The Role of CD86 Trafficking and Signaling on Myeloma Survival and Proliferation 公开

Abstract

Multiple myeloma is an incurable hematologic malignancy of long-lived antibody secreting plasma cells. Myeloma cells retain numerous features of plasma cell biology including a reliance on signals within the bone marrow microenvironment. In advanced stages, myeloma cells can become extramedullary and survive independently of the bone marrow due to autocrine cytokine signals and cell-cell interactions. One such physical interaction involves the CD28-CD86 module which signals bi-directionally to confer myeloma cell survival and drug resistance. My studies investigate how myeloma cells regulate this interaction and the expression of the CD86 at the plasma membrane. I also demonstrated a role for CD86 in proliferation and further elucidated downstream molecular changes induced by CD28 signaling.

I identified that while several regions of the tail are required for proper trafficking of CD86 out of the Golgi, a specific region for proper transport is a three amino acid-long PDZ binding motif at the C-terminus of the tail. BioID analysis uncovered two PDZ-domain containing proteins, SCRIB and DLG1 as proximal to the CD86 cytoplasmic tail. Deletion of SCRIB and DLG1 in myeloma cell lines results in a decrease in CD86 surface expression, myeloma proliferation and viability. These proteins are important for generating CD86 to the surface where it binds to CD28 and is stabilized. My studies also expand upon the role for CD86 in myeloma, showing a role for CD86 in myeloma proliferation and IMiD resistance. Furthermore, they demonstrate a potential CD28 pro-survival signaling mechanism via SYNTENIN upregulation and may reveal a compensatory role for the ICOS-L-CD28 axis in the absence of CD86.

Table of Contents

Chapter 1: Introduction 1

Abstract 2

Introduction to the Game of Bones 3

Spectrum of Plasma cell dyscrasias 3

The role of the bone marrow microenvironment 9

Myeloma takeover beyond the microenvironment 22

Conclusion 27

Initial Treatment Strategies 29

CD28-CD86 Signaling 33

References 37

Table 1: Defining Stages of Myeloma 49

Figure 1: Bone marrow interactions that promote

myeloma growth and survival. 51

Figure 2: Models for leukocyte and myeloma cell

extravasation. 53

Figure 3: Models for leukocyte and myeloma cell

extravasation. 55

Figure 4: CD28-CD86 binding in different immune

cell types. 56

Chapter 2: PDZ proteins, SCRIB and DLG1, regulate

myeloma surface CD86 expression, growth, and survival 58

Abstract 59

Introduction 60

Materials and Methods 62

Results 68

Discussion 79

Acknowledgments 83

Bibliography 85

Figure 1: CD86 cytoplasmic tail is important for

trafficking to cell surface 95

Figure 2: Multiple Regions of CD86 cytoplasmic

tail are important for trafficking to cell surface 98

Figure 3: CD86 contains a PDZ binding motif

important for surface expression 100

Figure 4: BioID proximity assay identifies numerous CD86

cytoplasmic tail interacting partners 102

Figure 5: SCRIB and DLG1 regulate CD86 surface expression 104

Figure 6: SCRIB and DLG1 are important for cell

growth and viability 106

Figure 7: SCRIB and DLG1 regulate CD86 prosurvival

Signaling 108

Visual Overview: 110

Supplemental Methods 111

Supplementary Table 1 112

Supplementary Figure 1 113

Supplementary Figure 2 115

Supplementary Figure 3 117

Supplementary Figure 4 119

Supplementary Figure 5 120

Supplementary Figure 6 122

Supplementary Figure 7 124

Supplemental Bibliography 125

Chapter 3: Differential expression of CD28

interacting partners CD86 and ICOS-L induces

molecular changes in myeloma cells 126

Introduction 127

Materials and Methods 128

Results 131

Figure 1: Overexpression of CD86FL and TL

results in increased syntenin expression. 132

Figure 2: Silencing of CD86 results in increased

syntenin expression. 134

Figure 3: ICOS-L does not contain a PDZ-binding motif. 135

Figure 4: ICOS-L is upregulated with CD86 and

SYNTENIN silencing. 136

Figure 5: ICOS-L has decreased expression in

CD86-overexpressing cell lines. 137

Discussion 137

Bibliography 141

Chapter 4: The Role of CD28 and CD86 in myeloma cell

Proliferation 143

Introduction 144

Materials and Methods 145

Results 147

Figure 1: Dividing cells express higher CD86. 148

Figure 2: Lenalidomide treatment decreases CD28 and

CD86 expression levels. 149

Discussion 149

Bibliography 151

Chapter 5: Discussion 153

Summary 154

Figure 1: Summary of main research findings. 158

Figure 2: Myeloma CD86 expression and IMiD outcome. 161

Future Directions 165

Concluding Remarks 170

Bibliography 172

About this Dissertation

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• Permission granted by the author to include this thesis or dissertation in this repository. All rights reserved by the author. Please contact the author for information regarding the reproduction and use of this thesis or dissertation.

School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Biochemistry, Cell & Developmental Biology
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Biology, General Biology, Molecular Biology, Cell
关键词	CD86 Myeloma Cancer
Committee Chair / Thesis Advisor	Boise, Lawrence, Emory University
Committee Members	Ford, Mandy, Emory University Seyfried, Nicholas, Emory University Moberg, Kenneth, Emory University Koval, Michael, Emory University

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