Dr. Lorincz’s laboratory is focused on the interplay between transcription, DNA methylation and histone modification in development, using the mouse as a model system.
A number of factors have been described that catalyze the post-translational addition or removal of specific moieties, such as acetyl or methyl groups, to/from specific residues on the core nucleosomal histones, features of the so-called “histone code”. Methylation of lysine 4 of the H3 tail, which is associated with the promoter regions of both actively transcribed and “poised” genes, was recently shown to inhibit de novo DNA methylation, thus serving to “protect” promoter regions from DNA methylation. Conversely, we have shown that trimethylation of H3K9 (H3K9me3), a mark associated with transcriptional repression that is inversely correlated with H3K4me3, is enriched in embryonic stem cells at the promoter regions of a subset of germline-specific genes and specific endogenous retroviruses ERVs. This mark is deposited by the H3K9 KMTase Setdb1, generally independent of DNA methylation and serves to maintain a number of ERVs in a silent state in cells deficient in all three DNA methyltransferases. Surprisingly, HP1 proteins, “readers” of the H3K9me3 mark, are dispensable for H3K9me3-mediated proviral silencing, raising the question, how does H3K9 methylation inhibit transcription?
Ongoing research in the lab is directed towards characterizing the interplay between readers and writers of covalent histone marks, chromatin remodeling factors and DNA methylation in transcriptional regulation of genes and retroelements (see Sharif et al. Cell Stem Cell, 2016), using knock-down, conventional and CRISPR-based genetic knock-out approaches. Employing Illumina next generation sequencing and bioinformatics pipelines developed in house (see Younesy et al, Bioinformatics, 2014), we systematically characterizing the role of histone H3K9 methyltransferases, H3K9me “readers” and chromatin remodeling factors in ESCs via RNAseq, ChIPseq, meDIPseq and hmeDIPseq analyses (see Liu et al., Genes & Development, 2014).
In complementary genome-wide studies, we are also characterizing the function of LTR elements as alternative genic promoters in ESCs and early embryos as well as germ cells (see Molecular Cell, Thompson et al., 2016) and studying the impact of specific elements on the transcriptome in distantly related mouse strains as well as in rats.
We also recently developed an ultra-low-input micrococcal nuclease-based native ChIP (ULI-NChIP) and sequencing method (See Brind’Amour et al, Nature Communications, 2015) to generate genome-wide histone mark profiles with high resolution from as few as 103 cells. We demonstrate that ULI-NChIP-seq generates high-quality maps of covalent histone marks from 103 to 106 embryonic stem cells. Subsequently, we showed that ULI-NChIP-seq H3K27me3 profiles generated from E13.5 primordial germ cells isolated from single male and female embryos show high similarity to recent data sets generated using 50–180x more material. We are currently applying this method to address fundamental questions about intergenerational inheritance of covalent histone marks and their role in enhancer function and the inheritance of DNA methylation in the early mouse embryo.
