Cell Discovery最新文献

Human cerebellum encompasses numerous neurons, exhibiting a distinct developmental paradigm from cerebrum. Here we conducted scRNA-seq, scATAC-seq and spatial transcriptomic analyses of fetal samples from gestational week (GW) 13 to 18 to explore the emergence of cellular diversity and developmental programs in the developing human cerebellum. We identified transitory granule cell progenitors that are conserved across species. Special patterns in both granule cells and Purkinje cells were dissected multidimensionally. Species-specific gene expression patterns of cerebellar lobes were characterized and we found that PARM1 exhibited inconsistent distribution in human and mouse granule cells. A novel cluster of potential neuroepithelium at the rhombic lip was identified. We also resolved various subtypes of Purkinje cells and unipolar brush cells and revealed gene regulatory networks controlling their diversification. Therefore, our study offers a valuable multi-omics landscape of human fetal cerebellum and advances our understanding of development and spatial organization of human cerebellum.

Adenine base editors (ABEs) and cytosine base editors (CBEs) enable the single nucleotide editing of targeted DNA sites avoiding generation of double strand breaks, however, the genomic features that influence the outcomes of base editing in vivo still remain to be characterized. High-throughput datasets from lentiviral integrated libraries were used to investigate the sequence features affecting base editing outcomes, but the effects of endogenous factors beyond the DNA sequences are still largely unknown. Here the base editing outcomes of ABE and CBE were evaluated in mammalian cells for 5012 endogenous genomic sites and 11,868 genome-integrated target sequences, with 4654 genomic sites sharing the same target sequences. The comparative analyses revealed that the editing outcomes of ABE and CBE at endogenous sites were substantially different from those obtained using genome-integrated sequences. We found that the base editing efficiency at endogenous target sites of both ABE and CBE was influenced by endogenous factors, including epigenetic modifications and transcriptional activity. A deep-learning algorithm referred as BE_Endo, was developed based on the endogenous factors and sequence information from our genomic datasets, and it yielded unprecedented accuracy in predicting the base editing outcomes. These findings along with the developed computational algorithms may facilitate future application of BEs for scientific research and clinical gene therapy.

Ketogenic diet (KD) alleviates refractory epilepsy and reduces seizures in children. However, the metabolic/cell biologic mechanisms by which the KD exerts its antiepileptic efficacy remain elusive. Herein, we report that KD-produced β-hydroxybutyric acid (BHB) augments brain gamma-aminobutyric acid (GABA) and the GABA/glutamate ratio to inhibit epilepsy. The KD ameliorated pentetrazol-induced epilepsy in mice. Mechanistically, KD-produced BHB, but not other ketone bodies, inhibited HDAC1/HDAC2, increased H3K27 acetylation, and transcriptionally upregulated SIRT4 and glutamate decarboxylase 1 (GAD1). BHB-induced SIRT4 de-carbamylated and inactivated glutamate dehydrogenase to preserve glutamate for GABA synthesis, and GAD1 upregulation increased mouse brain GABA/glutamate ratio to inhibit neuron excitation. BHB administration in mice inhibited epilepsy induced by pentetrazol. BHB-mediated relief of epilepsy required high GABA level and GABA/glutamate ratio. These results identified BHB as the major antiepileptic metabolite of the KD and suggested that BHB may serve as an alternative and less toxic antiepileptic agent than KD.

Class B1 G protein-coupled receptors (GPCRs) are important regulators of many physiological functions such as glucose homeostasis, which is mainly mediated by three peptide hormones, i.e., glucagon-like peptide-1 (GLP-1), glucagon (GCG), and glucose-dependent insulinotropic polypeptide (GIP). They trigger a cascade of signaling events leading to the formation of an active agonist-receptor-G protein complex. However, intracellular signal transducers can also activate the receptor independent of extracellular stimuli, suggesting an intrinsic role of G proteins in this process. Here, we report cryo-electron microscopy structures of the human GLP-1 receptor (GLP-1R), GCG receptor (GCGR), and GIP receptor (GIPR) in complex with G_s proteins without the presence of cognate ligands. These ligand-free complexes share a similar intracellular architecture to those bound by endogenous peptides, in which, the G_s protein alone directly opens the intracellular binding cavity and rewires the extracellular orthosteric pocket to stabilize the receptor in a state unseen before. While the peptide-binding site is partially occupied by the inward folded transmembrane helix 6 (TM6)-extracellular loop 3 (ECL3) juncture of GIPR or a segment of GCGR ECL2, the extracellular portion of GLP-1R adopts a conformation close to the active state. Our findings offer valuable insights into the distinct activation mechanisms of these three important receptors. It is possible that in the absence of a ligand, the intracellular half of transmembrane domain is mobilized with the help of G_s protein, which in turn rearranges the extracellular half to form a transitional conformation, facilitating the entry of the peptide N-terminus.

Histone variant H2A.Z is found at promoters and regulates transcription. The ATP-dependent chromatin remodeler SRCAP complex (SRCAP-C) promotes the replacement of canonical histone H2A-H2B dimer with H2A.Z-H2B dimer. Here, we determined structures of human SRCAP-C bound to H2A-containing nucleosome at near-atomic resolution. The SRCAP subunit integrates a 6-subunit actin-related protein (ARP) module and an ATPase-containing motor module. The ATPase-associated ARP module encircles half of the nucleosome along the DNA and may restrain net DNA translocation, a unique feature of SRCAP-C. The motor module adopts distinct nucleosome binding modes in the apo (nucleotide-free), ADP-bound, and ADP-BeF_x-bound states, suggesting that ATPase-driven movement destabilizes H2A-H2B by unwrapping the entry DNA and pulls H2A-H2B out of nucleosome through the ZNHIT1 subunit. Structure-guided chromatin immunoprecipitation sequencing analysis confirmed the requirement of H2A-contacting ZNHIT1 in maintaining H2A.Z occupancy on the genome. Our study provides structural insights into the mechanism of H2A-H2A.Z exchange mediated by SRCAP-C.