Spatiotemporal patterning of EpCAM is important for murine embryonic endo- and mesodermal differentiation

Abstract

Epithelial cell adhesion molecule EpCAM is expressed in pluripotent embryonic stem cells (ESC) in vitro, but is repressed in differentiated cells, except epithelia and carcinomas. Molecular functions ofEpCAM, possibly imposing such repression, were primarily studied in malignant cells and might not apply to non-pathologic differentiation. Here, we comprehensively describe timing and rationale for EpCAM regulation in early murine gastrulation and ESC differentiation using single cell RNA-sequencing datasets, in vivo and in vitro models including CRISPR-Cas9-engineered ESC-mutants. We demonstrate expression of EpCAM in inner cell mass, epiblast, primitive/visceral endoderm, and strict repression in the most primitive, nascent Flk1 + mesoderm progenitors at E7.0. Selective expression of EpCAM was confirmed at mid-gestation and perinatal stages. The rationale for strict patterning was studied in ESC differentiation. Gain/loss-of-function demonstrated supportive functions of EpCAM in achieving full pluripotency and guided endodermal differentiation, but repressive functions in mesodermal differentiation as exemplified with cardiomyocyte formation. We further identified embryonic Ras (ERas) as novel EpCAM interactor of EpCAM and an EpCAM/ERas/AKT axis that is instrumental in differentiation regulation. Hence, spatiotemporal patterning of EpCAM at the onset of gastrulation, resulting in early segregation of interdependent EpCAM + endodermal and EpCAM − /vimentin + mesodermal clusters represents a novel regulatory feature during ESC differentiation.Epithelial cell adhesion molecule EpCAM was originally described as a cell surface antigen highly expressed in human carcinomas 1 . Today, we know that EpCAM is present as a heart-shaped cis-dimer at the cell surface 2 , and that it has a broader but nevertheless sharply restricted expression pattern in undifferentiated pluripotent

High Expression of EpCAM and Sox2 is a Positive Prognosticator of Clinical Outcome for Head and Neck Carcinoma

“…Induction of EpCAM results in various cellular outcome including cell-cell adhesion 37 , enhanced tumor cell proliferation 19 , 38 and regulation of stem cell features 20 , 39 – 41 . Itself, EpCAM is prone to substantial regulation throughout cancer progression 27 , 42 , 43 , stem cell differentiation 22 , and during EMT related to treatment resistance, e . g .…”

“…In the present study, we analyzed the expression levels and patterns of EpCAM, Sox2 and vimentin in primary HNSCC for association with patient outcome. EpCAM is an epithelial cell marker involved in regulation of proliferation and stem cell differentiation 19 – 22 . EpCAM down-regulation in mesodermally differentiating embryonic stem cells that undergo EMT is an early and necessary event in gastrulation 22 , which is likewise observed in partial EMT occurring in cancer cells 23 – 28 .…”

“…EpCAM is an epithelial cell marker involved in regulation of proliferation and stem cell differentiation 19 – 22 . EpCAM down-regulation in mesodermally differentiating embryonic stem cells that undergo EMT is an early and necessary event in gastrulation 22 , which is likewise observed in partial EMT occurring in cancer cells 23 – 28 . Sex-determining region Y-box 2 (Sox2) is a transcription factor required for stem cell pluripotency and reprograming of somatic cells to induced pluripotent stem cells 29 .…”

Revisiting the Roles of Pro-Metastatic EpCAM in Cancer

“…There were also studies in mammalian cell lines utilizing genome editing tools to study EpCAM loss-of-function. For instance, one study generated single-cell clones of CRISPR/Cas9 mediated EpCAM knockout using E14TG2α embryonic stem cells markers [61]. The clones showed reduced expression of pluripotency genes such as Oct3/4, Sox2 and Nanog and displayed significant >50% reductions of endodermal markers.…”

A high-content screen for small-molecule regulators of epithelial cell-adhesion molecule (EpCAM) cleavage yields a robust inhibitor

“…This expression is reactivated or enforced in the vast majority of carcinoma (25) and in cancer stem cells (26). The maintenance of the undifferentiated state of embryonic stem cells is strongly connected with EpCAM expression levels (6,16,20,27). In carcinomas, EpCAM is highly overexpressed and (re-)distributed over the whole cell surface, which is frequently associated with cytoplasmic and nuclear staining (6, 28 -31).…”