Nature Cell Biology

Restoration of organellar membrane integrity is critical for maintaining cellular homeostasis. Lysosomal membrane damage activates local repair machineries and global stress responses, but how signaling lipid metabolism is engaged by damage sensors to support and mechanistically link these processes remains poorly understood. Here we show that the phosphoinositide 3-phosphatase MTMR14 is recruited to damaged lysosomes through calcium-dependent binding to sphingomyelin. At these sites, MTMR14 promotes local PI(3)P hydrolysis and supports PI(4)P accumulation, thereby facilitating formation of ER-lysosome contact sites associated with membrane repair, without affecting ESCRT recruitment. MTMR14-dependent lipid remodelling causes reduced mTORC1 signalling and a decrease in global protein synthesis, consistent with an acute proteostatic adaptation to lysosomal injury. Cells lacking MTMR14 display impaired damage-induced lipid remodelling, altered repair-associated structures, sustained protein synthesis, and increased sensitivity to lysosomal injury, all of which can be mitigated by mTORC1/S6K inhibition. Our findings identify damage-sensing recruitment of MTMR14 and local PI(3)P turnover on damaged lysosomes as a phosphoinositide module that promotes lysosomal membrane integrity and homeostasis while functionally linking nutrient signalling to proteostasis under membrane stress.

P-bodies are cytoplasmic membraneless organelles involved in mRNA storage, yet their role in cellular stress responses remains poorly understood. Here, we demonstrate that P-bodies are rapidly and selectively remodeled during the early response to endoplasmic reticulum (ER) stress in D. melanogaster oogenesis, positioning them as key early stress responders. Notably, this remodeling occurs within minutes of stress induction and precedes stress granule formation. This early remodeling is characterized by changes in P-body morphology and internal organization and promotes selective mRNA regulation. Specifically, ER stress leads to the recruitment and stabilization of maternal mRNAs and those encoding P-body components, while transcripts not associated with P-bodies are degraded. These observations indicate that P-body remodeling is not merely structural but functionally linked to the selective preservation of mRNA populations during stress. Mechanistically, we find that this process is driven by transcriptional upregulation of the RNA-binding protein, Bruno 1, downstream of ATF4-dependent stress signaling, thereby establishing a direct connection between the unfolded protein response and condensate regulation. Consistent with this model, loss of Bruno 1 abolishes, whereas its overexpression enhances P-body remodeling, demonstrating that stress-induced changes in RNA binding protein levels can actively reprogram condensate properties. Together, our findings reveal that P-bodies function as dynamic, stress-responsive hubs that integrate transcriptional signaling with post-transcriptional control, enabling the selective preservation of essential mRNAs during ER stress. More broadly, this work uncovers a previously unrecognized mechanism by which stress signaling pathways reorganize cytoplasmic architecture to shape mRNA fate.

How organelles communicate stress to the nucleus to coordinate adaptive responses remains a fundamental question in cell biology. Here, we identify a non-canonical retrograde signaling pathway in which stalling-induced UFMylation of ER-associated ribosomes anchors splicing regulators at the ER, directly coupling translational stress to nuclear RNA processing. Phylogenetic profiling linked the UFMylation machinery to a network of nuclear mRNA processing factors. Fractionation-based quantitative proteomics further supported this link and revealed that translational stress triggers UFM1-dependent retention of serine/arginine-rich (SR) splicing factors at the ER, depleting their nuclear pools. Mechanistically, UFMylated ribosomes physically tether SR proteins at the ER surface, driving widespread intron retention that preferentially targets transcripts encoding membrane lipid metabolism and endomembrane-associated processes--a response conserved from plants to mammals. These findings reframe UFMylation from a local ribosome repair signal to a systems-level coordinator of ER-nucleus communication that reprograms nuclear splicing and reshapes membrane-associated gene expression with implications for diverse human diseases linked to UFMylation defects.

Polyploid cells are increasingly recognized not only as hallmarks of cancer but also as features of regenerating tissues. During intestinal regeneration, polyploid cells are transient, yet the mechanisms underlying their clearance remain unknown. Using mouse intestinal organoids as a regeneration model, we show that, unlike in many cancer cell-line models, this clearance occurs without immediate cell-cycle arrest and is not driven by failure to establish spindle bipolarity. Instead, polyploid intestinal cells efficiently cluster supernumerary centrosomes to form bipolar spindles in an HSET-dependent manner, facilitated by delayed centrosome separation at mitotic onset. Despite this, polyploid divisions frequently produce chromosome segregation errors, including catastrophic chronocrisis. Lineage tracing reveals that progeny of such divisions is rapidly lost over subsequent generations. Increasing polyploidy during early regeneration disrupts organoid maturation, indicating that timely polyploidy clearance is required for successful regeneration. Polyploid cells are also detected in regenerating human colonic organoids, suggesting that transient polyploidy is a conserved feature of intestinal regeneration.

