Proteostasis collapse, a hallmark of aging and neurodegeneration like Alzheimer's disease (AD), causes irreversible damage in late life. Whether late-life proteostasis capacity is developmentally programmed remains unclear, as mechanistic studies requiring lifelong tracking and molecular manipulation are challenging or impossible in long-lived species. Using C. elegans as a lifelong, genetically tractable AD model, we uncover a critical early-life window during which reducing TIP60/NuA4 acetyltransferase complex activity enduringly enhances proteostasis and extends lifespan. Mechanistically, NuA4 reduction depletes H4K16ac, triggering a compensatory, early-life-biased, XBP-1-mediated unfolded protein response (UPR[ER]). This UPR[ER] activation remodels endoplasmic reticulum (ER) morphology and reprograms lipid metabolism, driving selective oleic acid (OA) accumulation. Crucially, this developmentally-installed OA reservoir confers lasting resilience against proteotoxic stress, an effect mimicked by OA supplementation. Together, these findings establish a chromatin-ER-lipid axis that developmentally primes adult proteostasis and suggest early-life interventions as a strategy to promote healthy aging and resilience to proteotoxic stress.

The impact of the metabolic microenvironment on epigenetically plastic cancer cells underpins phenotypic heterogeneity, a major cause of metastasis and therapy resistance. Nutrient limitation is a key microenvironmental stress, and can cause cells to transition from proliferative to invasive phenotypes, however, whether cancer cells have the capacity to delay phenotype switching remains unknown. Here, using melanoma as a model, we reveal that the ability to buffer glucose availability by accumulating and mobilizing glycogen can determine cancer cell phenotypic transitions. While proliferating cells contain high levels of glycogen, invasive cells are marked by depleted glycogen stores. Accordingly, the inability to store and metabolize glycogen leads to phenotype instability and a switch from proliferation to invasion. The amount of stored glycogen inversely correlates with tissue invasion depth in primary melanomas, and reduced expression of the glycogen phosphorylases PYGB/L and phosphoglucomutase 1 (PGM1) is associated with worse patient survival. Together, we identify metabolic glucose buffering as a determinant of invasive phenotype transitions in skin cancer, suggesting similar paradigms in other cancer types.

Costimulatory receptors of the TNFR superfamily are key components in synthetic immunoreceptors, yet how their intracellular domains coordinate membrane anchoring with adaptor recruitment remains unresolved. Here, using solution-state NMR and membrane-reconstituted FRET assays, we identify a conserved basic-residue-rich sequence (BRS) in the juxtamembrane region of OX40 that binds anionic phospholipids and adopts a membrane-inserted conformation. This membrane engagement enables a dual-site interaction with TRAF2, combining canonical PxQxx binding with a secondary membrane-proximal interface, to promote robust NF-κB activation. Disruption of the BRS impairs adaptor docking without reducing receptor surface expression or ligand-induced clustering. This regulatory architecture is further supported in 4-1BB and CD40, indicating broader relevance across TNFR costimulatory receptors. Our findings refine current models of TNFR signaling and offer a generalizable framework for receptor design. Incorporating membrane-anchoring motifs into synthetic constructs may enhance spatial control of adaptor recruitment, thereby improving signal fidelity, sensitivity, and performance in CAR-T applications.

Anabolic and catabolic processes are coordinated by a conserved regulatory network, which includes the nutrient-sensing protein kinase mTOR complex 1 (mTORC1) and the insulin- and stress-responsive transcription factor FoxO. In a physiological setting, these regulators align growth, storage, reproduction, and aging with nutrient availability. Here, we identify transcription factor Spalt-related (Salr), previously implicated in organogenesis, as a negative regulator of growth and lipid storage in Drosophila melanogaster. Salr activates catabolic gene expression and restricts mTORC1-mediated cell growth in the Drosophila fat body. The genomic binding of Salr overlaps extensively with that of FoxO, and a similar convergence is observed for their mammalian homologs, SALL1 and FOXO1. Both Salr and FoxO are activated upon fasting, but respond to distinct cues: while FoxO displays transient activation and is responsive to AKT inhibition, Salr is activated in a slow and sustained manner through the integrated stress response. Once activated, Salr counters nuclear localization of FoxO. Taken together, we show that Salr and FoxO are growth-inhibitory transcription factors that act in a convergent manner to respond to nutrient stress through distinct cues.

Mutations in KRAS are a dominant driver of pancreatic ductal adenocarcinoma (PDAC), with about 50% of patients presenting with KRAS[G12D] mutations. Small molecule inhibitors targeting KRAS[G12D] suppress PDAC; however, the contribution of the tumor microenvironment (TME) to the sustained efficacy of KRAS[G12D] inhibition and mechanisms of resistance to KRAS[G12D] suppression remain to be elucidated. Here, integrated spatial transcriptomics, single-cell RNA sequencing, and CODEX-based spatial proteomics analyses of PDAC mouse models uncover that while KRAS[G12D] inhibition by MRTX1133 initially increases CD11c[+] cells and T cell infiltration proximal to cancer cells, long-term treatment results in reversal of the immune responses leading to resistance promoted by multiprotein mediator complex associated kinase CDK8. CDK8 imparts this resistance via induction of CXCL2 chemokine secretion, inhibition of FAS expression, and remodeling of the TME to promote immune evasion. Targeting CDK8 by itself or in combination with αCTLA-4 immunotherapy overcomes resistance to KRAS[G12D] inhibition. We also provide evidence of CDK8 upregulation in PDX tumors resistant to inhibitors selective for RAS(ON) and RAS[G12D](ON): daraxonrasib and zoldonrasib, respectively, highlighting a common KRAS vulnerability node for TME resistance.

