Biology subjects

Thibault, P.

Publications and source records attributed to Thibault, P..

At least 19 recordsLinked to original sources

bioRxiv2026

Tumour-stroma signaling drives a reversible program with neural-like features in cancer-associated fibroblasts

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers, in part due to a dense and highly plastic tumor microenvironment dominated by cancer-associated fibroblasts (CAFs). In previous work, we identified a CAF population expressing a neural-associated NES+SOX4+NOTCH1+ program that increased with disease progression. The signals driving this state and the mechanisms regulating it remained unclear. Here, we investigated how tumor-derived signals activate this program and whether it represents a stable fibroblast identity or a reversible state. Using spatial transcriptomic data spanning normal pancreas to invasive carcinoma, we identified EGF, FGF2 and BDNF as candidate tumor-to-fibroblast signals associated with disease progression. Functional experiments in patient-derived CAFs and human fibroblast cell lines showed that EGF and FGF2 are sufficient to induce the NES+SOX4+NOTCH1+ program. Growth factor withdrawal reversed the NES+SOX4+NOTCH1+ program at both the protein and transcriptional levels, indicating that this state is dynamic and environment dependent. Single-cell transcriptomic analysis further revealed a multi-phase response involving early loss of fibroblast-associated features, transient activation of neural-associated genes, and a later stress-associated state. Regulatory analyses identified ETS-family transcription factors as key regulators of the program, while phosphoproteomic profiling and inhibitor studies showed that MAPK/ERK signaling plays a central role in its transcriptional activation. Finally, pan-cancer single-cell analysis showed that this state is enriched in CAFs and increases from normal to tumor tissue across multiple cancer types. Together, these findings define a reversible, growth factor-driven CAF program and identify signaling mechanisms that may provide opportunities to selectively modulate tumor-supportive fibroblast states.

cancer biology↗

bioRxiv2026

STING dampens the unfolded protein response to enable the presentation of self-antigens on MHC-I during inflammation.

A growing body of evidence supports the contribution of the long-lasting adaptive immune system in Parkinsons disease (PD). We showed that the PD-associated protein PINK1 negatively regulates the presentation of mitochondrial antigens (MitAP) on MHC-I molecules. In vivo evidence indicated that MitAP activation in mice, in the absence of PINK1, led to cytotoxic CD8+ T cell stimulation and severe motor impairments, reversible by L-DOPA. We show here that following TLR4 activation, MitAP is engaged through a pathway involving cGAS-STING, which acts as a rheostat to dampen the unfolded protein response (UPR). Without STING, the stress response is amplified, leading to a translational attenuation that inhibits the expression of XBP1s, a transcription factor required for MitAP. STING activity also regulates the repertoire of peptides displayed at the cell surface during inflammation, highlighting a potential role in immunosurveillance. These findings establish STING and the UPR as key immune regulators targetable for therapeutic intervention during autoimmune diseases and PD.

immunology↗

bioRxiv2026

Synthetic analogue of adrenocorticotropic hormone, ACTH(4-7)PGP delays neurological manifestations in diseases of mucopolysaccharidosis III spectrum by reducing neuroinflammation and rescuing neurotransmission, synaptogenesis, and axonal demyelination

Mucopolysaccharidosis III (MPS III or Sanfilippo disease) is a spectrum of 4 genetic disorders (MPS IIIA-D), caused by defects in the genes SGSH, NAGLU, HGSNAT and GNS encoding enzymes involved in degradation of heparan sulfate (HS). HS accumulates in brain tissues and causes neuronal dysfunction and neurodegeneration leading to neuropsychiatric problems, developmental delays, childhood dementia, blindness and death during the second decade of life. Previously, we demonstrated that pathophysiological mechanisms, underlying MPS IIIC in mouse models, involves functional pathological changes, affecting synaptogenesis and synaptic transmission and leading to learning and memory deficits. These results suggested that a treatment for MPS III could be developed by using compounds inducing synaptogenesis. In the current study, we tested the efficacy of a synthetic peptide ACTH(4-7)PGP, an analog of adrenocorticotropic hormone fragment, previously used as a neuroprotective and anti-inflammatory medication for treatment of acute neurological conditions, including stroke. We show that intranasal administration of ACTH(4-7)PGP restores defective synaptic transmission in CA1 pyramidal neurons of MPS IIIA and MPS IIIC mouse models and rescues the decrease in synaptic proteins in cultured MPS IIIC mouse hippocampal neurons and iPSC-derived neurons of human MPS IIIA, MPS IIIB and MPS IIIC patients. Furthermore, daily intranasal administration of ACTH(4-7)PGP to MPS IIIC and MPS IIIA mice reduces hyperactivity and rescues defects in working and spatial memory, delays progression of CNS pathology including neuroinflammation and axonal demyelination, and increases the lifespan. Together with the absence of any adverse reactions to ACTH(4-7)PGP in the MPS III and WT mice, our results justify testing the drugs efficacy in clinical settings.

