Investigating the role of histone modifications in stable differentiation and reprogramming in the Arabidopsis stomatal lineage
Abstract/Contents
- Abstract
- Plant somatic cells exhibit extraordinary developmental plasticity, yet they are able to stably maintain terminal identities. Although this capacity was recognized long ago, our mechanistic understanding of the establishment, maintenance, and erasure of cellular identities in plants remains limited. Manipulations of chromatin modifiers, particularly polycomb repressive complex 2 (PRC2), indicate chromatin-level regulation of cell identity is critical for establishing stable cell fates in plants. This is partially due to the fact that genes regulating embryo development and meristem identity are targets of PRC2-based repression in. Evidence also suggests developmental plasticity via reprogramming depends in part on plant differentiated cells' ability to re-establish stemness by accessing meristematic and embryonic gene expression programs. This happens naturally in some contexts; Kalanchoe daigremontiana clonally propagates by upregulating the expression of meristematic genes in leaf margins to reprogram differentiated cells to produce new seedlings. Reprogramming can also be induced experimentally; excision of the root stem cell niche in Arabidopsis causes differentiated root cells to revert to an embryo-like transcriptome to produce a new stem cell niche. Despite a growing body of literature describing reprogramming events and PRC2 in Arabidopsis thaliana, several questions remain. Namely, how do somatic cells supersede repressive chromatin states to reinstate developmental plasticity? If cells can so readily access these meristematic and embryonic transcriptional programs, how do plant cells specifically regulate when reprogramming can occur? In this dissertation, I use the Arabidopsis stomatal lineage to explore these phenomena in single cell types, thereby profiling cellular reprogramming with uniquely fine developmental and temporal resolution in plants. Stomata are structures on the surfaces of most land plants required for gas exchange between plants and their environment. Stomatal guard cells are the end product of a specialized lineage whose cell divisions and fate transitions ensure both production and pattern of cells in aerial epidermal tissues. These cell divisions and fate transitions are regulated by the sequential expression of a series of bHLH transcription factors that serve as master regulators of this developmental series: SPEECHLESS (SPCH), MUTE, and FAMA. The stomatal lineage is dynamic and flexible, altering stomatal production in response to environmental change. As such, the stomatal lineage is an excellent system to study how flexible developmental transitions are regulated in plants. In Chapter 1 I summarize the current knowledge of extrinsic and intrinsic regulation of stomatal development regulation. I also summarize current knowledge regarding chromatin-level regulation of cell fate stabilization in Arabidopsis and explain how the stomatal lineage can be a valuable model system for studying this phenomenon in plants. Guard cells can be manipulated such that they reprogram to the stomatal lineage initiation phase. In Chapter 2 I leverage a this to develop a cell-type specific reprogramming system than can be probed at the genome-wide scale for alterations in gene expression and histone modifications before and during reprogramming. I show that relationships among histone modification enrichments and gene expression in single cell types mirror trends from complex tissue, and that dynamic regulation by PRC2 is critical for maintenance of guard cell identity. Surprisingly, I found guard cells may sense and resist inappropriate reprogramming, in part through PRC2-mediated repression of a regulator of wound-induced callus formation. I propose a model where negative regulation of pro-reprogramming genes in the absence of a wounding signal is partially responsible cell fate maintenance in Arabidopsis. Furthering the utility of the reprogramming datasets I generated in chapter 2 requires cognate datasets of the stomatal lineage initiation phase. However, cells in this phase of stomatal development are not accessible to isolation with currently available tools. In Chapter 3, I describe the optimization of tools to render these cells accessible. To that end, I sought to optimize the Isolation of Nuclei Tagged in specific Cell Types (INTACT) system for the stomatal lineage to facilitate collection of rare stomatal lineage progenitor cells. In parallel, I applied the novel technique, Cleavage Under Targets and Release Using Nuclease (CUT& RUN) to Arabidopsis samples, enabling genome-wide chromatin mapping using small numbers of cells, which dramatically improves the signal to noise ratio of traditional protein-DNA interaction profiling. This chapter details the development of these tools and datasets, are promising for application to the stomatal lineage. In Chapter 4 I address further questions to be considered regarding cellular reprogramming in plants. I also comment on the outlook for INTACT and CUT& RUN Arabidopsis stomatal lineage and propose future applications for them.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2019; ©2019 |
| Publication date | 2019; 2019 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Lee, Laura Rose |
|---|---|
| Degree supervisor | Bergmann, Dominique |
| Thesis advisor | Bergmann, Dominique |
| Thesis advisor | Gozani, Or Pinchas |
| Thesis advisor | Morrison, Ashby J |
| Thesis advisor | Wang, Zhi-Yong, Dr |
| Degree committee member | Gozani, Or Pinchas |
| Degree committee member | Morrison, Ashby J |
| Degree committee member | Wang, Zhi-Yong, Dr |
| Associated with | Stanford University, Department of Biology. |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Laura Lee. |
|---|---|
| Note | Submitted to the Department of Biology. |
| Thesis | Thesis Ph.D. Stanford University 2019. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2019 by Laura Rose Lee
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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