Primordial follicle oocyte activation (PFA) and zygotic genome activation (ZGA) represent two major waves of transcription activation respectively required for oocyte growth and preimplantation embryo development. Although many shared molecular hallmarks between PFA and ZGA suggest potential common factors and mechanisms driving both waves of transcriptional activation, such factors are yet to be identified. Here we demonstrate that the pioneer factor NFYA belongs to such regulators. Oocyte-specific Nfya deletion impairs open chromatin establishment and transcriptional activation during PFA, which triggers non-canonical ferroptosis leading to early folliculogenesis failure. Moreover, acute NFYA depletion in zygotes causes defective ZGA and predominantly two-cell embryo arrest. Mechanistically, although NFYA exhibits distinct chromatin-binding preferences predominantly targeting promoters during PFA and enhancers during ZGA, pre-occupied NFYA regulates chaperones and histone genes in both PFA and ZGA through conserved promoter binding. Together, our studies establish NFYA as a multifaceted regulator of genome activation during both PFA and ZGA. HighlightsO_LINFYA deficiency impairs primordial follicle oocyte activation (PFA) and triggers non-canonical ferroptosis resulting in early folliculogenesis failure C_LIO_LINFYA depletion impairs zygotic genome activation (ZGA) and causes predominantly 2-cell embryo arrest C_LIO_LIConserved and distinct NFYA-chromatin interactions drive both PFA and ZGA C_LIO_LIChaperones are pre-occupied and regulated by NFYA and their inhibition impairs both PFA and ZGA. C_LI

Exportin 1 (XPO1/CRM1) is the principal nuclear export receptor for cargos bearing hydrophobic nuclear export sequences (NESs). Dysregulation of XPO1-dependent export is implicated in cancer, neurodegeneration, and other diseases, yet a comprehensive view of XPO1 function remains limited by the poor reliability of sequence-based NES prediction. Existing predictors are largely derived from a small set of XPO1-cargo structures and are therefore biased toward canonical docking geometries, limiting their ability to detect NESs that engage XPO1 through noncanonical pocket-occupancy patterns. We hypothesized that deep learning-based structural modeling could overcome this limitation by directly sampling binding geometries. Using AlphaFold 3, we modeled full-length cargo-XPO1-RanGTP complexes for more than 4,000 human proteins and identified over 3,000 previously uncharacterized, high-confidence NESs. Integration of AlphaFold predictions with unsupervised structural geometry analysis and experimental validation identified both canonical NESs and noncanonical sequence patterns exhibiting atypical anchor-residue usage, expanding the structural language of XPO1-recognized NESs. Groove-resolved contact maps further revealed helix rotation within the export groove as a regulatory feature that can rewire pocket usage without altering the core NES sequence, enabling PTM- and cofactor-sensitive tuning of export strength. This exportome atlas resolves many previously ambiguous or unidentified NESs in disease-associated proteins and across major cellular systems, including centrosome organization, mRNA processing, ubiquitin signaling, kinase networks, ribosome quality control, and macroautophagy. We further identified recurrent NES-NLS tandem motifs encoded in primary sequence, suggesting coordinated regulation of nucleocytoplasmic transport. Together, our deep learning-based exportome atlas, integrated with NLS maps and accessible through a web-searchable resource, defines an expanded and regulatable code of nuclear transport at proteome scale and offers a framework for dissecting nuclear trafficking and its dysregulation in human disease.

