Genome-wide studies of transcriptional dynamics in fear learning Open Access

Sharma, Sumeet (Fall 2020)

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The consolidation of fear-related memories requires synaptic plasticity dynamics in many brain regions, with the basolateral amygdala (BLA) thought to be the site where cue associations with fearful physiological outputs are made. Herein we employed mRNA-sequencing and 5-hydroxymethylcytosine profiling to obtain a genome-wide view of transcriptional and epigenetic dynamics in the BLA during fear memory consolidation. We find that differentially hydroxymethylated regions (DHMRs) reside primarily within introns, exons, enhancer elements, mitochondrial DNA, and repetitive DNA. We find 727 genes to be dynamically expressed (DE), of which 108 also contain differentially hydroxymethylated regions (DHMRs). These DE genes encompass pathways in mitochondrial function, gene transcription, protein translation, and heat shock response. Within the DHMR-containing DE genes, we replicate Fkbp5 dynamics in fear conditioning. Fkbp5 is a well-known gene previously implicated in stress responses, fear processing, and fear- and stress-related disorders in humans. We find that Fkbp5 is modulated at both the transcriptional and DNA methylation level, and we identify CHH-context cytosines within intron 1 that are casual drivers of Fkbp5 gene regulation. Finally, to better examine the dynamics of transcriptional dynamics in fear processing, we demonstrate the development of a robust novel behavioral procedure that results in stress enhanced fear learning and stress-dose dependent alterations in anxiety.

Table of Contents

I. Chapter 1. Introduction and Historical Background: Introduction to the study of gene pathways in PTSD and genetic dissection of learning and memory pathways (pg. 12-41)

a. Translational approaches to understanding disease (pg. 12-13)

b. Genetic Epidemiology (pg. 13-16)

c. Genetic Association Studies (pg. 16-19)

d. PTSD GWAS (pg. 19-21)

e. PTSD GxE Studies (pg. 21-23)

f. FKBP5 in genetic association studies (pg. 23-27)

g. Memory and Epigenetics (pg. 27-30)

h. Methylome Editing (pg. 30-31)

i. Figures (pg. 32-33)

j. References (pg. 34-41)

II. Chapter 2: Epigenetic and transcriptomic profiling of the BLA during fear memory consolidation: Unbiased approaches to elucidating molecular pathways associated with fear learning (pg. 42-86)

a. Introduction (pg. 42-44)

b. Results (pg. 44-54)

c. Discussion (pg. 54-57)

d. Methods (pg. 58-65)

e. Figures and Tables (pg. 66-80)

f. References (pg. 81-86)

III. Chapter 3: Behavioral model of a Stress x Fear Interaction: development of a behavioral paradigm exhibiting a significant interaction between stress and fear learning. (pg. 87-111)

a. Introduction (pg. 87-91)

b. Results (pg. 91-95)

c. Discussion (pg. 95-96)

d. Methods (pg. 96-100)

e. Figures (pg. 101-106)

f. References (pg. 107-111)

IV. Chapter 4: Targeted Profiling Fkbp5 in stress-enhanced fear learning: Transcriptional, epigenetic, and evaluation of putative GREs (pg. 112-141)

a. Introduction (pg. 112)

b. Results (pg. 113-117)

c. Discussion (pg. 117-118)

d. Tables and Figures (pg. 119-126)

e. References (pg. 127-141)

V. Discussion (pg. 142-)

a. Chapter 2. Findings, implications, limitations, future directions (pg. 142-145)

b. Chapter 3. Findings, implications, limitations, future directions (pg. 145-146)

c. Chapter 4. Findings, implications, limitations, future directions (pg. 147-148)

d. Conclusion (pg. 149-151)

e. Combined Future Directions (pg. 152-153)

f. Figures (pg. 154-155)

g. References (pg. 156)

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