Molecular analysis of LEFTY-expressing cells in early human embryoid bodies.
Human ESCs (HESCs) are self-renewing pluripotent cell lines that are derived from the inner cell mass of blastocyst-stage embryos. These cells can produce terminally differentiated cells representing the three embryonic germ layers. We thus hypothesized that during the course of in vitro differentiation of HESCs, progenitor-like cells are transiently formed. We demonstrated that LEFTY proteins, which are known to play a major role during mouse gastrulation, are transiently expressed during HESC differentiation. Moreover, LEFTY proteins seemed to be exclusively expressed by a certain population of cells in the early human embryoid bodies that does not overlap with the population expressing the ESC marker OCT4. We also showed that LEFTY expression is regulated at the cellular transcription level by molecular labeling of LEFTY-positive cells. A DNA microarray analysis of LEFTY-overexpressing cells revealed a signature of cell surface markers such as CADHERIN 2 and 11. Expression of LEFTY controlled by NODAL appears to have a substantial role in mesodermal origin cell population establishment, since inhibition of NODAL activity downregulated expression not only of LEFTY A and LEFTY B but also of BRACHYURY, an early mesodermal marker. In addition, other mesodermal lineage-related genes were downregulated, and this was accompanied by an upregulation in ectoderm-related genes. We propose that during the initial step of HESC differentiation, mesoderm progenitor-like cells appear via activation of the NODAL pathway. Our analysis suggests that in vitro differentiation of HESCs can model early events in human development.
Dvash
Tamar
T
Department of Genetics, Institute of Life Sciences, Hebrew University, Jerusalem, Israel.
Sharon
Nadav
N
Yanuka
Ofra
O
Benvenisty
Nissim
N
D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002469:Cell Separation; D002478:Cells, Cultured; D004149:Dioxoles / Q000494:pharmacology; D004622:Embryo, Mammalian / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D053595:Embryonic Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005326:Fetal Proteins / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D055458:Left-Right Determination Factors; D008648:Mesoderm / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D055457:Nodal Protein; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020825:T-Box Domain Proteins / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D000098908:Brachyury Protein
D001549:Benzamides; D015415:Biomarkers; D004149:Dioxoles; D005326:Fetal Proteins; D049452:Green Fluorescent Proteins; D055458:Left-Right Determination Factors; D055457:Nodal Protein; D012333:RNA, Messenger; D020825:T-Box Domain Proteins; D016212:Transforming Growth Factor beta; D000098908:Brachyury Protein; C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; C526161:LEFTY1 protein, human; C526007:NODAL protein, human; C526008:Nodal protein, mouse; C485184:enhanced green fluorescent protein