Role of OVOL2 and miR-126 in ETV2-regulated cardiovascular development Restricted; Files Only

Kim, Juyoung (Fall 2018)

Permanent URL: https://etd.library.emory.edu/concern/etds/df65v865d?locale=en
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Abstract

FLK1, the receptor for VEGF (vascular endothelial growth factor), has been identified as an essential factor for cardiovascular development and post-natal life. Accordingly, extensive efforts have been made to understand FLK1-mediated signaling, but the underlying mechanisms of Flk1 gene regulation remain largely unknown. In this regard, my laboratory revealed for the first time the critical function of ETV2, a member of the ETS transcription factor family, for cardiovascular development. Further, ETV2 was identified as a direct upstream regulator for Flk1 gene as well as other genes critical for both vascular endothelial and hematopoietic cell lineages development.

 

To understand the molecular mechanisms of ETV2 in regulating the development of the cardiovascular system, I set out two independent, but closely related projects for my Ph.D. thesis. First, I sought to identify the proteins that can interact with ETV2, since transcription factors often form the transcriptional complex with other transcription regulatory proteins to regulate the expression of the target genes. I revealed that ETV2 can directly interact with OVOL2, a zinc finger transcription factor. The interaction of ETV2-OVOL2 cooperatively regulates not only for the generation of FLK1+ cells, but also the generation of vascular endothelial and hematopoietic cell lineages from mouse embryonic stem cells (mESCs). Second, I performed experiments to identify ETV2-regulated microRNA (miRNA) using next generation sequencing and found a subset of miRNAs regulated by ETV2. These miRNAs were closely related to cardiovascular development; particularly miR-126 was of special interest, since my experiments showed that the expression of miR-126 was directly activated by ETV2. Further, I demonstrated that the miR-126/MAPK pathway is critical for the generation of FLK1+ cells from mESCs. Mechanistically, c-JUN/FOS (AP1 complex), a downstream of the MAPK pathway, binds to the enhancer region of Flk1 via AP1 binding sequences, leading to the activation of Flk1 expression.  

In my dissertation, I have revealed the novel functions of ETV2; the interaction of ETV2-OVOL2 and the role of the miR-126/MAPK pathway in ETV2-mediated FLK1+ cell generation. Therefore, these findings will advance our understanding on cardiovascular development, which could provide fundamental research directions for designing therapeutic approaches for cardiovascular disease. 

Table of Contents

Table of Contents

Abstract

 

Acknowledgements

Chapter 1: Introduction……………………………………………………………...….1

An Overview on ETV2 in hematopoietic and endothelial cell development…………......2

ETV2 is essential for vascular endothelial and hematopoietic cell development……........3

Molecular mechanisms of ETV2 in regulating cardiovascular development……………..5

Interplay between ETV2 and other ETS factors in establishing cardiovascular system….8

Cardiovascular cell fate determining role of ETV2……………………………………...10

Figure 1…………………………………………………………………………………..13

Figure legends……………………………………………………………………………14

Chapter II: OVOL2 is a critical regulator of ER71 in generating FLK1+ cells, hematopoietic and endothelial cells from embryonic stem cells……………………..15

 

Abstract…………………………………………………………………………………..18

 

Introduction………………………………………………………………………………19

 

Methods…………………………………………………………………………………..20

 

Results……………………………….…………………………………………………...28

 

Figures…...……………………………………………………………………………….34

 

Figure legends……………………………………………………………………………44

 

Supplemental tables……………………………………………………………………...47

 

Supplemental figures and legends……………………………………………………….50

 

Chapter III: ETV2/ER71 regulates the generation of FLK1+ cells from mouse embryonic stem cells through miR126-MAPK signaling…………………………….59

Abstract…………………………………………………………………………………..61

Introduction………………………………………………………………………………62

Materials and Methods…………………………………………………………………...65

Results……………………………………………………………………………………72

Figures……………………………………………………………………………………78

Table ……….……………………………………………………………………………90

Figure legends……………………………………………………………………………91

Chapter IV: Discussion and Perspective………………………………………………93

Discussion………………………………………………………………………………..94

Perspective……………………………………………………………………….……..100

References……………………………………………………………………………...103

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