Cells in vivo experience mechanically diverse microenvironments in which physical confinement is a pervasive but poorly understood regulator of their behavior and fate. Whether and how mechanical confinement governs immune cell differentiation remains unknown. Here, we reveal that a mechanical cue -- long-term confinement is sufficient to drive monocyte-to-macrophage differentiation through a mechanoepigenetic pathway. In vivo, differentiating monocytes exhibited flattened nuclei in the liver capsule, indicative of confinement by surrounding stromal and parenchymal structures. Using a custom cell confiner to recapitulate this confined niche, we found that confinement induces macrophage-like protrusive architectures, enhances motility, and upregulates macrophage-associated genes in RAW264.7 and THP-1 monocyte-lineage cells. Notably, extending this paradigm to primary murine bone-marrow, human umbilical-cord, and tissue-derived hepatic-associated monocytes yielded similar outcomes, thus enhancing phagocytic capacity, directly demonstrating that mechanical confinement can program monocytes into macrophages. Mechanistically, we found that confinement activates KDM6B, leading to H3K27me3 demethylation, which derepresses macrophage-specific transcriptional programs. Pharmacological inhibition of KDM6B with GSK-J4 restored H3K27me3 and blocked macrophage differentiation both in vitro and in vivo. These findings define a KDM6B-H3K27me3 axis that links nuclear mechanics to transcriptional reprogramming, positioning mechanical confinement as a "super-enhancer-like" cue for engineer macrophage function in therapeutic and bioengineering contexts.

Although function often follows form, the causal role of tissue geometry is difficult to disentangle in complex embryonic development. Brain organoids generated using diverse protocols and species display striking morphological variability, particularly in lumen shape; however, whether and how lumen geometry influences neural development remains unclear. Here, we manipulate lumen sphericity in human cerebral organoids by acutely inducing apical constriction and reveal its impact on the division orientation of apical progenitors. Rapid protein stabilization or optogenetic reconstitution of the apical constriction regulator Shroom3 induces pronounced lumen rounding accompanied by a reduction in apical surface area. In organoids with rounded lumens, apical progenitor divisions shift toward horizontal cleavage planes compared with control organoids, consistent with geometric constraints from the reduced apical surface. Accordingly, rounded-lumen organoids exhibit increased cell delamination and an earlier emergence of basal progenitors in the abventricular region. These findings identify lumen geometry as an instructive regulator of progenitor division mode and lineage progression during early brain development.

Colonic stem cells reside in a microenvironment enriched in epidermal growth factor, which is essential for their survival and can activate both PI3K-AKT and MAPK-ERK pathways. This predicts co-activation of both pathways within the growth factor-high stem cell compartment at the base of crypts. However, in patient-derived human colonic organoids and normal human tissue, stem cells maintain robust AKT activity while suppressing ERK signaling despite active EGFR engagement. As stem cells differentiate, they activate pulsatile Erk signaling, which is essential for migration, survival, and maintenance of barrier function. We show that AKT-dependent phosphorylation of Raf-1 at serine 259 establishes a post-receptor checkpoint that maintains ERK temporal dynamics in stem cells. Acute activation of ERK in stem cells triggers rapid global differentiation. Disruption of the ERK checkpoint via mutation of serine 259 leads to sustained AKT and ERK co-activation in stem cells. Unlike ERK/AKT coactivation driven by apoptosis, co-activation in the stem cell compartment results in the emergence of a neoplastic, architecturally disorganized cell population dominating the cell fate profile. Incredibly, introducing brief ERK pulses through Akt inhibition or ERK activation triggers re-differentiation of neoplastic cells. Consistent with duration-dependent MAPK encoding principles, these data demonstrate that regardless of baseline signaling amplitude, ERK signaling dynamics are epistatic to total kinase signaling load in human colonic stem cells.

Stress granules (SGs) are dynamic RNA-RBP condensates that form during stress and inflammation, yet how they modulate inflammatory signalling remains unclear. We uncover a rapid, protective SG-mediated mechanism that preserves mitochondrial integrity. During stress and translation inhibition, mitochondrial fragmentation releases double-stranded RNA (dsRNA), which activates PKR and its downstream effector DRP1, generating a self-amplifying loop of mitochondrial fragmentation and inflammation. We find that released dsRNA nucleates nanoSGs within minutes at ER-mitochondria contact sites--the very sites of mitochondrial division. These nanoSGs grow into macroSGs, effectively sequestering PKR-activating dsRNA from the cytosol. By depleting dsRNA, SGs suppress PKR-DRP1-driven positive-feedback inflammation and maintain mitochondrial integrity and function. Our findings reveal SGs as key guardians of mitochondrial homeostasis and position condensate biology at the centre of chronic mitochondrial-driven inflammation relevant to autoimmunity, ageing, and neurodegenerative disease.

