Squeezing out the histone modifications data Wieslawa Mentzen with Matteo Floris and Paolo Uva Connections between epigenetics and microRNAs during embryonic stem cell differentiation, an update:

The regulatory program underlying differentiation involves a distinct network of transcription factors and microRNAs. Expression of many differentiation-associated genes is regulated by the epigenetic modifications. As much as one third of a genome differs in chromatin structure between pluripotent and lineage-committed cell lines. We focus on the degree to which miRNAs are epigenetically regulated.

Cell lines Undifferentiated: h1ESC, human embryonic stem cells Differentiated: IMR90, fetal lung fibroblasts miRNA precursors and their putative promoters Marson et al, Connecting microRNA genes to the core transcriptional regulatory circuitry of embryonic stem cells, Cell, 2008 Modifications -DNA methylation data Lister et al., Human DNA methylomes at base resolution show widespread epigenomic differences, Nature Histone modifications: H3K9me3 and H3K27me3 Hawkins et al., Distinct epigenomic landscapes of pluripotent and lineage-comitted human cells, Cell Stem Cell 2010 Chromosome coordinates of chromatin domains with increased level of each modification. For each miRNA we look for overlaps with DNA hypermethylation or histone modification domain in the gene body and promoter regions.

H3K9me3 H3K27me3 ESC IMR gene promoter Presence of modifications 320 out of 550 miRNA genes have a K9me3 or K27me3 mark in either cell line 74

01 Correlation between (#mods in IMR - #mods ESC) and expression ratio ESC/IMR , p<2.2e-16 Expression ratio ESC/IMR Presence of modification ESCIMRESC/IMR

K9 K27 in ESC in IMR expression ratio ESC/IMR

K9 K27 in ESC in IMR expression ratio ESC/IMR K9 in IMR, high expression ratio K9 and K27 in ESC, low expression K27 in IMR, high expression ratio K27 in ESC, low expression K9 in ESC, low expression

appearance loss shrinkage presence in both cell lines expansion ESC IMR

gene promoter H3K27me3 H3K9me3 IMR both ESC expands in IMR shrinks in IMR Change of modification during differentiation

ESC IMR

ESC IMR

H3K9 H3K27

miRNA known from literature to be involved in cell differentiation Are they under epigenetic regulation?

miRNA expressed in ESC miR-375 Expression higher in ESC Expansion of K27 domain in IMR

miR9-1 miRNA expressed in ESC Expression higher in ESC Expansion of K27 domain in IMR, appearance of K9 domain in IMR

miR-363 cluster miRNA expressed in ESC Expression higher in ESC Appearance of K9 and K27 domains in IMR Regulation specific for this cluster!

miR homologs in mouse constitute 70% of miRNA population expressed in ESC (Marson et al., 2008). miRNA expressed in ESC specific for this and neighboring miR-371 cluster Large bipartite cluster on chr 19 Expression higher in ESC Expansion of K9 domain in IMR

miR-135b miRNA expressed in ESC Expression higher in ESC Expansion of K27 domain in IMR

miR miRNA expressed in ESC Expression higher in ESC Appearance of K9 domain in IMR

miR-17 cluster miRNA expressed in ESC Expression higher in ESC Appearance of K27 domain in IMR Specific for promoter region of the cluster

Chr 14 clusters miR-370 and miR-770 miRNA expressed in IMR

miR-370 cluster Expression higher in IMR Presence of K9 or K27 domain in ESC

miRNA expressed in IMR Expression higher in IMR Presence of K27 domain in ESC in miRNAs within RTL1 gene K27 domain in ESC in some of those genes that are within RTL1 gene, specific RTL1: This gene is a retrotransposon-derived, paternally expressed imprinted gene that is highly expressed at the late fetal stage in both the fetus and placenta. It has an overlapping maternally expressed antisense transcript, which contains several microRNAs targeting the transcripts of this gene through an RNA interference (RNAi) mechanism. This gene is essential for maintenance of the fetal capillaries. miR-770 cluster

miR-152 miRNA expressed in IMR Expression higher in IMR K9 domain in ESC

miRNA known from literature to be involved in cell differentiation

Summary -320 out of 550 miRNA genes have a K9me3 or K27me3 mark in either cell line -most of the analyzed miRNAs have histone modification pattern consistent with these genes being epigenetically regulated -presence of a repressive modification in gene or promoter region of miRNA is associated with a reduced expression level in this cell line -both the appearance and the expansion of K27 and K9 domains have a repressive effect - this regulation in many cases is miRNA-specific and not the spill-over from the regulation of nearby genes

MERRY CHRISTMAS AND HAPPY NEW YEAR THE END