GSBS Dissertations and Theses

ORCID ID

0000-0003-0253-0809

Approval Date

8-8-2017

Document Type

Doctoral Dissertation

Academic Program

Interdisciplinary Graduate Program

Department

Department of Molecular, Cell and Cancer Biology

First Thesis Advisor

Thomas Fazzio

Keywords

Embryonic stem cells, chromatin, gene regulation, MBD3/NuRD, H3K56ac

Abstract

Central to the control of virtually all cellular activity is the regulation of gene expression. In eukaryotes, this regulation is greatly influenced by chromatin structure, which is itself regulated by numerous chromatin-remodeling complexes. These are typically large protein complexes with interchangeable subunits that allow for highly specialized functions in different cell types. Moreover, additional specificity can be gained through complexes formed from different subunit isoforms. Histone modifications also regulate chromatin by recruiting remodeling complexes to particular genomic regions.

In this thesis we characterize MBD3C, an isoform of the Nucleosome Remodeling and Deacetylase (NuRD) complex subunit MBD3. MBD3 is essential for pluripotency and development, but MBD3C appears to be expressed only in embryonic stem cells (ESCs), and whether it forms a distinct NuRD complex, how its expression is regulated, and its precise function(s) remain unknown. We show that MBD3C forms a complete NuRD complex that functions redundantly with the other MBD3 isoforms in ESC gene regulation. Furthermore, MBD3C binds the SET/MLL complex subunit WDR5 through a conserved motif within its unique N-terminal region, and this interaction is necessary for the regulation of >2,000 ESC genes. Together, these findings indicate that ESCs can utilize isoforms of the same protein to achieve similar functions through diverse mechanisms.

The second part of this thesis focuses on the role of the histone modification H3.3K56ac in pluripotency and differentiation. Although H3K56ac is well-studied in yeast, in mammalian cells it is far less abundant and its functions are largely unknown. Our data indicate that the H3.3K56R mutant is largely normal for ESC maintenance and loss of pluripotency markers during differentiation, but H3.3K56ac is necessary for proper lineage commitment. Ongoing studies will characterize the H3.3K56Q phospho-mimetic mutant during differentiation, and examine H3.3K56ac function at lineage-specific genes.

DOI

10.13028/M2DQ1G

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Licensed under a Creative Commons license

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This work is licensed under a Creative Commons Attribution-Noncommercial 4.0 License

Available for download on Thursday, September 27, 2018

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