Nucleoporins are increasingly recognized as tissue-specific regulators beyond their structural roles in the nuclear pore complex. Here, we identify nucleoporin Nup358 as a critical repressor of Wnt signaling required for intestinal epithelium integrity. Ablation of Nup358 in adult mice causes a catastrophic loss of crypt-villus architecture and disrupts the intestinal epithelial layer. Notably, while the intestinal stem cell (ISC) pool remains stable, the transit-amplifying (TA) progenitor compartment is depleted. Mechanistically, we show that the interaction of Nup358 with Dvl1 through its N-terminal domain inhibits Dvl1 spontaneous phase separation. In the absence of Nup358, Dvl1 biomolecular condensates promote Tankyrase-mediated degradation of Axin1, leading to the constitutive stabilization of {beta}-catenin and ligand-independent Wnt activation, negatively impacting cell differentiation and TA progenitor survival. Our results demonstrate that Nup358 acts as a molecular safeguard that dampens Wnt signaling levels in intestinal crypts. By preventing Dvl1-mediated Wnt signal amplification, Nup358 allows ISCs to transition into the TA compartment and initiate the differentiation programs essential for intestinal homeostasis.

STING is a key innate immune adaptor, classically activated by cytosolic DNA via cGAS-cGAMP to induce type I interferon signaling. While its cytoplasmic role is well defined, recent studies reveal that STING participates in non-canonical signaling pathways and localizes at the nuclear envelope and chromatin, where its functions remain poorly understood. In Hutchinson Gilford Progeria Syndrome (HGPS), a premature aging disease caused by expression of lamin A mutant protein named progerin, STING accumulates in the nucleus and drives chronic inflammation. Here, we show that replication stress (RS) is a trigger of STING nuclear accumulation and binding to chromatin. In addition, we uncover a previously unrecognized role for nuclear STING binding to nascent DNA and promoting RS in progeria and tumor cells. Mechanistically, STING contributes to replication fork slowing and stalling by limiting dNTPs availability. In addition, STING hinders replication fork protection/stability upon stalling, by facilitating MRE11-mediated nascent DNA degradation (NDD). We also find that STING contribution to depletion of dNTPs and NDD is mediated by SAMHD1. As such, SAMHD1 knockdown phenocopies STING abrogation in progeria cells and rescues replication fork speed and stability in STING-overexpressing tumor cells. These findings define a pathological STING-SAMHD1 axis that drives RS and genome instability in both progeria cells and tumor cells with elevated STING activity, uncovering a feedforward loop between innate immune signaling and impaired DNA replication. HighlightsO_LIReplication stress in human fibroblasts triggers STING nuclear accumulation and an IFN response C_LIO_LISTING upregulation and nuclear accumulation hinders replication in progeria fibroblasts and U2OS tumor cells C_LIO_LISTING-induced replication stress features fork slowing/stalling and nascent DNA degradation C_LIO_LISTING-induced fork slowing/stalling is mediated by the dNTPase SAMHD1 C_LIO_LISAMHD1-enabled MRE11 activity is responsible for STING-induced nascent DNA degradation C_LI

When and how lineage competence first emerges in the epiblast remains a central question in mammalian development. During the transition from naive to formative pluripotency, epiblast cells acquire responsiveness to lineage-inducing cues, yet whether transcriptional heterogeneity in this window reflects a regulated programme of lineage emergence or stochastic variation, and which molecular regulators shape developmental competence and potential lineage biases, remain poorly defined. Here, using a targeted CRISPRa screen, we identify the developmental regulator Wilms tumor 1 (WT1) as an unexpectedly early regulator of formative pluripotency. WT1 is transiently induced during the transition to formative pluripotency in vitro and in vivo, with peak expression coinciding with the emergence of lineage-associated transcriptional biases. Precocious Wt1 induction overrides the naive transcriptional network and advances cells toward a post-implantation epiblast identity, even under naive-stabilizing conditions. Genome-wide binding analyses show that WT1 engages active regulatory elements of the emerging post-implantation gene regulatory network together with core formative transcription factors, including Otx2 and Oct4. Alternative WT1 splice isoforms encode distinct lineage-biased transcriptional programmes associated with anterior and posterior fates. In the E5.5 epiblast, WT1 expression and splice composition align with lineage-biased transcriptional states, linking isoform usage to anterior-posterior transcriptional tendencies in vivo. Isoform-dependent gene expression modules are conserved in human pluripotent cells, indicating that this regulatory logic is preserved across species. Together, our findings indicate that lineage-associated transcriptional programmes begin to diversify during formative pluripotency and identify WT1 as an isoform-tuned regulator that biases these transcriptional outputs.