genetics↗

bioRxiv2026

Redundant roles of YES and SRC tyrosine kinases in driving malignant peripheral nerve sheath tumors

Malignant peripheral nerve sheath tumors (MPNST) are highly aggressive soft tissue sarcomas that are largely incurable with no clinically effective systemic therapies or immunotherapies for advanced disease. Here, we identify the SRC-family kinases (SFKs) YES and SRC as redundant, essential drivers of MPNST growth. Dual inhibition of YES/SRC activity by genetic silencing or pharmacological SFK inhibitors markedly suppressed the proliferation of multiple NF1-mutant MPNST cell lines. In vivo, conditional genetic depletion of YES/SRC in MPNST cells abrogated tumor growth in subcutaneous and orthotopic models, and dasatinib treatment delayed tumor progression and improved overall survival. Integrated transcriptomic and phosphotyrosine proteomic analyses revealed that YES/SRC inactivation extensively rewires MPNST signaling, coordinately repressing multiple oncogenic signaling pathways and downstream cell cycle transcriptional programs. Unexpectedly, YES/SRC inhibition also upregulated interferon and antigen processing and presentation pathways and increased cell-surface MHC class I expression, consistent with tumor-intrinsic immune reactivation. Clinically, analysis of a large sarcoma cohort demonstrated that YES1 is significantly overexpressed in MPNST compared to benign soft tissue tumors. Collectively, our findings establish YES/SRC as non-oncogene vulnerabilities in MPNST.

cancer biology↗

bioRxiv2026

A Chemical-Genetic Interaction Matrix Reveals Drug Mechanism and Genetic Architecture

To probe drug mechanism of action (MOA) and interrogate the genetic architecture of human cells, we carried out isogenic genome-wide CRISPR/Cas9 knockout screens against 310 diverse drugs, bioactive compounds, and stress conditions. Stringent statistical correction for gene knockout fitness defects yielded a large-scale matrix of >12,000 high confidence chemical-genetic interactions (CGIs). This dataset revealed many previously unappreciated off-target effects for well-characterized compounds and novel MOAs for uncharacterized compounds. The CGI matrix uncovered dense genetic modules that yielded new biological insights into phospholipidosis, mitotic regulation, metabolism, the DNA damage response, and mTOR signaling. The dataset allowed identification of multi-drug sensitization and resistance mechanisms, inference of gene function, elaboration of cross-process connectivity, evaluation of the cell type specificity of CGIs, prediction of chemical synergism, and extensive annotation of understudied genes. This resource provides a map of the genetic landscape in human cells and a framework to help guide drug discovery.

systems biology↗

bioRxiv2025

Splicing Isoforms Associated to TGFb-Induced Myofibroblast Activation

Myofibroblast differentiation is a key process in developmental biology and involved in numerous physiopathology. The gene expression program orchestrating fibroblast to myofibroblast differentiation, as well as its recapitulation by TGF{beta} stimulation in vitro, is relatively well characterized. Intriguingly, it is known that the splicing isoform EDA+FN1 is a marker and driver of myofibroblast differentiation, but the alternative splicing landscape of myofibroblast is unknown. Here, we performed a high-throughput transcriptomic approach by RNA-Seq in a primary skin fibroblast line and uncover more than 250 splicing isoforms associated with TGF{beta}-induced myofibroblasts using two different bioinformatic pipelines. This splicing profile highlights a distinct layer of regulation when compared to the global gene expression profile of myofibroblasts. A 5 alternative splicing event (ASE) signature [ACTN1-19A/19B; COL5A1-64A/64B; COL6A3 exon 4; FLNA exon 30 and TPM1-6a/6b] was further validated by ddPCR and AS-PCR and retrieved in publicly available RNA-Seq datasets describing other TGF{beta}-stimulated lung and skin fibroblasts. Surprisingly, TGF{beta} does not induce an EDA+FN1 splicing shift, although it stimulates global fibronectin expression. Thus, we conclude that the 5 ASEs signature may be used as putative universal myofibroblast markers and be of functional significance to myofibroblast formation and biology.

