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Title: Transcriptional activator DOT1L putatively regulates human embryonic stem cell differentiation into the cardiac lineage
Authors: Pursani, V
Bhartiya, D
Tanavde, V
Bashir, M
Sampath, P 
Keywords: histone methyltransferase
histone methyltransferase DOT1L
homeobox protein Nkx-2.5
octamer transcription factor 4
small interfering RNA
transcription factor NANOG
unclassified drug
basic helix loop helix transcription factor
calcium binding protein
DOT1L protein, human
helix-loop-helix protein, eHAND
membrane protein
muscle protein
MYOF protein, human
NFE2L2 protein, human
NKX2-5 protein, human
transcription factor Nrf2
cardiac cell line
cardiac stem cell
cell activity
cell differentiation
chromatin immunoprecipitation
controlled study
enzyme localization
gene activation
gene expression regulation
gene knockdown
gene sequence
gene targeting
HAND1 gene
heart development
HES3 cell
human cell
human embryonic stem cell
in vitro study
KIND1 cell
marker gene
MYOF gene
NANOG gene
NKX25 gene
NR2F2 gene
pluripotent stem cell
POU5F1 gene
priority journal
transcription initiation site
transcription regulation
cardiac muscle cell
cell culture
cell line
cell lineage
human embryonic stem cell
Basic Helix-Loop-Helix Transcription Factors
Calcium-Binding Proteins
Cell Differentiation
Cell Line
Cell Lineage
Cells, Cultured
Homeobox Protein Nkx-2.5
Human Embryonic Stem Cells
Membrane Proteins
Muscle Proteins
Myocytes, Cardiac
NF-E2-Related Factor 2
Issue Date: 2018
Citation: Pursani, V, Bhartiya, D, Tanavde, V, Bashir, M, Sampath, P (2018). Transcriptional activator DOT1L putatively regulates human embryonic stem cell differentiation into the cardiac lineage. Stem Cell Research and Therapy 9 (1) : 97. ScholarBank@NUS Repository.
Rights: Attribution 4.0 International
Abstract: Background: Commitment 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. Methods: The 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. Results: ChIP 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. Conclusions: Collectively, our data suggests the crucial role of H3K79me2 methyltransferase DOT1L for activation of NKX2.5 during the cardiac differentiation of hES cells. © 2018 The Author(s).
Source Title: Stem Cell Research and Therapy
ISSN: 17576512
DOI: 10.1186/s13287-018-0810-8
Rights: Attribution 4.0 International
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