Regeneration enables organisms to repair damaged tissues, yet this capacity is strikingly limited in the cochlear sensory epithelium, essential for sound detection. A major cause of hearing loss arises from the irreversible loss of sensory hair cells (HCs) in the cochlea. While supporting cells (SCs) have a latent ability to transdifferentiate into HCs, this regenerative potential is rapidly lost after development. Using live imaging and single-cell multi-omics of cochlear explants, we uncovered the cellular and molecular heterogeneity underlying the limited regenerative capacity of the neonatal mouse cochlea. Notch repression broadly silenced key SC genes, yet only a rare subpopulation of Deiters cells (DC), termed transdifferentiating DCs (tDCs), initiated the transdifferentiation into HC fate. These cells underwent coordinated transcriptional and enhancer remodeling, linking epigenetic priming with morphological plasticity, while other SCs remained refractory despite robust Notch targets downregulation. Our study provides a molecular definition of an early induced transitional DC to HC state, revealing Notch inhibition as a selective trigger that unmasks rare regenerative competence.

Plant immune responses must balance effective pathogen restriction to prevent excessive tissue damage and rely on potentiation between pattern-triggered immunity (PTI) and effector-triggered immunity (ETI). However, how these immune responses are spatially organized across different cell-types and coordinated by intracellular pathways remains incompletely understood. Here, we show that autophagy functions as a central organizer of this spatial immune response during Pseudomonas syringae infection in Arabidopsis thaliana. Combining single-cell transcriptomics, cell-type-specific complementation, and live-cell imaging, we uncover distinct and opposing roles of autophagy across tissues. In guard cells, autophagy promotes pathogen-induced stomatal reopening by supressing abscisic acid (ABA) signaling through selective degradation of the ABA receptor PYL4. In contrast, in mesophyll cells, autophagy restricts immune activation and is required for effective immune execution: its loss enhances expression of the EDS1-PAD4-ADR1 immune pathway but compromises canonical PTI outputs, likely impacting PTI-ETI potentiation. This uncoupling reveals that immune activation alone is insufficient for effective defense. Together, our findings resolve the longstanding ambiguity surrounding the role of autophagy in plant immunity and establish autophagy as a spatial organizer that partitions immune strategies between stomata and mesophyll during bacterial infection.

Macrophages are highly plastic innate immune cells that can form multinucleated giant cells (MGCs) under physiological and pathological conditions, such as osteoclasts and foreign body giant cells. The mechanisms governing macrophage multinucleation remain incompletely understood. Here, we identify activating transcription factor 3 (Atf3) as an essential regulator of MGC formation via cell-cell fusion in response to persistent Interleukin 4 (IL-4) and STAT6 signaling, characteristic of the foreign body reaction. Atf3-deficient macrophages activate STAT6-dependent transcriptional programs in response to IL-4, including fusion-associated genes, yet fail to undergo multinucleation. This defect is associated with impaired actin cytoskeleton remodeling, abnormal nuclear morphology, reduced lamin A/C expression, and genome instability. Mechanistically, loss of Atf3 derepresses the Cholesterol-25-hydroxylase (Ch25h), elevating 25-hydroxycholesterol (25-HC), suppressing the mevalonate pathway, and reducing cholesterol and isoprenoid biosynthesis essential for cytoskeletal dynamics. Deletion of Ch25h in Atf3-deficient macrophages restores cholesterol levels and actin turnover, but not lamin A/C or fusion. These findings establish Atf3 as a central transcriptional node integrating lipid metabolism with cytoskeletal and nucleoskeletal remodeling to control IL-4-driven macrophage multinucleation.

Mammalian somatic haploid cells offer advantages for genome engineering, yet rapid diploidization limits their utility. Here, we reveal that a haploidy-specific attenuation of mitotic spindle bipolarization, independent of previously characterized centrosome loss, underlies haploid instability in human cells. Comparative imaging and structure-function analyses demonstrate that the halved absolute dosage of the pericentriolar scaffolding protein Cep192 prevents its centrosomal accumulation to the threshold required for Aurora A-Eg5 axis. Consequently, haploids exhibit innate fragility in centrosome separation and spindle maintenance. Supplementing Cep192 restored spindle bipolarization to diploid levels and, when combined with genetic enhancement of the acentrosomal spindle pathway, profoundly stabilized the haploid state. Moreover, a genome-wide CRISPR-activation screen leveraging the above principle identified novel haploid-stabilizing genes, including the glutamate transporter SLC1A2. Our findings uncover an absolute-dosage scaling limit of mitotic scaffolding in haploids and establish genetic enhancement of spindle fidelity as an effective strategy for engineering stable animal haploid bioresources. TeaserMolecular elucidation of haploidy-linked fragility in mitotic spindle architecture enables engineering stable human haploid cells.