genomics↗

bioRxiv2025

LRRK2G2019S acts as a dominant interfering mutant in the context of iron overload

Altered iron homeostasis has long been implicated in Parkinsons Disease (PD), although the mechanisms have not been clear. Given the critical role of PD-related activating mutations in LRRK2 (leucine-rich repeat protein kinase 2) within membrane trafficking pathways we examined the impact of a homozygous mutant LRRK2G2019S on iron homeostasis within the RAW macrophage cell line with high iron capacity. Proteomics analysis revealed a dysregulation of iron-related proteins in steady state with highly elevated levels of ferritin light chain and a reduction of ferritin heavy chain. LRRK2G2019S mutant cells showed efficient ferritinophagy upon iron chelation, but upon iron overload there was a near complete block in the degradation of the ferritinophagy adaptor NCOA4. These conditions lead to an accumulation of phosphorylated Rab8 at the plasma membrane, which is selectively inhibited by LRRK type II kinase inhibitors. Iron overload then leads to increased oxidative stress and ferroptotic cell death. These data implicate LRRK2 as a key regulator of iron homeostasis and point to the need for an increased focus on the mechanisms of iron dysregulation in PD.

cell biology↗

bioRxiv2025

Mono-allelic p.R37H Dehydrodolichyl Diphosphate Synthase variants lead to protein glycosylation defects, aberrant lipid profiles and interneuron scarcity in a novel mouse model of progressive epileptic encephalopathy.

Developmental delay and seizures with or without movement abnormalities (OMIM 617836) caused by heterozygous pathogenic variants in the DHDDS gene (DHDDS-CDG) is a rare genetic disease that belongs to the progressive encephalopathy spectrum. It results in cognitive delay in affected children, accompanied by myoclonus, seizures, ataxia and tremor, which worsens over time. DHDDS encodes a subunit of a DHDDS/NUS1 cis-prenyltransferase (cis-PTase), a branch point enzyme of the mevalonate pathway essential for N-linked glycosylation. We describe the first mouse model of this disease, DhddsR37H+/-strain, heterozygous for the human recurrent de novo c.110G>A:p.R37H pathogenic variant. DhddsR37H+/- mice present with seizures, myoclonus and memory deficits associated with reduced density or/and maturity of inhibitory interneurons in the cortex. Multiomics analyses of mouse CNS tissues, together with the enzymatic/structural characterization of the R37H DHDDS mutant protein, reveal that the variant produces a catalytically inactive enzyme and results in a brain dolichol deficit, aberrant glycosylation of brain glycoproteins, including those involved in synaptic transmission and major perturbations in the CNS proteome and lipidome. Acetazolamide, a carbonic anhydrase inhibitor clinically approved for treatment of glaucoma, epilepsy, and intracranial hypertension, and successfully used "off-label" to treat genetic movement disorders, reduces seizure susceptibility to pentylenetetrazol in DhddsR37H+/- mice, suggesting potential therapeutic value of using this drug in human DHDDS-CDG patients. Together, our results define cis-PTase as a master regulator of CNS development and function and establish that its monoallelic debilitating variants cause a novel congenital disorder of glycosylation associated with aberrant levels of neuronal proteins and lipids.

neuroscience↗

bioRxiv2025

Enriching for Extracellular Vesicles from Human Bone

Extracellular vesicles (EVs) are nano-sized membrane-bound structures thought to be secreted by all cells and increasingly recognized as key mediators of intercellular communication. Established EV isolation protocols for bodily fluids-primarily focus on blood with limited insights into methods optimized for EVs from other hematopoietic regions. In this study, we present a novel protocol for the isolation and enrichment of EVs from human trabecular bone and bone marrow. This method employs a two-step purification strategy, combining iodixanol density cushion (IDC) ultracentrifugation with size exclusion chromatography (SEC), and enables EV recovery from fresh tissue hours after collection. Importantly, this approach facilitates the enrichment of bone-derived EVs without the need for enzymatic digestion or long-term culture, preserving native EV populations. This protocol offers a valuable tool for researchers investigating EVs derived from the diverse cellular constituents of the bone microenvironment.

