Characterization of epithelial cell adhesion molecule as a surface marker on undifferentiated human embryonic stem cells.
Human embryonic stem cells (hESCs) have the capacity to remain pluripotent and self-renew indefinitely. To discover novel players in the maintenance of hESCs, we have previously reported the generation of monoclonal antibodies that bind to cell surface markers on hESCs, and not to mouse embryonic stem cells or differentiated embryoid bodies. In this study, we have identified the antigen target of one such monoclonal antibody as the epithelial cell adhesion molecule (EpCAM). In undifferentiated hESCs, EpCAM is localized to Octamer 4 (OCT4)-positive pluripotent cells, and its expression is down-regulated upon differentiation. To further understand its biological function in hESCs, endogenous EpCAM expression was silenced using small interfering RNA. EpCAM knockdown had marginal negative effects on OCT4 and TRA-1-60 expression, however cell proliferation was decreased by >40%. Examination of lineage marker expression showed marked upregulation of endoderm and mesoderm genes in EpCAM-silenced cells, under both pluripotent and differentiating conditions. These results were validated using a hESC line whose EpCAM expression has been stably knocked down. Data from the stable line confirmed that downregulation of EpCAM decreases cell growth and increases gene expression in the endoderm and mesoderm lineages. In vivo, hESCs lacking EpCAM were able to form teratomas containing tissues representing the three germ layers, and gene expression analysis yielded marked increase in the endoderm marker alpha fetoprotein compared with control. Together these data demonstrate that EpCAM is a surface marker on undifferentiated hESCs and plays functional roles in proliferation and differentiation.
Ng
Valerie Y
VY
Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Biopolis, Singapore 138668, Singapore. valerie.y.ng@gmail.com
Ang
Sheu Ngo
SN
Chan
Jia Xin
JX
Choo
Andre B H
AB
D000818:Animals; D000951:Antigens, Neoplasm / Q000235:genetics / Q000378:metabolism*; D000954:Antigens, Surface / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D019070:Cell Lineage; D049109:Cell Proliferation; D053595:Embryonic Stem Cells / Q000276:immunology* / Q000473:pathology; D000071858:Epithelial Cell Adhesion Molecule; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D050814:Octamer Transcription Factor-3 / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000276:immunology* / Q000473:pathology; D011509:Proteoglycans / Q000378:metabolism; D034622:RNA Interference; D013724:Teratoma / Q000276:immunology / Q000473:pathology
D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D015415:Biomarkers; D015815:Cell Adhesion Molecules; C499379:EPCAM protein, human; D000071858:Epithelial Cell Adhesion Molecule; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D011509:Proteoglycans; C107064:TRA-1-60 antigen, human