Genomic and microscopic dissection of nuclear compartmentalization and function

The spatial genome organization in the cell nucleus is non-random and important for gene expression regulation. Abnormal nuclear organization is related to disease. Microscopy measurements of individual genes reveal a population-based, statistical gradient of increased gene activity from the nuclear periphery to the center. Sequencing-based methods (Hi-C, DamID) suggest the genome could be partitioned into two compartments: transcriptionally active and inactive chromatin positioned at the interior and periphery of the nucleus, respectively. Newer genomic methods (SPRITE, GAM) suggest multi-way interactions occurring more frequently in the nuclear interior, specifically at nucleoli and nuclear speckles. But it is still elusive how such nuclear compartments (bodies) are involved in genome organization. Nuclear speckles are a type of nuclear body enriched in RNA metabolic factors and correlated with active gene expression. A first genome-wide mapping of chromosomal distances from nuclear speckles by a novel genomic method developed in our lab, TSA-Seq, reveals ~5% of the genome deterministically positions at the periphery of speckles, as one major gene expression “hot-zone” in the nuclear interior. Yet the function and dynamics of genome organization relative to nuclear speckles are still unclear. In this dissertation, I applied TSA-Seq combined with microscopy to dissect genome organization relative to nuclear compartments (speckles, lamina, and nucleolus), and its dynamics and function. In Chapter 2, I first improved TSA-Seq with 10-20-fold higher sensitivity (TSA-Seq 2.0) to enable efficient mapping of genome organization relative to nuclear speckles. I applied TSA-Seq 2.0 to mapping nuclear speckles in four human cell lines and in one cell line upon heat shock. We found a largely conserved genome organization relative to speckles with small position changes tightly correlated with gene expression changes. In Chapter 3, I addressed the question of whether transcription is required for speckle association of certain chromatin domains. Speckle TSA-Seq mapping upon transcription inhibition revealed largely unchanged genome organization relative to speckles in either cycling or synchronized early-G1 phase cells, suggesting speckle association is independent of strong transcription. In Chapter 4, I optimized and extended TSA-Seq mapping to nuclear lamina in the same four human cell lines. Regions changing position relative to speckles show opposite changes relative to lamina. But there are many more changes relative to lamina including differential localization of heterochromatin regions between nuclear lamina and nucleolus. Our results reveal an initial 3D picture of nuclear compartmentalization. Further extension of TSA-Seq mapping into multiple nuclear compartments in different cell types and states will help further dissect nuclear organization and function.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Cell and Developmental Biology

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2020

Author dc:creator

• Zhang, Liguo

Contributors dc:contributor

• Belmont, Andrew
• Prasanth, Kannanganattu
• Brieher, William
• Stubbs, Lisa
• Zhao, Huimin

• Genome organization
• Nuclear position
• Nuclear compartment
• Nuclear body
• Nuclear speckles
• SON
• SC35
• Nuclear lamina
• Nucleolus
• TSA-Seq
• Microscopy
• Next generation sequencing
• Bioinformatics
• Gene expression
• Transcription
• Cell synchronization
• Histone modification
• Epigenetics
• Epigenomics
• Chromatin structure
• Chromatin decondensation
• Heat shock
• HSPH1
• Stem cell

Statement dc:rights

• © 2020 Liguo Zhang

Language dc:language

en

Handle dc:identifier

http://hdl.handle.net/2142/108234

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/108234