Cell surface proteins are major targets of biomedical research due to their utility as cellular markers and their extracellular accessibility for pharmacological intervention. However, information about the cell surface protein repertoire (the surfaceome) of individual cells is only sparsely available. Here, we applied the Cell Surface Capture (CSC) technology to 41 human and 31 mouse cell types to generate a mass-spectrometry derived Cell Surface Protein Atlas (CSPA) providing cellular surfaceome snapshots at high resolution. The CSPA is presented in form of an easy-to-navigate interactive database, a downloadable data matrix and with tools for targeted surfaceome rediscovery (http://wlab.ethz.ch/cspa). The cellular surfaceome snapshots of different cell types, including cancer cells, resulted in a combined dataset of 1492 human and 1296 mouse cell surface glycoproteins, providing experimental evidence for their cell surface expression on different cell types, including 136 G-protein coupled receptors and 75 membrane receptor tyrosine-protein kinases. Integrated analysis of the CSPA reveals that the concerted biological function of individual cell types is mainly guided by quantitative rather than qualitative surfaceome differences. The CSPA will be useful for the evaluation of drug targets, for the improved classification of cell types and for a better understanding of the surfaceome and its concerted biological functions in complex signaling microenvironments.

Oct4 is a widely recognized pluripotency factor as it maintains Embryonic Stem (ES) cells in a pluripotent state, and, in vivo, prevents the inner cell mass (ICM) in murine embryos from differentiating into trophectoderm. However, its function in somatic tissue after this developmental stage is not well characterized. Using a tamoxifen-inducible Cre recombinase and floxed alleles of Oct4, we investigated the effect of depleting Oct4 in mouse embryos between the pre-streak and headfold stages, ∼E6.0–E8.0, when Oct4 is found in dynamic patterns throughout the embryonic compartment of the mouse egg cylinder. We found that depletion of Oct4 ∼E7.5 resulted in a severe phenotype, comprised of craniorachischisis, random heart tube orientation, failed turning, defective somitogenesis and posterior truncation. Unlike in ES cells, depletion of the pluripotency factors Sox2 and Oct4 after E7.0 does not phenocopy, suggesting that ∼E7.5 Oct4 is required within a network that is altered relative to the pluripotency network. Oct4 is not required in extraembryonic tissue for these processes, but is required to maintain cell viability in the embryo and normal proliferation within the primitive streak. Impaired expansion of the primitive streak occurs coincident with Oct4 depletion ∼E7.5 and precedes deficient convergent extension which contributes to several aspects of the phenotype.

Myocardial infarction resulting in irreversible loss of cardiomyocytes (CMs) remains a leading cause of heart failure. Although cell transplantation has modestly improved cardiac function, major challenges including increasing cell survival, engraftment, and functional integration with host tissue, remain. Embryonic stem cells (ESCs), which can be differentiated into cardiac progenitors (CPs) and CMs, represent a candidate cell source for cardiac cell therapy. However, it is not known what specific cell type or condition is the most appropriate for transplantation. This problem is exasperated by the lack of efficient and predictive strategies to screen the large numbers of parameters that may impact cell transplantation. We used a cardiac tissue model, engineered heart tissue (EHT), and quantitative molecular and electrophysiological analyses, to test transplantation conditions and specific cell populations for their potential to functionally integrate with the host tissue. In this study, we validated our analytical platform using contractile mouse neonatal CMs (nCMs) and noncontractile cardiac fibroblasts (cFBs), and screened for the integration potential of ESC-derived CMs and CPs (ESC-CMs and -CPs). Consistent with previous in vivo studies, cFB injection interfered with electrical signal propagation, whereas injected nCMs improved tissue function. Purified bioreactor-generated ESC-CMs exhibited a diminished capacity for electrophysiological integration; a result correlated with lower (compared with nCMs) connexin 43 expression. ESC-CPs, however, appeared able to appropriately mature and integrate into EHT, enhancing the amplitude of tissue contraction. Our results support the use of EHT as a model system to accelerate development of cardiac cell therapy strategies.

The composition of cell-surface proteins changes during lineage specification, altering cellular responses to their milieu. The changes that characterize maturation of early neural stem cells (NSCs) remain poorly understood. Here we use mass spectrometry-based cell surface capture technology to profile the cell surface of early NSCs and demonstrate functional requirements for several enriched molecules. Primitive NSCs arise from embryonic stem cells upon removal of Transforming growth factor-b signaling, while definitive NSCs arise from primitive NSCs upon Lif removal and FGF addition. In vivo aggregation assays revealed that N-cadherin upregulation is sufficient for the initial exclusion of definitive NSCs from pluripotent ectoderm, while c-kit signaling limits progeny of primitive NSCs. Furthermore, we implicate EphA4 in primitive NSC survival signaling and Erbb2 as being required for NSC proliferation. This work elucidates several key mediators of NSC function whose relevance is confirmed on forebrain-derived populations and identifies a host of other candidates that may regulate NSCs. STEM CELLS 2014;32:258-268

SummaryThe identification of cell surface proteins on stem cells or stem cell derivatives is a key strategy for the functional characterization, isolation, and understanding of stem cell population dynamics. Here, using an integrated mass spectrometry- and microarray-based approach, we analyzed the surface proteome and transcriptome of cardiac progenitor cells (CPCs) generated from the stage-specific differentiation of mouse and human pluripotent stem cells. Through bioinformatics analysis, we have identified and characterized FZD4 as a marker for lateral plate mesoderm. Additionally, we utilized FZD4, in conjunction with FLK1 and PDGFRA, to further purify CPCs and increase cardiomyocyte (CM) enrichment in both mouse and human systems. Moreover, we have shown that NORRIN presented to FZD4 further increases CM output via proliferation through the canonical WNT pathway. Taken together, these findings demonstrate a role for FZD4 in mammalian cardiac development.