cell biology↗

bioRxiv2024

Mechanisms of PP2A-Ankle2 dependent nuclear reassembly after mitosis

In animals, mitosis involves the breakdown of the nucleus. The reassembly of a nucleus after mitosis requires the reformation of the nuclear envelope around a single mass of chromosomes. This process requires Ankle2 (also known as LEM4 in humans) which interacts with PP2A and promotes the function of Barrier-to-Autointegration Factor (BAF). Upon dephosphorylation, BAF dimers cross-bridge chromosomes and bind lamins and transmembrane proteins of the reassembling nuclear envelope. How Ankle2 functions in mitosis is incompletely understood. Using a combination of approaches in Drosophila, along with structural modeling, we provide several lines of evidence that suggest that Ankle2 is a regulatory subunit of PP2A, explaining how it promotes BAF dephosphorylation. In addition, we discovered that Ankle2 interacts with the endoplasmic reticulum protein Vap33, which is required for Ankle2 localization at the reassembling nuclear envelope during telophase. We identified the interaction sites of PP2A and Vap33 on Ankle2. Through genetic rescue experiments, we show that the Ankle2/PP2A interaction is essential for the function of Ankle2 in nuclear reassembly and that the Ankle2/Vap33 interaction also promotes this process. Our study sheds light on the molecular mechanisms of post-mitotic nuclear reassembly and suggests that the endoplasmic reticulum is not merely a source of membranes in the process, but also provides localized enzymatic activity.

cell biology↗

bioRxiv2024

Comprehensive Proteomic Analysis of HCoV-OC43 Virions and Virus-Modulated Extracellular Vesicles

Viruses are obligate parasites that depend on the cellular machinery for their propagation. Several viruses also incorporate cellular proteins that facilitate viral spread. Defining these cellular proteins is critical to decipher viral life cycles and delineate novel therapeutic strategies. While numerous studies have explored the importance of host proteins in coronavirus spread, information about their presence in mature virions is limited. In this study, we developed a protocol to highly enrich mature HCoV-OC43 virions and characterize them by proteomics. Recognizing that cells release extracellular vesicles whose content is modulated by viruses, and given our ability to separate virions from these vesicles, we also analyzed their protein content in both uninfected and infected cells. We uncovered 69 unique cellular proteins associated with virions including 31 high confidence hits. These proteins primarily regulate RNA metabolism, enzymatic activities, vesicular transport, cell adhesion, metabolite interconversion and translation. We further discovered that the virus had a profound impact on exosome composition, incorporating 47 novel cellular proteins (11 high confidence) and excluding 92 others (61 high confidence) in virus-associated extracellular vesicles compared to uninfected cells. Moreover, a dsiRNA screen revealed that 11 of 18 select targets significantly impacted viral yields, including proteins found in virions or extracellular vesicles. Overall, this study provides new and important insights into the incorporation of numerous host proteins into HCoV-OC43 virions, their biological significance and the ability of the virus to modulate extracellular vesicles. ImportanceIn recent years, coronaviruses have dominated global attention, making it crucial to develop methods to control them and prevent future pandemics. Besides viral proteins, host proteins play a significant role in viral propagation and offer potential therapeutic targets. Targeting host proteins is advantageous because they are less likely to mutate and develop resistance compared to viral proteins, a common issue with many antiviral treatments. In this study, we examined the protein content of the less virulent biosafety level 2 HCoV-OC43 virus as a stand-in for the more virulent SARS-CoV-2. Our findings reveal that several cellular proteins incorporated into the virion regulate viral spread. Additionally, we report that the virus extensively modulates the content of extracellular vesicles, enhancing viral dissemination. This underscores the critical interplay between the virus, host proteins, and extracellular vesicles.

