Chromatin Dynamics and the Specification of Cell Fate

Roger Deal, Fred Hutchinson Cancer Research Center

November 3, 2010 @ 05:00 pm to 06:00 pm

101 Althouse

How are chromatin-based mechanisms used to establish and maintain specific gene expression programs that give rise to specialized cell types? The answer to this question is critical for understanding how multicellular organisms build their bodies and also for learning to treat diseases, such as cancer, that can arise from defects in cell fate specification. However, reaching an understanding of this problem has been hindered by technical limitations such as the difficulty of analyzing the dynamic behavior of chromatin in real time, and the lack of simple methods for epigenomic analysis of specific cell types within a tissue. I have developed and applied two new technologies that overcome these hurdles and have allowed previously intractable questions to be addressed. First, as a means of studying the relationship between chromatin dynamics and genome activity, I used metabolic labeling with an amino acid analog to affinity tag newly synthesized histones, allowing measurement of the locations and rates of nucleosome eviction and replacement, or turnover, across the genome. Application of this method to cultured Drosophila cells revealed that nucleosome turnover occurs rapidly over transcribed genes, regulatory elements, and at DNA replication origins. The turnover of nucleosomes was found to be much faster at sites of trithorax group (TrxG) activator binding than at sites of polycomb group (PcG) silencer binding, suggesting that the opposing activities of these epigenetic regulators are mediated through their differential effects on nucleosome turnover, and the resulting differences in DNA exposure. Second, to address how chromatin-based regulation is used to effect cell differentiation, I used affinity tagging of the nuclear envelope in specific cell types within a tissue, allowing purification of the labeled nuclei for profiling of gene expression and chromatin features in the cell type of interest. I employed this method to study the hair and non-hair cell types of the Arabidopsis root epidermis, which arise from a common progenitor. I identified hundreds of genes that are preferentially expressed in each cell type and found that cell type-specific expression is often associated with differences between cell types in the TrxG-mediated trimethylation of histone H3 lysine 4 and PcG-mediated trimethylation of histone H3 lysine 27 at these genes. Collectively, these findings suggest that TrxG and PcG proteins stabilize gene expression states by modulating the rate of nucleosome turnover, and that regulation of their histone methylation activities is a critical element of plant cell differentiation. Importantly, this work also provides generally applicable new tools to address fundamental problems in epigenomics. I will also discuss my future plans to study the mechanics of genome reprogramming during cell differentiation, the coordination of multiple differentiation events during organ formation, and the relationships between regulated nucleosome turnover and the control of processes such as transcription and replication that use DNA as a template. Hosted by Biochemistry and Molecular Biology

Contact

Tamara Housel
txh9@psu.edu
814-865-3072