Project description:We performed microarray analyses on human ESCs-derived NPs and neurons carrying loss-of-function mutation in the MeCP2 gene. Neural precursors and differentiating neurons at 2 and 4-week were used for RNA extraction and affymetrix microarrays. We added ERCC RNA spike-in controls to normalize to cell number input.
Project description:To define the sequence of events that lead to the generation of somatic motor neurons (MNs), we took advantage of ESCs, which can be directed to differentiate into spinal neural progenitors (NPs) in vitro (Gouti et al. 2014). This method relies on the exposure of ESCs, cultured as a monolayer, to a brief pulse of Wnt signalling prior to neural induction. Subsequently, removal of Wnt signalling and exposure to retinoic acid (RA) results in the generation of NPs. Exposure of these NPs to Shh signalling agonist SAG induces the expression of genes expressed in the ventral spinal cord and the induction of MNs.
Project description:Expression data comparing transformed or neoplastic human neural precursors following OTX2 knockdown to transformed human neural precursor cells
Project description:Human telencephalon is an evolutionary advanced brain structure associated with many uniquely human behaviors and disorders. However, cell lineages and molecular pathways implicated in human telencephalic development remains largely unknown. We generated human telencephalic organoids from stem cell-derived single neural rosettes (SNRs) and investigated telencephalic development under normal and pathological conditions. SNR-derived organoids contained pallial and subpallial neural progenitors (NPs), excitatory and inhibitory neurons, as well as macroglial and periendothelial cells, and demonstrated predictable organization and cytoarchitecture. We comprehensively characterized the properties of neurons in SNR-derived organoids and identified transcriptional programs associated with the specification of excitatory and inhibitory lineages from a common pool of NPs early in telencephalic development. We also demonstrated that neurons in organoids with a hemizygous deletion of an autism- and intellectual disability associated gene SHANK3 exhibit intrinsic and excitatory synaptic deficits associated with impaired expression of clustered protocadherins. Collectively, this study validates SNR-derived organoids as a reliable new model for studying human telencephalic development and identifies novel molecular pathways disrupted by SHANK3 hemizygosity in human telencephalic tissue.
Project description:Human telencephalon is an evolutionary advanced brain structure associated with many uniquely human behaviors and disorders. However, cell lineages and molecular pathways implicated in human telencephalic development remains largely unknown. We generated human telencephalic organoids from stem cell-derived single neural rosettes (SNRs) and investigated telencephalic development under normal and pathological conditions. SNR-derived organoids contained pallial and subpallial neural progenitors (NPs), excitatory and inhibitory neurons, as well as macroglial and periendothelial cells, and demonstrated predictable organization and cytoarchitecture. We comprehensively characterized the properties of neurons in SNR-derived organoids and identified transcriptional programs associated with the specification of+B50:B51 excitatory and inhibitory lineages from a common pool of NPs early in telencephalic development. We also demonstrated that neurons in organoids with a hemizygous deletion of an autism- and intellectual disability associated gene SHANK3 exhibit intrinsic and excitatory synaptic deficits associated with impaired expression of clustered protocadherins. Collectively, this study validates SNR-derived organoids as a reliable new model for studying human telencephalic development and identifies novel molecular pathways disrupted by SHANK3 hemizygosity in human telencephalic tissue.
Project description:Human telencephalon is an evolutionary advanced brain structure associated with many uniquely human behaviors and disorders. However, cell lineages and molecular pathways implicated in human telencephalic development remains largely unknown. We generated human telencephalic organoids from stem cell-derived single neural rosettes (SNRs) and investigated telencephalic development under normal and pathological conditions. SNR-derived organoids contained pallial and subpallial neural progenitors (NPs), excitatory and inhibitory neurons, as well as macroglial and periendothelial cells, and demonstrated predictable organization and cytoarchitecture. We comprehensively characterized the properties of neurons in SNR-derived organoids and identified transcriptional programs associated with the specification of+B50:B51 excitatory and inhibitory lineages from a common pool of NPs early in telencephalic development. We also demonstrated that neurons in organoids with a hemizygous deletion of an autism- and intellectual disability associated gene SHANK3 exhibit intrinsic and excitatory synaptic deficits associated with impaired expression of clustered protocadherins. Collectively, this study validates SNR-derived organoids as a reliable new model for studying human telencephalic development and identifies novel molecular pathways disrupted by SHANK3 hemizygosity in human telencephalic tissue.
Project description:We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3’ and/or 5’ end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5’ differences and in support of this we report that a 5’ isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5’ isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes Sequence library of miRNAs from a single sample of human foetal mesenchymal stem cells. Results tested and confirmed by northern blotting. Please note that only raw data files are available for the embryonic and neual samples and thus, directly submitted to SRA (SRX547311, SRX548700, respectively under SRP042115/PRJNA247767)