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Status |
Public on Jun 30, 2023 |
Title |
Genome-wide transcriptomic profiling of cardiomyocyte differentiation from Wild-type and PERK-KO H1 hESCs [RNA-seq] |
Organism |
Homo sapiens |
Experiment type |
Expression profiling by high throughput sequencing
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Summary |
Cardiac development involves large-scale rearrangements of the proteome. How the developing cardiac cells maintain the integrity of the proteome during the rapid lineage transition remains unclear. Here we show that proteotoxic stress visualized by the misfolded protein aggregates appears during early cardiac differentiation of human embryonic stem cells (hESCs) and is resolved by activation of the PERK branch of the unfolded protein response (UPR). PERK depletion increases misfolded protein accumulation, leading to pluripotency exit defect and impaired mesendoderm specification of hESCs. Mechanistically, we found that PERK safeguards cardiogenesis through its conserved downstream effector ATF4, which subsequently activates a novel transcriptional target WARS1, to cope with the differentiation-induced proteotoxic stress. Our results indicate that protein quality control represents a previously unrecognized core component of the cardiogenic regulatory network. Broadly, these findings provide a framework for understanding how UPR is integrated into the developmental program by activating the PERK-ATF4-WARS1 axis.
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Overall design |
Undifferentiated hESCs cultured in E8 medium were dissociated into single cell suspension by Accutase (Stem Cells Technology, 7920) and reseeded onto Matrigel-coated 24-well plate at a density of 105 cells/per well in E8 medium containing 10 μM Y-27632. When reached ~80% confluence 2-3 days after plating, CM differentiation was initiated by switching to the cardiac differentiation medium (DMEM/F-12 (Gibco, 11330032) supplemented with 50 U ml-1 Penicillin-Streptomycin (Gibco, 15070063), Chemically Defined Lipid Concentrate (1:100, Gibco, 11905031), 10.7 μg ml-1 holo-Transferrin human (Sigma, T0665), 71 μg ml-1 L-Ascorbic acid (Sigma, A8960), 14 ng ml-1 Sodium selenite (Sigma, S5261)) and renewed every day. 5 μM CHIR99021 (Selleck, S1263) and 3 μM IWP2 (Selleck, S7085) was added into the cardiac differentiation medium from days 0-1 and days 2-5, respectively. 3 μg ml-1 Heparin was added into the cardiac differentiation medium from days 1-7. 20 μg ml-1 Insulin (Sigma, 91077C) was added into the cardiac differentiation medium from day 7 onward. The medium was changed daily until day 7 and renewed every 2-3 days from day 7 onward. At day 0, 2 and 6 during the differentiation period, and cells were collected using using the NucleoZol reagent. mRNA libraries were prepared using the VAHTS® mRNA-seq V2 Library Prep Kit for Illumina from Vazyme and sequenced on an Illumina HiSeq 2000 with 150 bp paired-end reads by GENEWIZ.
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Contributor(s) |
Cheng W, Cao N |
Citation missing |
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Submission date |
Nov 04, 2021 |
Last update date |
Jun 30, 2023 |
Contact name |
Weisheng Cheng |
E-mail(s) |
chengwsh@outlook.com
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Organization name |
Sun yat-sen University
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Street address |
2nd Zhongshan Road
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City |
Guangzhou |
ZIP/Postal code |
510080 |
Country |
China |
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Platforms (1) |
GPL11154 |
Illumina HiSeq 2000 (Homo sapiens) |
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Samples (36)
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This SubSeries is part of SuperSeries: |
GSE188123 |
RNA-seq and ChIP-seq studies of cardiomyocyte differentiation from H1 hESCs |
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Relations |
BioProject |
PRJNA777808 |
SRA |
SRP344521 |
Supplementary file |
Size |
Download |
File type/resource |
GSE188111_RAW.tar |
85.9 Mb |
(http)(custom) |
TAR (of TXT) |
GSE188111_Raw_gene_counts_matrix.txt.gz |
2.4 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
Processed data are available on Series record |
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