EGFR hotspot mutations (mEGFR), including primary L858R, exon 19 deletion, and secondary T790M, are pivotal oncogenic drivers in human non-small cell lung cancer (NSCLC). At the same time, NSCLC resistance to third-generation tyrosine kinase inhibitors (TKIs) is a major clinical challenge and remains mechanistically unresolved. Here, we uncover a previously unrecognized tumor cell-intrinsic mechanism in which mutant EGFR (mEGFR) exploits innate immune signaling via the cGAS-STING-TBK1 pathway to sustain oncogenic signaling and therapeutic resistance. Mechanistically, mutant EGFR kinase aberrantly associates with STING signalosomes and phosphorylates STING (Y245/Y314) and TBK1 (Y577/Y677), stabilizing and hyperactivating TBK1 and establishing an unexpected kinase loop critical for DNA damage repair. Genetic or pharmacological disruption of mEGFR-STING-TBK1 coupling sensitizes resistant patient-derived NSCLC organoids to chemotherapy. Combining TBK1 inhibition with cisplatin suppressed mEGFR-driven tumors in murine models of spontaneous and immunocompetent NSCLC and in patient-derived organoids. Our findings suggest a new function of cGAS-STING in DNA damage tolerance, its paradoxical exploitation by oncogenic driver mutations, and an innate immune therapeutic vulnerability in NSCLC.

Research on mitosis has predominantly examined cell-autonomous mechanisms, yet its regulation and consequences within the broader physiological context remain largely unexplored. It remains, therefore, unknown whether and how spindle assembly and chromosome segregation are influenced by tissue properties. We have investigated this question in the mammalian embryonic brain, using high-resolution microscopy combined with pharmacological perturbations and minimal cell systems to break down the contribution of tissue environment to mitotic fidelity. We found that cell density imposes biomechanical constraints upon the apical radial glial (aRG) cell population during their highly proliferative period, which enhances spindle pole microtubule polymerization rates and potentiates chromosome mis-segregation. Mechanistically, we show that cortical tension relying on branched actin organization at the cell cortex conveys mechanical stress exerted by cell density. We identified the microtubule depolymerase MCAK/Kif2C as a critical effector downstream of cortical actin, linking actin dynamics to spindle pole activity. Altogether, our findings demonstrate that aRG are mechanosensitive during their expansion phase, and that excessive biomechanical stress, imposed by high cell density in developing tissues, can disrupt mitotic fidelity.

2-cell like cells (2CLC) are a transiently cycling population of cells with totipotent-associated features. Although CTCF depletion induces 2CLC conversion in mouse ESC, whether this reprogramming is a consequence of disrupted higher-order chromatin organization or of CTCF-specific functions remained unclear. Here, we show that depletion of the cohesin release factor WAPL in ESC also promotes 2CLC reprogramming, which is increased by CTCF co-depletion. Single-cell RNA-seq/ATAC-seq analyses in CTCF/WAPL-depleted ESC revealed that chromatin accessibility precedes 2C-associated gene expression. Moreover, we identified ARID3A as a transcription factor that regulates the extent of 2CLC conversion following WAPL/CTCF depletion. Although WAPL or CTCF depletion induces distinct transcriptional changes in human ESC, these do not resemble transcriptional programs of early human embryogenesis, suggesting limited evolutionary conservation. Finally, we demonstrate that 2CLC conversion mediated by alterations in chromatin organization depends on the DPPA2/DUX axis and correlates with nucleolar integrity. Together, these findings establish a mechanistic link between higher-order chromatin organization and totipotency-like cell identity in mice.

Telomeres preserve stable eukaryotic chromosomes by protecting the natural chromosome ends from DNA repair but pose a persistent challenge to the replication machinery and an endogenous source of replication stress. Different features have been implicated in causing this effect but how the canonical replication process is altered at telomeres remains poorly understood. To address this question, we have reconstituted telomere replication with purified human proteins. Our system reveals that G-rich telomeric DNA can directly and specifically block lagging strand replication in a manner counteracted by BLM helicase. Unexpectedly, we also identify shelterin as inhibitory for the lagging strand. Biochemical experiments and electron microscopy imaging show that POT1-containing shelterin complexes induce Okazaki fragment skipping by binding the lagging strand template, generating large single-stranded gaps that are left behind on otherwise fully replicated molecules. Our study defines how the core components of telomeres interfere with the canonical replication process, identifying multiple potential sources of replication stress.