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Energetic stress compels cells to evolve adaptive mechanisms to adjust their metabolism. Inhibition of mTOR kinase complex 1 (mTORC1) is essential for cell survival during glucose starvation. How mTORC1 controls cell viability during glucose starvation is not well understood. Here we show that the mTORC1 effectors eukaryotic initiation factor 4E binding proteins 1/2 (4EBP1/2) confer protection to mammalian cells and budding yeast under glucose starvation. Mechanistically, 4EBP1/2 promote NADPH homeostasis by preventing NADPH-consuming fatty acid synthesis via translational repression of Acetyl-CoA Carboxylase 1 (ACC1), thereby mitigating oxidative stress. This has important relevance for cancer, as oncogene-transformed cells and glioma cells exploit the 4EBP1/2 regulation of ACC1 expression and redox balance to combat energetic stress, thereby supporting transformation and tumorigenicity in vitro and in vivo. Clinically, high EIF4EBP1 expression is associated with poor outcomes in several cancer types. Our data reveal that the mTORC1-4EBP1/2 axis provokes a metabolic switch essential for survival during glucose starvation which is exploited by transformed and tumor cells. How cells adapt to glucose starvation is still elusive. Here, Levy et al. show that the mTOR substrate 4EBP1 protects human, mouse, and yeast cells from glucose starvation and is exploited by cancer cells to promote tumorigenesis.

DNA methylation regulation involves multi-layered chromatin interactions that require remodeling proteins like the helicase, lymphoid-specific (HELLS). Here, we generate HELLS and DNA methyltransferase 3A and B (DNMT3A/B) knockout human pluripotent stem cells and report telomere-to-telomere maps of whole genome bisulfite sequencing data combined with ATAC-sequencing. Disrupting HELLS induces a global loss of DNA methylation that is distinct from the DNMTs, in particular over peri/centromeric satellite repeats as defined in the telomere-to-telomere genome assembly. However, HELLS appears dispensable for local enhancer remodeling and the potential to differentiate into the three embryonic germ layers. Taken together, our results further clarify the genomic targets and role of HELLS in human cells.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13059-025-03681-9.

SummaryThe medial ganglionic eminence (MGE) gives rise to parvalbumin (PV)- and somatostatin (SST)-expressing cortical interneurons essential for regulating cortical excitability. Although PV interneurons are linked to various neurodevelopmental and neurodegenerative disorders, reliably generating them from human pluripotent stem cells (hPSCs) has been extremely challenging. We present a robust, reproducible protocol for generating single-rosette MGE organoids (MGEOs) from hPSCs. Transcriptomic analyses reveal that MGEOs exhibit MGE regional identity and faithfully model the developing human fetal MGE. As MGEOs mature, they generate abundant PV-expressing cortical interneurons, including putative basket and axoaxonic cells, at a scale not previously achieved in vitro. When fused with hPSC-derived cortical organoids, these interneurons rapidly migrate into cortical regions, integrate into excitatory networks, and contribute to complex electrophysiological patterns and the emergence of large numbers of fast-spiking neurons. MGEOs thus offer a powerful in vitro approach for probing human MGE-lineage cortical and subcortical GABAergic neuron development, modeling various neuropsychiatric disorders, and advancing cell-based therapies for neurodevelopmental and neurodegenerative disorders. Graphical abstract

ABSTRACTMedium chain acyl?CoA dehydrogenase deficiency (MCADD) is an inherited metabolic disease, characterized by biallelic variants in the ACADM gene. Interestingly, even with the same genotype, patients often present with very heterogeneous symptoms, ranging from fully asymptomatic to life?threatening hypoketotic hypoglycemia. The mechanisms underlying this heterogeneity remain unclear. Therefore, there is a need for in vitro models of MCADD that recapitulate the clinical phenotype as a tool to study the pathophysiology of the disease. Fibroblasts of control and symptomatic MCADD patients with the c.985A>G (p.K329E) were reprogrammed into induced pluripotent stem cells (iPSCs). iPSCs were then differentiated into hepatic expandable organoids (EHOs), further matured to Mat?EHOs, and functionally characterized. EHOs and Mat?EHOs performed typical hepatic metabolic functions, such as albumin and urea production. The organoids metabolized fatty acids, as confirmed by acyl?carnitine profiling and high?resolution respirometry. MCAD protein was fully ablated in MCADD organoids, in agreement with the instability of the mutated MCAD protein. MCADD organoids accumulated medium?chain acyl?carnitines, with a strongly elevated C8/C10 ratio, characteristic of the biochemical phenotype of the disease. Notably, C2 and C14 acyl?carnitines were found decreased in MCADD Mat?EHOs. Finally, MCADD organoids exhibited differential expression of genes involved in ??oxidation, mitochondrial ??oxidation, TCA cycle, and peroxisomal coenzyme A metabolism, particularly upregulation of NUDT7. iPSC?derived organoids of MCADD patients recapitulated the major biochemical phenotype of the disease. Mat?EHOs expressed relevant pathways involved in putative compensatory mechanisms, notably CoA metabolism and the TCA cycle. The upregulation of NUDT7 expression may play a role in preventing excessive accumulation of dicarboxylic acids in MCADD. This patient?specific hepatic organoid system is a promising platform to study the phenotypic heterogeneity between MCADD patients.

SummaryKnowledge of cell signaling pathways that drive human neural crest differentiation into craniofacial chondrocytes is incomplete, yet essential for using stem cells to regenerate craniomaxillofacial structures. To accelerate translational progress, we developed a differentiation protocol that generated self-organizing craniofacial cartilage organoids from human embryonic stem cell-derived neural crest stem cells. Histological staining of cartilage organoids revealed tissue architecture and staining typical of elastic cartilage. Protein and post-translational modification (PTM) mass spectrometry and snRNA-seq data showed that chondrocyte organoids expressed robust levels of cartilage extracellular matrix (ECM) components: many collagens, aggrecan, perlecan, proteoglycans, and elastic fibers. We identified two populations of chondroprogenitor cells, mesenchyme cells and nascent chondrocytes, and the growth factors involved in paracrine signaling between them. We show that ECM components secreted by chondrocytes not only create a structurally resilient matrix that defines cartilage, but also play a pivotal autocrine cell signaling role in determining chondrocyte fate. Graphical abstract Highlights•Craniofacial cartilage organoids were grown from human neural crest stem cells•These organoids exhibited elastic cartilage architecture and characteristic markers•Paracrine signaling drove chondrogenesis in mesenchyme cells and nascent chondrocytes•ECM components cemented chondrocyte cell fate through autocrine signaling Natural sciences; Biological sciences; Biochemistry; Cell biology; Stem cells research; Specialized functions of cells