Interplay of Notch, BCR, and CD40 signaling during B cell differentiation, activation, and lymphomagenesis [Elektronische Ressource] / Franziska Hampel. Betreuer: Dirk Eick

Franziska Hampel

Sujets

• Gesundheit

Informations

• Publié par: ludwig-maximilians-universitat_munchen
• Publié le: 01 janvier 2010
• Nombre de lectures: 37
• Langue: Deutsch
• Poids de l'ouvrage: 2 Mo

Extrait

Interplay of Notch, BCR, and CD40 signaling
during B cell differentiation, activation, and
lymphomagenesis





Dissertation der Fakultät für Biologie
der Ludwig-Maximilians-Universität München
zur Erlangung des Doktorgrades






Franziska Hampel

April 2010


















To Thomas
& my parents


Erklärung
Diese Dissertation wurde von Mai 2007 bis April 2010 am Institut für Klinische
Molekularbiologie und Tumorgenetik des Helmholtz Zentrums München in der Arbeitsgruppe
von PD Dr. Ursula Zimber-Strobl erstellt und von Prof. Dr. Dirk Eick betreut.










Eidesstattliche Versicherung
Hiermit erkläre ich, Franziska Hampel, geboren am 7. April 1982 in Fürstenfeldbruck,
ehrenwörtlich, dass ich die vorliegende Arbeit selbständig und nur unter Verwendung der
angegebenen Hilfsmittel und Quellen angefertigt habe.

München, den 13. April 2010



Franziska Hampel











Erstgutachter: Prof. Dr. Dirk Eick
Zweitgutachterin: Prof. Dr. Elisabeth Weiß

Tag der Einreichung: 13. April 2010
Tag der mündlichen Prüfung: 1. Oktober 2010

TABLE OF CONTENTS
Table of contents...............................................................................................................I
List of figures ...................................................................................................................V
List of tables.....................................................................................................................V
List of abbreviations....................................................................................................... VI
1 INTRODUCTION ............................................................................................................1
1.1. The immune system1
1.1.1 T cell development..................1
1.1.2 Early B cell development ..............................................................................................................2
1.1.3 Late B cell development – Marginal zone versus follicular B cell differentiation ................3
1.1.4 B cell activation...............................................................................................................................5
1.2 Central signaling pathways in B lymphocytes ................................................................6
1.2.1 CD40 receptor signaling................................................................................................................7
1.2.2 NF-κB signaling..............................................................................................................................7
1.2.3 B cell receptor signaling.................................................................................................................9
1.3 The role of the Notch pathway in lymphocytes............................................................ 11
1.3.1 The Notch signaling cascade ......................................................................................................11
1.3.2 Notch signaling in lymphocytes .................................................................................................13
1.3.3 The role of Notch in marginal zone B cell development.......................................................13
1.3.4 Notch signaling in lymphomagenesis........................................................................................15
1.4 Interplay of signaling pathways in B cell lymphomas .................................................. 16
1.5 Model systems ...............................................................................................................17
1.5.1 The Notch1/2IC-expressing EREB2.5 cell lines ....................................................................17
1.5.2 The LMP1/CD40-transgenic mouse strain..............................................................................19
1.5.3 The Ig β-deficient mouse strain ..................................................................................................19
2 GOAL ............................................................................................................................... 21
3 RESULTS.........................................................................................................................22
3.1 Comparison of Notch1IC, Notch2IC, and EBNA2 target genes in B cells in vitro.....22
3.1.1 The experimental setting .............................................................................................................22
3.1.2 Notch1/2 lead to cell cycle entry due to the induction of cell cycle-associated genes ......22
3.1.3 Differential expression of pro-apoptotic and anti-apoptotic genes in Notch1IC-,
Notch2IC-, and EBNA2-expressing B cells ......................................................................................24
I
3.2 The role of Notch2 in B cell differentiation in vivo ......................................................27
3.2.1 Generation of a transgenic mouse line conditionally expressing Notch2IC .......................27
3.2.2 Notch2IC protein is translocated into the nucleus and is functional in B cells of
+/-Notch2IC//CD19Cre mice .............................................................................................................32
+/-3.2.3 Early B cell development is blocked in Notch2IC//mb1Cre mice..................................33
3.2.4 Notch2IC expression dependent on mb1-Cre causes an aberrant T cell-like differentiation
in the bone marrow..................34
+/-3.2.5 Early B cell development is normal in Notch2IC//CD19Cre mice ................................37
+/-3.2.6 Marginal zone B cells are the dominant B cell population in Notch2IC//CD19Cre
mice..........................................................................................................................................................38
3.2.7 Notch2IC-expressing B cells are located in the marginal zone and display a pre-activated
phenotype................................................................................................................................................43
+/-3.2.8 Analysis of marginal zone B cell precursors in Notch2IC//CD19Cre mice ..................45
+/-3.2.9 Notch2IC//CD19Cre B cells are hyper-responsive to LPS and α-CD40 stimulation in
vitro but do not form germinal centers................................................................................................47
+/-3.2.10 Signaling pathways active in Notch2IC//CD19Cre B cells ............................................50
3.2.11 Constitutive Notch2 signaling overcomes CD19 deficiency during marginal zone B cell
differentiation.........................................................................................................................................53
3.3 The role of B cell receptor signaling in LMP1/CD40-activated B cells.......................58
3.3.1 Genetic ablation of Igβ in LMP1/CD40-transgenic mice .....................................................58
3.3.2 Constitutive CD40 signaling rescues Ig β-deficient B cells to a certain extent ....................59
ΔGFP/ Δ3.3.3 LMP1/CD40//Igβ B cells display an activated phenotype.........................................60
3.3.4 LMP1/CD40 expression prolongs the survival of Ig β-deficient B cells in vitro and in vivo
..................................................................................................................................................................61
ΔGFP/ Δ3.3.5 Analysis of signaling pathways in LMP1/CD40//Ig β B cells.....................................63
4 DISCUSSION ..................................................................................................................67
4.1 Notch1/2IC activate proliferation-associated as well as pro-apoptotic genes in human
B cells in vitro......................................................................................................................67
4.2 Constitutive Notch2 signaling is instructive for marginal zone B cell differentiation in
vivo ......................................................................................................................................69
4.2.1 Notch2IC expression dependent on mb1-Cre blocks early B cell differentiation and leads
to ectopic T cell development in the bone marrow..........................................................................69
4.2.2 Notch2IC expression dependent on CD19-Cre induces marginal zone B cell
differentiation at the expense of follicular B cells.............................................................................71
II
4.2.3 The basal activity of the non-canonical NF- κB pathway is reduced in Notch2IC-
expressing B cells ...................................................................................................................................74
+/-4.2.4 Notch2IC//CD19Cre B cells display high c-Myc levels as well as enhanced MAPK and
PI3K/Akt activity......