Transcriptional activator DOT1L putatively regulates human embryonic stem cell differentiation into the cardiac lineage

Abstract

BackgroundCommitment of pluripotent stem cells into differentiated cells and associated gene expression necessitate specific epigenetic mechanisms that modify the DNA and corresponding histone proteins to render the chromatin in an open or closed state. This in turn dictates the associated genetic machinery, including transcription factors, acknowledging the cellular signals provided. Activating histone methyltransferases represent crucial enzymes in the epigenetic machinery that cause transcription initiation by delivering the methyl mark on histone proteins. A number of studies have evidenced the vital role of one such histone modifier, DOT1L, in transcriptional regulation. Involvement of DOT1L in differentiating pluripotent human embryonic stem (hES) cells into the cardiac lineage has not yet been investigated.MethodsThe study was conducted on in-house derived (KIND1) and commercially available (HES3) human embryonic stem cell lines. Chromatin immunoprecipitation (ChIP) was performed followed by sequencing to uncover the cardiac genes harboring the DOT1L specific mark H3K79me2. Following this, dual immunofluorescence was employed to show the DOT1L co-occupancy along with the cardiac progenitor specific marker. DOT1L was knocked down by siRNA to further confirm its role during cardiac differentiation.ResultsChIP sequencing revealed a significant number of peaks characterizing H3K79me2 occupancy in the proximity of the transcription start site. This included genes like MYOF, NR2F2, NKX2.5, and HAND1 in cardiac progenitors and cardiomyocytes, and POU5F1 and NANOG in pluripotent hES cells. Consistent with this observation, we also show that DOT1L co-localizes with the master cardiac transcription factor NKX2.5, suggesting its direct involvement during gene activation. Knockdown of DOT1L did not alter the pluripotency of hES cells, but it led to the disruption of cardiac differentiation observed morphologically as well as at transcript and protein levels.ConclusionsCollectively, our data suggests the crucial role of H3K79me2 methyltransferase DOT1L for activation of NKX2.5 during the cardiac differentiation of hES cells.Electronic supplementary materialThe online version of this article (10.1186/s13287-018-0810-8) contains supplementary material, which is available to authorized users.

The methyltransferase DOT1L limits activation and the Th1 program in CD4+T cells during infection and inflammation

“…Our finding that DOT1L-dependent H3K79me2 is associated with a subset of highly expressed genes is consistent with several previous studies that correlate the presence of H3K79me2 with gene expression and transcriptional elongation (Chen et al, 2018;Guenther et al, 2007;Im et al, 2003;Kryczek et al, 2014;Pursani et al, 2018;Steger et al, 2008;Wang et al, 2008). Despite our own findings and previous reports that correlate H3K79me2 with gene expression (Bernt et al, 2011;Steger et al, 2008), H3K79me2 is not necessarily required for gene expression.…”

The Methyltransferase DOT1L Controls Activation and Lineage Integrity in CD4+ T Cells during Infection and Inflammation

“…Our finding that DOT1L-dependent H3K79me2 is associated with a subset of highly expressed genes is consistent with several previous studies that correlate the presence of H3K79me2 with gene expression and transcriptional elongation (Chen et al, 2020;Guenther et al, 2007;Im et al, 2003;Kryczek et al, 2014;Pursani et al, 2018;Steger et al, 2008;Wang et al, 2008). Despite our own findings and previous reports that correlate DOT1L-dependent H3K79me2 with gene expression (Bernt et al, 2011;Steger et al, 2008), H3K79me2 is not necessarily required for gene expression.…”

DOT1L-mediated murine neuronal differentiation associates with H3K79me2 accumulation and preserves SOX2-enhancer accessibility

“…Various epigenetic enzymes are important for the orchestration of neuronal differentiation 2,8 . Among these, Disruptor of Telomeric silencing 1 Like (DOT1L) has been recently identified as a critical player in the differentiation process [9][10][11][12] . DOT1L is a highly conserved histone methyltransferase that catalyzes the mono-, di-and trimethylation of lysine 79 of histone H3 (H3K79me1/me2/me3) 13 .…”