microbiology↗

bioRxiv2024

O-GlcNAcylation of FOXK1 orchestrates the E2F pathway and promotes oncogenesis

Gene transcription is a highly regulated process, and deregulation of transcription factors activity underlies numerous pathologies including cancer. Albeit near four decades of studies have established that the E2F pathway is a core transcriptional network that govern cell division in multi-cellular organisms1,2, the molecular mechanisms that underlie the functions of E2F transcription factors remain incompletely understood. FOXK1 and FOXK2 transcription factors have recently emerged as important regulators of cell metabolism, autophagy and cell differentiation3-6. While both FOXK1 and FOXK2 interact with the histone H2AK119ub deubiquitinase BAP1 and possess many overlapping functions in normal biology, their specific functions as well as deregulation of their transcriptional activity in cancer is less clear and sometimes contradictory7-13. Here, we show that elevated expression of FOXK1, but not FOXK2, in primary normal cells promotes transcription of E2F target genes associated with increased proliferation and delayed entry into cellular senescence. FOXK1 expressing cells are highly prone to cellular transformation revealing important oncogenic properties of FOXK1 in tumor initiation. High expression of FOXK1 in patient tumors is also highly correlated with E2F gene expression. Mechanistically, we demonstrate that FOXK1, but not FOXK2, is specifically modified by O-GlcNAcylation. FOXK1 O-GlcNAcylation is modulated during the cell cycle with the highest levels occurring during the time of E2F pathway activation at G1/S. Moreover, loss of FOXK1 O-GlcNAcylation impairs FOXK1 ability to promote cell proliferation, cellular transformation and tumor growth. Mechanistically, expression of FOXK1 O-GlcNAcylation-defective mutants results in reduced recruitment of BAP1 to gene regulatory regions. This event is associated with a concomitant increase in the levels of histone H2AK119ub and a decrease in the levels of H3K4me1, resulting in a transcriptional repressive chromatin environment. Our results define an essential role of O-GlcNAcylation in modulating the functions of FOXK1 in controlling the cell cycle of normal and cancer cells through orchestration of the E2F pathway.

molecular biology↗

bioRxiv2024

Modeling gene-environment interactions in Parkinson's Disease: Helicobacter pylori infection of Pink1-/- mice induces CD8 T cell-dependent motor and cognitive dysfunction.

Parkinsons disease (PD) is a chronic neurodegenerative disorder characterized by progressive loss of motor function. Diagnosis occurs late: after motor symptom development downstream of the irreparable loss of a large proportion of the dopaminergic neurons in the substantia nigra of the brain. Understanding PD pathophysiology in its pre-motor prodromal phase is needed for earlier diagnosis and intervention. Genetic risk factors, environmental triggers, and dysregulated immunity have all been implicated in PD development. Here, we demonstrate in a mouse model deficient in the PD-associated gene Pink, that infection with the human PD-associated gastric bacterium Helicobacter pylori leads to development of motor and cognitive signs resembling prodromal features of PD. This was also associated with proliferation and activation of primary mitochondria-reactive CD8 T cells and infiltration of CD8 T cells into the brain. Development of the motor and cognitive phenotypes in the infected Pink1-/- mice was abrogated when CD8 T cells were depleted prior to infection. We anticipate that this new model, which integrates genetic PD susceptibility, a PD-relevant environmental trigger, and specific immune changes that are required for symptom development, will be a valuable tool for increasing our understanding of this complex disease.

immunology↗

bioRxiv2023

Building RNA Backbone in Constant Time by Numerical Approximation

We present a numerical approximation method designed to swiftly create RNA ribose conformations in constant time. The methods parameterization relies on the atomic coordinates of a given base along with its two neighboring phosphate groups. Such a parameterization suits three dimensional modeling engines that determine the RNA conformational search space using base operations instead of backbone sampling. These engines consider phosphate groups as part of the base rigid bodies. Reconstructing ribose conformations result in less than 1 [A] of RMSD (root-mean-square deviation) compared to original conformations derived from high-resolution X-ray crystallographic structures. By incorporating this ribose construction method into MC-Sym, a well-established RNA three dimensional modeling software, we streamline the modeling process into two phases. This enhances the search algorithms speed and improves model consistency and precision. Additionally, we employed the method to pinpoint 27 irregular ribose stereoisomers in high-resolution RNA X-ray crystal structures.

