The long-range goal of the proposed studies is to determine how chromatin modulates gene expression during embryogenesis. The focus is on the role of two histone modifications that are associated with developmental control genes. Trimethylation of lysine 27 of histone H3 (H3K27me3) is commonly associated with silenced genes, whereas trimethylation of lysine 4 of histone H3 (H3K4me3) is commonly associated with genes that are active or permissive to be activated. Studies in pluripotent embryonic cells have revealed that both these marks are associated with developmental control genes before these genes are expressed. These bivalent H3K4me3/H3K27me3 marks have been proposed to contribute to the quantitative, temporal or spatial precision of developmental gene expression, but the field lacks a technology that would allow a direct test of these hypotheses. This project aims to (1) develop a method to induce specific histone modifications at defined genes and (2) apply this technology to determine the role of bivalent histone marks during zebrafish embryogenesis. The method involves the design of zinc finger fusion proteins that have histone methyltransferase or demethylase activities. These proteins will be targeted to defined genes to change H3K4 or H3K27 methylation. The technology will then be used to analyze the consequences of disrupting or introducing H3K4me3 or H3K27me3 marks at specific genes. Since specific histone modifications accompany developmental and physiological states in all eukaryotes, developing a technology for the gene-specific modulation of histone marks would help elucidate the role of such modifications in numerous settings. The proposed technology could also have direct impact in studying and modulating disease processes. For example, abnormal histone modifications have been implicated in several cancers. Local manipulation of histone modifications might help to determine which of the many misregulated genes are responsible for tumor formation. It might even become possible to use the proposed technology to specifically activate disease-protective genes or repress disease-inducing genes.
This project addresses how the myriad of different cell types form during animal development. Abnormal regulation of this process leads to birth defects and cancer, whereas methodical manipulation of this process can help understand and treat degenerative diseases.
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