Project description:To identify the “time-lapse” TF networks during B lineage commitment, we established multipotent progenitors harboring a tamoxifen-inducible form of Id3, an in vitro system where virtually all cells became B cells within 6 days by simply withdrawing 4-OHT. In this study, single cell transcriptomic analysis at pre- and post-commitment was performed using the culture system. In addition, we also performed single cell RNA-seq analysis of B cell precursor populations (LMPP, CLP and pro-B cells) in murine bone marrow.
Project description:Single cell studies elucidating the transcriptional continuum underpinning lineage commitment in the human bone marrow (BM) have advanced the understanding of hematopoiesis beyond the classic hierarchical model. However, T-lineage commitment, which occurs in the thymus remains incompletely defined. We mapped single cell transcriptomes spanning post-natal human thymopoiesis including the earliest progenitors. Instead of previously described discrete populations, transcriptional and lineage assay data resolved a continuum of novel cell states within CD34+ thymic progenitor cells. The initial stages of thymopoiesis involve multilineage priming of single cells followed by a gradual transition to T-commitment, a continuum previously undescribed outside the BM. Early progenitors express a hematopoietic stem cell (HSC) like profile but unlike HSC show initiation of T-priming. Species related differences exist in the earliest progenitor cells.
Project description:Proper neural commitment is essential for ensuring the appropriate development of the human brain and for preventing neurodevelopmental diseases such as autism spectrum disorders, schizophrenia, and intellectual disorders. However, the molecular mechanisms underlying the neural commitment in humans remain elusive. Here, we report the establishment of a neural differentiation system based on human embryonic stem cells (hESCs) and on comprehensive RNA-Seq analysis of transcriptome dynamics during early hESC differentiation. Using weighted gene co-expression network analysis, we reveal that the hESC neurodevelopmental trajectory has five stages: pluripotency (day 0), differentiation initiation (days 2, 4, and 6), neural commitment (days 8–10), neural progenitor cell proliferation (days 12, 14, and 16), and neuronal differentiation (days 18, 20, and 22). These stages were characterized by unique module genes, which may recapitulate the early human cortical development. Moreover, a comparison of our RNA-Seq data with several other transcriptome profiling datasets from mice and humans indicated that module 3 associated with the day 8–10 stage is a critical window of fate switch from the pluripotency to the neural lineage. Interestingly, at this stage, no key extrinsic signals were activated. In contrast, using CRISPR-Cas9–mediated gene knockouts, we also found that intrinsic hub transcription factors, including the schizophrenia-associated SIX3 gene and septo-optic dysplasia–related HESX1 gene, are required to program hESC neural determination. Our results improve the understanding of the mechanism of neural commitment in the human brain and may help elucidate the etiology of human mental disorders and advance therapies for managing these conditions.