bioinformatics↗

bioRxiv2023

Transposable elements regulate thymus development and function

Transposable elements (TE) are repetitive sequences representing [~]45% of the human and mouse genomes and are highly expressed by medullary thymic epithelial cells (mTEC). In this study, we investigated the role of TEs on T-cell development in the thymus. We performed multi-omic analyses of TEs in human and mouse thymic cells to elucidate their role in T cell development. We report that TE expression in the human thymus is high and shows extensive age- and cell lineage-related variations. TE expression correlates with multiple transcription factors in all cell types of the human thymus. Two cell types express particularly broad TE repertoires: mTECs and plasmacytoid dendritic cells (pDC). In mTECs, transcriptomic data suggest that TEs interact with transcription factors essential for mTEC development and function (e.g., PAX1 and REL), and immunopeptidomic data showed that TEs generate MHC-I-associated peptides implicated in thymocyte education. Notably, AIRE, FEZF2, and CHD4 regulate small yet non-redundant sets of TEs in murine mTECs. Human thymic pDCs homogenously express large numbers of TEs that likely form dsRNA, which can activate innate immune receptors, potentially explaining why thymic pDCs constitutively secrete IFN LJ/{beta}. This study highlights the diversity of interactions between TEs and the adaptive immune system. TEs are genetic parasites, and the two thymic cell types most affected by TEs (mTEcs and pDCs) are essential to establishing central T-cell tolerance. Therefore, we propose that orchestrating TE expression in thymic cells is critical to prevent autoimmunity in vertebrates.

immunology↗

bioRxiv2023

SPAT: Surface Protein Annotation Tool

Given the particular attractivity of antibody-based immunotherapies, in vitro experimental approaches aiming to identify and quantify proteins directly located at the cell surface, such as the surfaceome, have been recently developed and improved. However, the "surface" enriched, yet noisy output obtained from available methods makes it challenging to accurately evaluate which proteins are more likely to be located at the surface of the plasma membrane and which are simple contaminants. To that purpose, we developed the in silico Surface Protein Annotation Tool (SPAT), which unifies established annotations to grade proteins according to the chance they have to be located at the cell surface. SPAT accuracy was tested using in-house acute myeloid leukemia data, as well as public datasets, and despite using publicly available annotations, showed good performances when compared to more complex surfaceome predictors. Given its simple input requirement, SPAT is easily usable for the annotation of any gene/protein lists. Its output, in addition to the "surface" score, provides additional annotations including a "secretion" flag, references to verified antibodies targeting annotated proteins, as well as expression data and protein levels in essential human organs, making it a user-friendly tool for the community.

bioinformatics↗

bioRxiv2023

LRRK2 regulates the activation of the unfolded protein response and antigen presentation in macrophages during inflammation

While the contribution of inflammation in the pathological process leading to Parkinsons disease (PD) is well established, a growing body of evidence supports a role for the long-lasting adaptive immune system in the disease. We showed that, in inflammatory conditions, the PD-proteins PINK1 and Parkin negatively regulate the presentation of mitochondrial antigens on MHC I molecules, a process referred to as MitAP (Mitochondrial Antigen Presentation). In the absence of PINK1, over-activation of this pathway in antigen-presenting cells (APCs) engages autoimmune mechanisms leading to the establishment of cytotoxic CD8+ T cells. Co-culture of dopaminergic neurons (DNs) with these T cells led to neuronal cell death, suggesting that the MitAP in DNs made them susceptible to T cell-mediated destruction. In the present study, we used a pharmacological and genetic approach to characterize the MitAP pathway at the molecular level. We showed that this antigen presentation pathway is induced in APCs in response to inflammatory signals through the sequential activation of TLR4, cGAS-STING and the Unfolded Protein Response (UPR). A "UPR motif" present on a STING cytoplasmic domain was shown to specifically activate the UPR sensor IRE1. Remarkably, the PD-related protein LRRK2, acted with STING upstream of the UPR to regulate the transition from innate to adaptive immunity, thereby identifying this PD-related protein as a key player in the immune response during inflammation.

immunology↗

bioRxiv2023

K29-linked unanchored polyubiquitin chains disrupt ribosome biogenesis and direct ribosomal proteins to the Intranuclear Quality control compartment (INQ)

Ribosome assembly requires precise coordination between the production and assembly of ribosomal components. Mutations in ribosomal proteins that inhibit the assembly process or ribosome function are often associated with Ribosomopathies, some of which are linked to defects in proteostasis. In this study, we examine the interplay between several yeast proteostasis enzymes, including deubiquitylases (DUBs), Ubp2 and Ubp14, and E3 ligases, Ufd4 and Hul5, and we explore their roles in the regulation of the cellular levels of K29-linked unanchored polyubiquitin (polyUb) chains. Accumulating K29-linked unanchored polyUb chains associate with maturing ribosomes to disrupt their assembly, activate the Ribosome assembly stress response (RASTR), and lead to the sequestration of ribosomal proteins at the Intranuclear Quality control compartment (INQ). These findings reveal the physiological relevance of INQ and provide insights into mechanisms of cellular toxicity associated with Ribosomopathies.

cell biology↗