Making Connections: Endoplasmic reticulum (ER)–mitochondria connections have been implicated in many physiological processes, including calcium homeostasis, signaling, membrane biogenesis, and apoptosis. Kornmann et al. (p. 477 , published online 25 June; see the Perspective by Wiedemann et al. ) looked for a proteinaceous link between the ER and mitochondria and, using combinations of synthetic biology and classical yeast genetics, found a protein complex that tethers the two organelles. A large-scale genetic interaction map suggests that these ER-mitochondria connections are important for interorganellar phospholipid exchange.

Embryonic stem (ES) cells are lines of cells that are isolated from blastocysts. The murine ES cells were demonstrated to be true pluripotent cells as they differentiate into all embryonic lineages. Yet, in vitro differentiation of rhesus ES cells was somewhat inconsistent and disorganized. The recent isolation of human ES cells calls for exploring their pluripotential nature. Human ES cells were grown in suspension to induce their differentiation into embryoid bodies (EBs). The differentiation status of the human ES cells and EBs was analyzed by following the expression pattern of several lineage-specific molecular markers using reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization. Here we report the induction in vitro of cystic embryoid bodies from human ES cells. Our findings demonstrate induction of expression of cell-specific genes during differentiation of the human ES cells into EBs. In the human EBs, we could show a characteristic regional expression of embryonic markers specific to different cellular lineages, namely, ζ-globin (mesoderm), neurofilament 68Kd (ectoderm), and α-fetoprotein (endoderm). Moreover, we present a synchronously pulsing embryoid body that expresses the myocardium marker α-cardiac actin. In addition, dissociating the embryoid bodies and plating the cells as monolayers results in multiple morphologies, among them cells with neuronal appearance that express neurofilament 68Kd chain. Human ES cells can reproducibly differentiate in vitro into EBs comprising the three embryonic germ layers. The ability to induce formation of human embryoid bodies that contain cells of neuronal, hematopoietic and cardiac origins will be useful in studying early human embryonic development as well as in transplantation medicine.

Human embryonic stem (ES) cells are pluripotent cells derived from the inner cell mass of in vitro fertilized human blastocysts. We examined the potential of eight growth factors [basic fibroblast growth factor (bFGF), transforming growth factor β1 (TGF-β1), activin-A, bone morphogenic protein 4 (BMP-4), hepatocyte growth factor (HGF), epidermal growth factor (EGF), β nerve growth factor (βNGF), and retinoic acid] to direct the differentiation of human ES-derived cells in vitro . We show that human ES cells that have initiated development as aggregates (embryoid bodies) express a receptor for each of these factors, and that their effects are evident by differentiation into cells with different epithelial or mesenchymal morphologies. Differentiation of the cells was assayed by expression of 24 cell-specific molecular markers that cover all embryonic germ layers and 11 different tissues. Each growth factor has a unique effect that may result from directed differentiation and/or cell selection, and we can divide the overall effects of the factors into three categories: growth factors (Activin-A and TGFβ1) that mainly induce mesodermal cells; factors (retinoic acid, EGF, BMP-4, and bFGF) that activate ectodermal and mesodermal markers; and factors (NGF and HGF) that allow differentiation into the three embryonic germ layers, including endoderm. None of the growth factors directs differentiation exclusively to one cell type. This analysis sets the stage for directing differentiation of human ES cells in culture and indicates that multiple human cell types may be enriched in vitro by specific factors.