Mammalian fertilization commences with the essential interaction between sperm IZUMO1 and its oocyte-surface receptor, JUNO. Following gamete fusion, JUNO is rapidly shed from the oocyte to establish a definitive membrane block to polyspermy, a pathological condition that remains a major hurdle in porcine in vitro fertilization (IVF). Despite its biological importance, the molecular networks driving JUNO cleavage has remained elusive. Here, by integrating proteomics, dual-color live-cell imaging, and functional perturbations, we identify the extended existence of JUNO and the GPI-specific phospholipase D1 (GPLD1) as the requisite enzyme mediating JUNO shedding in porcine oocytes. Targeted knockdown or pharmacological inhibition of GPLD1 stabilizes oocyte JUNO, prolongs the window of oocyte receptivity, and significantly exacerbates polyspermy, ultimately compromising embryonic developmental competence. Conversely, GPLD1 overexpression restricts redundant sperm adherence and enhances the efficiency of monospermic zygote formation and blastocyst development. Live-cell imaging reveals that fertilization triggers a transient, pulsed recruitment of GPLD1 in the oocyte, which precisely coincides with the biphasic kinetics of JUNO depletion. Our findings establish that the enzymatic cleavage of the GPI-anchor by GPLD1 is critical for JUNO release, defining a fundamental mechanism for the membrane-level block to polyspermy. This work provides a molecular framework for ensuring sperm-oocyte recognition and improving in vitro fertilization outcomes in mammals.

Skeletal muscle stem cells (MuSCs) rely on precisely coordinated metabolic and nuclear transitions to exit quiescence, enter the cell cycle, and regenerate tissue. How these processes are coupled remains poorly defined. Here, we identify PARKIN as a critical integrator of mitochondrial quality control and nuclear RNA processing programs that together enable balanced MuSC lineage progression. Using a MuSC-specific, inducible Park2 knockout model, we show that PARKIN supports mitophagy in quiescent MuSCs, and its loss triggers premature mitochondrial polarization and fragmentation -- hallmarks of metabolic activation -- that compromise appropriate self-renewal and fate specification. Unexpectedly, MuSCs harbor a constitutive nuclear pool of PARKIN that rises rapidly upon activation and localizes to interchromatin regions, with focal association with nuclear speckles. Park2-deficient MuSCs exhibit transcriptomic signatures consistent with widespread RNA isoform switching and intron retention, particularly affecting splicing machinery components, accompanied by altered nuclear speckle organization and impaired cell cycle progression. These findings reveal that PARKIN safeguards both mitochondrial homeostasis and the RNA processing architecture essential for activation, thereby coordinating metabolic and nuclear reprogramming during early MuSC state transitions. Our work positions PARKIN as a dual compartment regulator required for robust skeletal muscle regeneration.

Regulation of protein synthesis is essential for maintaining cellular homeostasis during stress. The integrated stress response (ISR) is a conserved signaling pathway that modulates global mRNA translation through four eIF2 kinases--GCN2, PKR, PERK, and HRI. However, how these kinases are selectively activated and tuned to distinct stress signals to direct appropriate cell fate decisions remains poorly understood. Here, we employ ultra-deep mutagenesis screens to systematically map regulators of protein synthesis across diverse stress perturbations in human cells. This comparative approach identifies stress-specific translational control factors, including a previously unrecognized role for the E3 ubiquitin ligase RNF25 in selectively sustaining translation following UV irradiation and other RNA-damaging treatments. In this context, we demonstrate that RNF25 operates independently of its partner RNF14, and that its ubiquitin ligase activity, as well as its RWD-domain, is required to restrain excessive activation of the eIF2 kinase GCN2. Accordingly, loss of RNF25 results in hyperactivation of GCN2, exacerbated translation shutdown, and impaired cell proliferation following RNA damage--phenotypes that can be fully reversed by genetic or pharmacological inhibition of GCN2. Together, these findings uncover a previously unappreciated RNF25-GCN2 signaling axis and identify ISR-driven toxicity as a potential vulnerability in combination with RNA-damaging chemotherapeutics.