.5 3 3.5 4

• IV.3

3G8 Nuclear Staining (Fold Increase)

Phagocytosis

0.5 1 1.5 2 2.5 3 3.5 4 4.5 5 FcRIIA FcRIIIB Fold Increase

• IV.3

3G8

FcRIIIB signal to NF-κB is independent of Syk and MEK

0.5 1 1.5 2 2.5 3 3.5 4 IV.3 3G8 Nuclear Staining (Fold Increase) DMSO Piceatannol PD98059

Signaling to Nucleus in PMN

FcγRIIA FcγRIIIB

Syk PI3K MEK ERK NF-κB NF-κB IκB PMN Nucleus

Signaling to Nucleus in PMN

FcγRIIA FcγRIIIB

Syk PI3K MEK ERK NF-κB NF-κB IκB PMN Nucleus ERK P

?

P

Nuclear phospho-ERK

0.5 1 1.5 2 2.5 3 3.5 4 4.5 IV.3 3G8 Nuclear Staining (Fold Increase)

Nuclear phospho-ERK

ERK

• 3G8

F(ab’)2 3G8 IV.3 p-ERK

Nuclear phopho-ERK is independent Syk, MEK, and PI 3-K

1 2 3 4 5 6 DMSO Piceatannol PD98059 Wortmannin Nuclear Staining (Fold Increase) 3G8

Signaling to Nucleus in PMN

FcγRIIA FcγRIIIB

Syk PI3K MEK ERK NF-κB NF-κB IκB PMN Nucleus ERK P

Nuclear ERK does not change after FcγRIIIB crosslinking

Nuclear ERK

MEK is not present in PMN nucleus

Nuclear MEK

Nuclear phospho-Elk-1

0.5 1 1.5 2 2.5 3 3.5 4 4.5 IV.3 3G8 Nuclear Staining (Fold Increase)

Nuclear phospho-Elk-1

p-Elk-1 ERK

• 3G8

F(ab’)2 3G8

Nuclear phopho-Elk-1 is independent Syk, MEK, and PI 3-K

1 2 3 4 5 6 DMSO Piceatannol PD98059 Wortmannin Nuclear Staining (Fold Increase) 3G8

FcγR activation of ERK in PMN cytoplasm

FcγRIIA FcγRIIIB

Syk PI3K MEK ERK NF-κB NF-κB IκB PMN Nucleus ERK P

?

FcγRIIIB activates nuclear factors in PMN

FcγRIIA FcγRIIIB

NF-κB NF-κB IκB PMN Nucleus ERK P Elk-1 P

Conclusions

• FcγRIIA and FcγRIIIB signal

differently in the PMN

• FcγRIIA induces efficient

phagocytosis

• FcγRIIIB induces efficient ERK and

Elk-1 activation

• FcγRIIIB signal transduction does

not involve Syk, MEK, PI 3-K, Src family kinases, or Tec family kinases

Acknowledgments

• Gabriela Sánchez-Mejorada
• Elsa Merit Reyes
• Erick García
• Nancy Mora
• Alejandro Ortiz Stern
• Georgina Nieto
• Melissa Ruiz
• Marco Hernández
• Selma Rivas
• Victor Salas