个人简介

Ph.D., Stanford University B.S., Reed College

研究领域

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Biochemistry/Molecular Biology

Epigenetics, DNA methylation, genome dynamics, gene silencing, position effects. macrondidia Figure 1. The filamentous fungus Neurospora crassa. Image courtesy of M. Springer and N. Raju Overview We are interested in how the eukaryotic genome is structured, how it functions, and how it changes. Our current research concentrates on gene silencing in eukaryotes. We are particularly interested in mechanisms involving special states of chromatin (e.g. heterochromatin) and DNA methylation. Methylation alters properties of DNA, affects DNA-protein interactions, represses genes in animals, plants, and fungi and is essential for normal development in plants and mammals. Remarkably little is understood, however, about what determines which chromosomal regions are methylated. We are using genetic and biochemical approaches, primarily with the filamentous fungus Neurospora crassa (Fig. 1) as a model, to elucidate the mechanism, regulation and function of DNA methylation. In addition, we are using these approaches to explore silencing associated with chromosome ends, centromeres and other specialized regions of the genome. Inactivation of duplicated genes by RIP (hatch boxes) occurs prior to meiosis in Neuorspora. Figure 2. Inactivation of duplicated genes by RIP (shading) occurs prior to meiosis in Neurospora (see: Selker 1990 Ann. Rev. Gen. 24, 579-613). DNA Methylation DNA methylation is essential for normal development in a wide range of organisms including mammals and plants but is absent in some organisms including many popular model eukaryotes (e.g., yeasts, Drosophila, C. elegans). We showed that ~2% of cytosines in Neurospora DNA are methylated and that DNA methylation is not essential for development or viability in this organism. This set the stage for us to exploit this model eukaryote to elucidate the control and function of DNA methylation. We found that most methylated regions of Neurospora are relics of transposons inactivated by RIP (repeat-induced point mutation), a premeiotic homology-based genome defense system that litters duplicated sequences with C:G to T:A mutations (Figs. 2 & 3). Inactivation of duplicated genes by RIP (hatch boxes) occurs prior to meiosis in Neuorspora. Figure 3. Detailed analyses of DNA methylation in the Neurospora genome revealed that it is mostly in AT-rich centromeric regions, subtelomeric regions and dispersed relics of RIP, as shown here in red for one chromosome (see Lewis et al. 2008 Genome Research 19, 427-437). Our genetic and biochemical studies on the control of DNA methylation revealed clear ties between DNA methylation and chromatin modifications. The DIM-2 DNA methyltransferase is directed by heterochromatin protein 1 (HP1), which in turn recognizes trimethyl-lysine 9 on histone H3, placed by the DIM-5 histone H3 methyltransferase (Fig. 4). Inactivation of duplicated genes by RIP (hatch boxes) occurs prior to meiosis in Neuorspora. Figure 4. Localization of HP1-GFP depends on H3K9m3 by DIM-5. Note that the foci of heterochromatin in nuclei of wildtype are lost in the dim-5 mutant. (see Freitag M et. Al. 2004 Mol Cell 13, 427-34.) DNA methylation is modulated by a variety of additional factors. For example, in dmm-1 (DNA methylation modulator-1) mutants, methylation spreads from inactivated transposable elements, which can silence adjacent genes and lead to poor growth. Additional studies in the laboratory are providing other insights into the workings of DNA methylation and other silencing processes. Inactivation of duplicated genes by RIP (hatch boxes) occurs prior to meiosis in Neuorspora. Figure 5. Heterochromatin formation and DNA methylation. DIM-7 recruits the DIM-5 histone methyltransferase to A:T rich DNA (red), to form DCDC (Dim-Cul4-DDB1 Complex) resulting in methylation of K9 of histone H3. HP-1 then recognizes this histone mark and recruits the DIM-2 DNA methyltransferase (see Lewis et al. 2010. PLoS Genetics 6, e1001196).