LSD1 regulates marginal zone B cell development, B cell proliferation, and plasmablast differentiation Restricted; Files Only

Abstract

LSD1 is a histone demethylase that primarily targets H3K4me1/2 and H3K9me1/2, leading to transcriptional repression and activation, respectively. Through its demethylase function, it promotes cellular processes such as autophagy, cell cycle progression, and inflammation, and drives the development and differentiation of multiple cell types including adipocytes, embryonic stem cells, blood cells, myocytes, neurons, and gametocytes. Additionally, LSD1 has been shown to contribute to human diseases such as cancer and viral infection. Despite the extensive research on the role of LSD1 throughout normal and disease pathways, much remains to be discovered. This thesis will focus on the role of LSD1 in B cell development and differentiation into antibody-secreting plasma cells. A 2009 study showed that LSD1 directly interacted with the key plasma cell transcription factor Blimp-1, suggesting an in vivo role for LSD1 during B cell differentiation. Here I show that B cell-conditional deletion of LSD1 in mice results in diminished B cell proliferation and differentiation in response to the antigen lipopolysaccharide (LPS). Genome-wide transcriptome and chromatin accessibility analyses showed that LSD1 repressed hundreds of genes in LPS-induced plasma cells. These genes were in close proximity to binding sites of the key B cell differentiation transcription factors Blimp-1, IRF4, and PU.1. LSD1 suppressed chromatin accessibility and H3K4me1 at these target binding sites, implying that LSD1 directly regulates multiple transcription factor networks throughout B cell differentiation. Quantification of developing B cell populations revealed that LSD1-deficient bone marrow B cell development is normal, but marginal zone B cell development in the spleen is impaired. Similar to its role in B cell differentiation, LSD1 repressed hundreds of genes in marginal zone B cells. Chromatin accessibility analysis showed that LSD1 repressed accessibility at the binding sites of transcription factors involved in splenic B cell development, including NF-kB. In vitro marginal zone B cell development experiments solidified a key role for LSD1 in regulating non-canonical NF-kB signaling induced by BAFF. Indeed, LSD1 directly interacted with the non-canonical NF-kB transcription factor p52. Overall, these studies not only define LSD1 as a critical epigenetic and transcriptional regulator during B cell development and differentiation, but also provide novel mechanistic insights into how LSD1 regulates these processes. The revealing of this new B cell branch of LSD1 function will be critical to understanding how epigenetic modifying proteins contribute to B cell-based diseases and may give rise to innovative treatment options for such diseases.

Table of Contents

Chapter 1: Introduction to LSD1 ……………………………………………………..........1

Part A – The role of LSD1 throughout cellular development, differentiation, function, and disease…………….2

           Rationale…………………………………………………………………..2

Biochemical and functional characterization of LSD1………….................2

           Cellular development, differentiation, and function……………….............8

                       Adipogenesis and fat cell function…………………........................8

                       Autophagy…………………………………………………………9

                       Cell cycle………………………………………............................11

                       Embryogenesis…………………………………...........................12

                       Hematopoiesis and blood cell function…………………………...14

                       Inflammation……………………………………………………..16

                       Inner ear development…………………………............................17

                       Myogenesis and muscle cell function…………………………….18

                       Neurogenesis and neuron/brain function………............................19

                       Spermatogenesis……………………………………………........23

                       Other pathways…………………………………………………...24

           Disease and its treatment………………………………………................25

                       Cancer……………………………………………………………25

                       Genetic disease resulting from KDM1A mutation………………..27

                       Neurodegenerative disease……………………………………….28

                       Sickle cell disease………………………………...........................29

                       Viral infection……………………………………………………30

                       Pharmacological targeting………………………………………..32

           Part B – B cell development and differentiation……………………….................36

                       B cell function…………………………………………………................36

                       B cell development…………………………………………….................37

                       B cell differentiation……………………………………………………...40

Chapter 2: LSD1 regulates B cell proliferation and plasmablast differentiation………….42

           Abstract…………………………………………………………………………..43

           Introduction………………………………………………………………………44

           H3K4 methylation is remodeled during B cell differentiation……………………46

           LSD1 is required for normal plasmablast formation……………………………..48

           LSD1 regulates the plasmablast transcriptional program………………………...52

           Blimp-1 target repressed genes are regulated by LSD1…………………………..56

           LSD1 promotes B cell proliferation………………………………………………57

           LSD1 regulates chromatin accessibility at ETS and IRF transcription factor motifs…60

           LSD1 restricts chromatin accessibility at naïve B cell enhancers in plasmablasts..62

           Aberrant accumulation of H3K4me1 at LSD1-regulated loci……………………67

Chapter 3: LSD1 cooperates with NF-κB to regulate marginal zone B cell development..72

           Abstract…………………………………………………………………………..73

           Introduction………………………………………………………………………74

           LSD1 regulates marginal zone B cell development………………………………76

           LSD1 functions as a transcriptional repressor in marginal zone B cells…………..82

           LSD1 represses chromatin accessibility at NF-kB motifs………………………..86

LSD1 regulates ex vivo marginal zone B cell development induced by NOTCH2 and non-canonical NF-kB signaling………..93

           LSD1 and NF-kB cooperate to regulate marginal zone B cell development……..99

Chapter 4: Work in Progress…………………………………………………………….104

Chapter 5: Discussion…………………………………………………………………...108

Bibliography……………………………………………………………………………116

About this Dissertation

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Genetics and Molecular Biology
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Biology, Genetics Biology, Microbiology Biology, Molecular
Keyword	LSD1 Genetics B cells Immunology
Committee Chair / Thesis Advisor	Jeremy M. Boss, Emory University
Committee Members	Roger B. Deal, Emory University Ignacio Sanz, Emory University Paula M. Vertino, University of Rochester Jacob E. Kohlmeier, Emory University

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