Discovery of novel determinants of endothelial lineage using chimeric heterokaryons.
Wing Tak WongGianfranco MatroneXiaoYu TianSimion Alin TomoiagaKin Fai AuShu MengSayumi YamazoeDaniel SievekingKaifu ChenDavid M BurnsJames K ChenHelen M BlauJohn P CookePublished in: eLife (2017)
We wish to identify determinants of endothelial lineage. Murine embryonic stem cells (mESC) were fused with human endothelial cells in stable, non-dividing, heterokaryons. Using RNA-seq, it is possible to discriminate between human and mouse transcripts in these chimeric heterokaryons. We observed a temporal pattern of gene expression in the ESCs of the heterokaryons that recapitulated ontogeny, with early mesodermal factors being expressed before mature endothelial genes. A set of transcriptional factors not known to be involved in endothelial development was upregulated, one of which was POU class 3 homeobox 2 (Pou3f2). We confirmed its importance in differentiation to endothelial lineage via loss- and gain-of-function (LOF and GOF). Its role in vascular development was validated in zebrafish embryos using morpholino oligonucleotides. These studies provide a systematic and mechanistic approach for identifying key regulators in directed differentiation of pluripotent stem cells to somatic cell lineages.
Keyphrases
- endothelial cells
- single cell
- rna seq
- gene expression
- pluripotent stem cells
- high glucose
- cell therapy
- vascular endothelial growth factor
- embryonic stem cells
- high throughput
- dna methylation
- stem cells
- transcription factor
- small molecule
- genome wide
- bone marrow
- induced pluripotent stem cells
- case control
- genome wide identification
- nucleic acid