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The complex 16p11.2 deletion syndrome (16pdel) is accompanied by neurological disorders, including epilepsy, autism spectrum disorder, and intellectual disability. We demonstrated that 16pdel iPSC differentiated neurons from affected people show augmented local field potential activity and altered ceramide-related lipid species relative to unaffected. FAM57B, a poorly characterized gene in the 16p11.2 interval, has emerged as a candidate tied to symptomatology. We found that FAM57B modulates ceramide synthase (CerS) activity, but is not a CerS per se. In FAM57B mutant human neuronal cells and zebrafish brain, composition and levels of sphingolipids and glycerolipids associated with cellular membranes are disrupted. Consistently, we observed aberrant plasma membrane architecture and synaptic protein mislocalization, which were accompanied by depressed brain and behavioral activity. Together, these results suggest that haploinsufficiency of FAM57B contributes to changes in neuronal activity and function in 16pdel syndrome through a crucial role for the gene in lipid metabolism.

AbstractUnderstanding the molecular underpinnings of pluripotency is a prerequisite for optimal maintenance and application of embryonic stem cells (ESCs). While the protein-protein interactions of core pluripotency factors have been identified in mouse ESCs, their interactome in human ESCs (hESCs) has not to date been explored. Here we mapped the OCT4 interactomes in naïve and primed hESCs, revealing extensive connections to mammalian ATP-dependent nucleosome remodeling complexes. In naïve hESCs, OCT4 is associated with both BRG1 and BRM, the two paralog ATPases of the BAF complex. Genome-wide location analyses and genetic studies reveal that these two enzymes cooperate in a functionally redundant manner in the transcriptional regulation of blastocyst-specific genes. In contrast, in primed hESCs, OCT4 cooperates with BRG1 and SOX2 to promote chromatin accessibility at ectodermal genes. This work reveals how a common transcription factor utilizes differential BAF complexes to control distinct transcriptional programs in naïve and primed hESCs.

Microglia are essential for maintenance of normal brain function, with dysregulation contributing to numerous neurological diseases. Protocols have been developed to derive microglia-like cells from human induced pluripotent stem cells (hiPSCs). However, primary microglia display major differences in morphology and gene expression when grown in culture, including down-regulation of signature microglial genes. Thus, in vitro differentiated microglia may not accurately represent resting primary microglia. To address this issue, we transplanted microglial precursors derived in vitro from hiPSCs into neonatal mouse brains and found that the cells acquired characteristic microglial morphology and gene expression signatures that closely resembled primary human microglia. Single-cell RNA-sequencing analysis of transplanted microglia showed similar cellular heterogeneity as primary human cells. Thus, hiPSCs-derived microglia transplanted into the neonatal mouse brain assume a phenotype and gene expression signature resembling that of resting microglia residing in the human brain, making chimeras a superior tool to study microglia in human disease., National Institutes of Health (Grant R01 AG058002-01), National Institutes of Health (Grant R01 MH104610), National Institutes of Health (Grant R37 CA084198), National Institutes of Health (Grant U19 AI131135)

Embryonic stem cells (ESCs) of mice and humans have distinct molecular and biological characteristics, raising the question of whether an earlier, “naive” state of pluripotency may exist in humans. Here we took a systematic approach to identify small molecules that support self-renewal of naive human ESCs based on maintenance of endogenous OCT4 distal enhancer activity, a molecular signature of ground state pluripotency. Iterative chemical screening identified a combination of five kinase inhibitors that induces and maintains OCT4 distal enhancer activity when applied directly to conventional human ESCs. These inhibitors generate human pluripotent cells in which transcription factors associated with the ground state of pluripotency are highly upregulated and bivalent chromatin domains are depleted. Comparison with previously reported naive human ESCs indicates that our conditions capture a distinct pluripotent state in humans that closely resembles that of mouse ESCs. This study presents a framework for defining the culture requirements of naive human pluripotent cells., Simons Foundation (Grant SFLIFE 286977), National Institutes of Health (U.S.) (Grant RO1-CA084198), National Science Foundation (U.S.). Graduate Research Fellowship, Jerome and Florence Brill Graduate Student Fellowship

Knowledge of both the global chromatin structure and the gene expression programs of human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) should provide a robust means to assess whether the genomes of these cells have similar pluripotent states. Recent studies have suggested that ESCs and iPSCs represent different pluripotent states with substantially different gene expression profiles. We describe here a comparison of global chromatin structure and gene expression data for a panel of human ESCs and iPSCs. Genome-wide maps of nucleosomes with histone H3K4me3 and H3K27me3 modifications indicate that there is little difference between ESCs and iPSCs with respect to these marks. Gene expression profiles confirm that the transcriptional programs of ESCs and iPSCs show very few consistent differences. Although some variation in chromatin structure and gene expression was observed in these cell lines, these variations did not serve to distinguish ESCs from iPSCs.

