Three-dimensional chromatin structure and its role in cellular function Open Access
Sauria, Michael Everett Gehring (2014)
Abstract
Cellular function is controlled by a complex interplay between genomic sequence and its surrounding context. A large force in establishing genomic context is the physical partitioning of the genome into defined neighborhoods that allows coordination of transcriptional activity, DNA interactions with RNA and proteins, and chemical modifications. Spatial organization is also involved in X-inactivation, cell fate determination, and senescence. Recent high-throughput resequencing technologies have allowed investigation of chromatin architecture on a scale and resolution overcoming the previous limits of microscopy and inference at individual loci from less direct assessments. It is now possible to create a genome-wide map of DNA fragment interactions or investigate a protein-specific DNA interaction network. These approaches have revealed a complex hierarchical organization ranging from whole chromosomes down to short-range associations between adjacent features. Because these technologies generate large amounts of data representing a complex system, pose significant computational and analytical challenges. We developed HiFive, a framework for analyzing HiC and 5C data, to address these challenges. HiFive allows handling of large amounts of data in an efficient manner and easy access to subsets of data for downstream analysis and plotting. We have also included an approximation approach to normalization that allows processing of data for a fraction of the computational cost and time. Compared to other available methodologies, HiFive performs as well or better across a variety of measures. To further validate the approaches used in HiFive, we also present downstream analyses locating significant structural signatures and analyzing gene spatial arrangements. We are able to increase sensitivity to detection of subdomain structures and their associated features. We also present a new approach to three-dimensional modeling that reveals a spatial partitioning of genes organized around transcriptional activity. Our results are consistent with our current understanding of chromatin architecture and suggest exciting possible avenues for future exploration.
Table of Contents
Chapter 1 - Introduction 1
Organization of the animal nucleus 1
Function of the nucleus 1
Chromosome territories 2
The fractal globule 4
Topological domains 4
Organization of the nucleus 6
The nucleolus 7
Polycomb bodies 8
Lamina associated domains 9
RNA polymerase II-dependent transcriptional foci 11
Assaying chromatin structure 14
Microscopic detection of chromatin structures 14
Chromatin conformation capture 15
3C-derived approaches 18
PMLA data analysis 20
Chapter 2 - Determining chromatin conformation from interaction frequency data using a probabilistic modeling approach 22
Introduction 22
Material and Methods 25
Mapping of interaction data 25
Software Implementation 26
Data Filtering 28
Estimation of Distance-Dependent Signal 33
HiFive Data Normalization 37
HiFive-Express Iterative Approximation for Bias Correction 40
Neighboring Fend Correlations 42
Results 43
HiC Unit of Interaction 43
HiFive's 5C Normalization Performance 45
HiFive's HiC Normalization Performance 49
Discussion 54
Chapter 3 - Validation of HiFive through method comparisons and biological findings 57
Introduction 57
Materials and methods 59
Acquiring and Mapping Data 59
5C Data Normalization with HiFive 61
5C Data Normalization with Alternate Methods 62
HiC Data Normalization with HiFive 63
HiC Data Normalization with Alternate Methods 63
Annotation Data Processing 65
Dynamic Binning 65
5C Data Correlations with HiC Data 67
HiC Inter-Dataset Correlations 68
Calculating the boundary index 68
Boundary index comparison to the directionality index 71
Three-dimensional chromatin modeling 72
Calculating gene spatial arrangements 75
Results 76
5C Method Comparison 76
HiC Method Comparison 78
The boundary index captures more significant features than the directionality index 79
Three dimensional chromatin models 81
Spatial partitioning of genes by transcriptional activity 88
Discussion 91
Chapter 4 - Discussion 95
Explaining nuclear organization 95
Areas of future inquiry 98
Delineating the conservation of boundaries 98
Defining boundary types and elements 100
Deciphering between targeted and stochastic association 102
Applications for chromatin structural understanding 106
Associations between chromatin structure and disease 106
Synthetic biology 108
Conclusions 110
Chapter 5 - References 111
Chapter 6 - Non-printed sources 128
Chapter 7 - Abbreviations 130
About this Dissertation
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hifive_documentation.pdf () | 2018-08-28 14:12:17 -0400 |
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