Dataset Information

SINGLE CELL ANALYSIS OF IN VITRO ERYTHROPOIESIS

ABSTRACT: We are developing a protocol to differentiate mouse and human induced pluripotent stem (IPS) and embryonic stem (ES) cells towards the haematopoietic pathway to generate erythrocytes in vitro. This system has many applications such as the study of the role of specific genes and human polymorphisms in infectious diseases such as malaria, as well as haematological diseases such as myelodysplastic syndrome. The nature of the in vitro differentiation process means that a heterogeneous population of cells is generated. In order to understand the types of cells produced with our protocol, we have performed a single cell analysis, which has the power to reveal the different populations of cells and their characteristics. For this, a cDNA library has been made that needs to be sequenced to obtain the gene expression profiles of the different cells. With this information we will be able to assess the quality of the differentiation protocol and improve it in order to produce better cells for the downstream applications. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/

PROVIDER: EGAS00001000764 | EGA |

REPOSITORIES: EGA

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Project description:The activation of receptors displayed on the surface of cells typically initiates cytoplasmic signalling cascades that lead to changes in cellular transcriptional responses so that a cell expresses proteins that enable it to behave appropriately as a function of their position within an organism. Understanding these signals and learning how to either prevent or ectopically activate them could have many therapeutic benefits and applications. In some cases these signals are highly localised, being delivered by directly neighbouring cells that express ligands which specifically bind to and activate receptors; for example, stem cells display receptors for ligands provided by cells within their niche to both ensure continual production of cells, and that their cellular progeny are directed towards appropriate fates. While for some cell types we know something of these signals enabling the in vitro differentiation of stem cells towards certain cellular fates, the differentiation processes is often not complete and so immature cell types which are not fully functional are produced, implying that some important signals are missing. Here, we propose to systematically identify receptor-triggered signalling pathways by stimulating receptors on the surface of human stem cells by using a library of soluble recombinant oligomerised protein ligands and measuring the consequent transcriptional perturbations using RNA-seq. We propose to screen an existing library of ~50 oligomerised human ligands previously used in receptor protein interaction screens and first test them for their ability to bind to human stem cells to identify those ligands for which the stem cells express a cell surface receptor. For those ligands which interact with the stem cells (we estimate between 2 and 5), they will be added to stem cell cultures and their effects on transcription quantified. The budget requested is sufficient for us to include controls and test important parameters such different time points after addition of the proteins. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/

Dataset Information

SINGLE CELL ANALYSIS OF IN VITRO ERYTHROPOIESIS

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