Glioblastoma (GBM) harbors subpopulations of therapy-resistant tumor-initiating cells (TICs) that are self-renewing and multipotent. To understand the regulation of the TIC state, we performed an image-based screen for genes regulating GBM TIC maintenance and identified ZFHX4, a 397 kDa transcription factor. ZFHX4 is required to maintain TIC-associated and normal human neural precursor cell phenotypes in vitro, suggesting that ZFHX4 regulates differentiation, and its suppression increases glioma-free survival in intracranial xenografts. ZFHX4 interacts with CHD4, a core member of the nucleosome remodeling and deacetylase (NuRD) complex. ZFHX4 and CHD4 bind to overlapping sets of genomic loci and control similar gene expression programs. Using expression data derived from GBM patients, we found that ZFHX4 significantly affects CHD4-mediated gene expression perturbations, which defines ZFHX4 as a master regulator of CHD4. These observations define ZFHX4 as a regulatory factor that links the chromatin-remodeling NuRD complex and the GBM TIC state., American Cancer Society (Postdoctoral Fellowship), American Brain Tumor Association (Discovery Grant), National Institutes of Health (U.S.) (Grant P30CA016672), National Institutes of Health (U.S.) (Grant U24CA143883), European Leukodystrophy Association, Japan. Ministry of Education, Culture, Sports, Science and Technology (MEXT/JSPS (KAKENHI 256701), Broad Institute of MIT and Harvard, National Institutes of Health (U.S.) (NIH (R01GM089652), National Institutes of Health (U.S.) (NIH (U01CA168426), National Institutes of Health (U.S.) (NIH U54CA121852), National Institutes of Health (U.S.) (NIH (R01HG002668), National Institutes of Health (U.S.) (NIH R01CA146455), National Institutes of Health (U.S.) (NIH (R01CA170592), National Institutes of Health (U.S.) (NIH P01CA095616), Sontag Foundation, Goldhirsh Foundation, National Institutes of Health (U.S.) (NIH R01NS080944), Starr Foundation, National Institutes of Health (U.S.) (NIH (R01CA129105), David H. Koch Institute for Integrative Cancer Research at MIT, National Institutes of Health (U.S.) (NIH P01CA142536), National Institutes of Health (U.S.) (NIH U01CA176058), National Institutes of Health (U.S.) (NIH K08NS062907), National Institutes of Health (U.S.) (NIH K12CA090354), American Association for Cancer Research, National Brain Tumor Society (fellowship), Dana-Farber Cancer Institute (Pediatric Low Grade Astrocytoma Program grant), Broad Institute of MIT and Harvard (SPARC grant), Howard Hughes Medical Institute (Investigator)

Glioblastoma (GBM) harbors subpopulations of therapy-resistant tumor-initiating cells (TICs) that are self-renewing and multipotent. To understand the regulation of the TIC state, we performed an image-based screen for genes regulating GBM TIC maintenance and identified ZFHX4, a 397 kDa transcription factor. ZFHX4 is required to maintain TIC-associated and normal human neural precursor cell phenotypes in vitro, suggesting that ZFHX4 regulates differentiation, and its suppression increases glioma-free survival in intracranial xenografts. ZFHX4 interacts with CHD4, a core member of the nucleosome remodeling and deacetylase (NuRD) complex. ZFHX4 and CHD4 bind to overlapping sets of genomic loci and control similar gene expression programs. Using expression data derived from GBM patients, we found that ZFHX4 significantly affects CHD4-mediated gene expression perturbations, which defines ZFHX4 as a master regulator of CHD4. These observations define ZFHX4 as a regulatory factor that links the chromatin-remodeling NuRD complex and the GBM TIC state., American Cancer Society (Postdoctoral Fellowship), American Brain Tumor Association (Discovery Grant), National Institutes of Health (U.S.) (Grant P30CA016672), National Institutes of Health (U.S.) (Grant U24CA143883), European Leukodystrophy Association, Japan. Ministry of Education, Culture, Sports, Science and Technology (MEXT/JSPS (KAKENHI 256701), Broad Institute of MIT and Harvard, National Institutes of Health (U.S.) (NIH (R01GM089652), National Institutes of Health (U.S.) (NIH (U01CA168426), National Institutes of Health (U.S.) (NIH U54CA121852), National Institutes of Health (U.S.) (NIH (R01HG002668), National Institutes of Health (U.S.) (NIH R01CA146455), National Institutes of Health (U.S.) (NIH (R01CA170592), National Institutes of Health (U.S.) (NIH P01CA095616), Sontag Foundation, Goldhirsh Foundation, National Institutes of Health (U.S.) (NIH R01NS080944), Starr Foundation, National Institutes of Health (U.S.) (NIH (R01CA129105), David H. Koch Institute for Integrative Cancer Research at MIT, National Institutes of Health (U.S.) (NIH P01CA095616), National Institutes of Health (U.S.) (NIH P01CA142536), National Institutes of Health (U.S.) (NIH U01CA176058), National Institutes of Health (U.S.) (NIH K08NS062907), National Institutes of Health (U.S.) (NIH K12CA090354), American Association for Cancer Research, National Brain Tumor Society (fellowship), Dana-Farber Cancer Institute (Pediatric Low Grade Astrocytoma Program grant), Broad Institute of MIT and Harvard (SPARC grant), Howard Hughes Medical Institute (Investigator)