ABSTRACT
NP105-113-B*07:02-specific CD8+ T cell responses are considered among the most dominant in SARS-CoV-2-infected individuals. We found strong association of this response with mild disease. Analysis of NP105-113-B*07:02-specific T cell clones and single-cell sequencing were performed concurrently, with functional avidity and antiviral efficacy assessed using an in vitro SARS-CoV-2 infection system, and were correlated with T cell receptor usage, transcriptome signature and disease severity (acute n = 77, convalescent n = 52). We demonstrated a beneficial association of NP105-113-B*07:02-specific T cells in COVID-19 disease progression, linked with expansion of T cell precursors, high functional avidity and antiviral effector function. Broad immune memory pools were narrowed postinfection but NP105-113-B*07:02-specific T cells were maintained 6 months after infection with preserved antiviral efficacy to the SARS-CoV-2 Victoria strain, as well as Alpha, Beta, Gamma and Delta variants. Our data show that NP105-113-B*07:02-specific T cell responses associate with mild disease and high antiviral efficacy, pointing to inclusion for future vaccine design.
Subject(s)
HLA-B7 Antigen/immunology , Immunodominant Epitopes/immunology , Nucleocapsid Proteins/immunology , SARS-CoV-2/immunology , T-Lymphocytes, Cytotoxic/immunology , Aged , Amino Acid Sequence , Antibodies, Viral/immunology , Antibody Affinity/immunology , COVID-19/immunology , COVID-19/pathology , Cell Line, Transformed , Female , Gene Expression Profiling , Humans , Immunologic Memory/immunology , Male , Middle Aged , Receptors, Antigen, T-Cell/immunology , Severity of Illness Index , Vaccinia virus/genetics , Vaccinia virus/immunology , Vaccinia virus/metabolismABSTRACT
Regulatory interactions mediated by transcription factors (TFs) make up complex networks that control cellular behavior. Fully understanding these gene regulatory networks (GRNs) offers greater insight into the consequences of disease-causing perturbations than can be achieved by studying single TF binding events in isolation. Chromosomal translocations of the lysine methyltransferase 2A (KMT2A) gene produce KMT2A fusion proteins such as KMT2A-AFF1 (previously MLL-AF4), causing poor prognosis acute lymphoblastic leukemias (ALLs) that sometimes relapse as acute myeloid leukemias (AMLs). KMT2A-AFF1 drives leukemogenesis through direct binding and inducing the aberrant overexpression of key genes, such as the anti-apoptotic factor BCL2 and the proto-oncogene MYC However, studying direct binding alone does not incorporate possible network-generated regulatory outputs, including the indirect induction of gene repression. To better understand the KMT2A-AFF1-driven regulatory landscape, we integrated ChIP-seq, patient RNA-seq, and CRISPR essentiality screens to generate a model GRN. This GRN identified several key transcription factors such as RUNX1 that regulate target genes downstream of KMT2A-AFF1 using feed-forward loop (FFL) and cascade motifs. A core set of nodes are present in both ALL and AML, and CRISPR screening revealed several factors that help mediate response to the drug venetoclax. Using our GRN, we then identified a KMT2A-AFF1:RUNX1 cascade that represses CASP9, as well as KMT2A-AFF1-driven FFLs that regulate BCL2 and MYC through combinatorial TF activity. This illustrates how our GRN can be used to better connect KMT2A-AFF1 behavior to downstream pathways that contribute to leukemogenesis, and potentially predict shifts in gene expression that mediate drug response.
ABSTRACT
Cyclic GMP-AMP (cGAMP) is an immunostimulatory molecule produced by cGAS that activates STING. cGAMP is an adjuvant when administered alongside antigens. cGAMP is also incorporated into enveloped virus particles during budding. Here, we investigate whether inclusion of cGAMP within viral vaccine vectors enhances their immunogenicity. We immunise mice with virus-like particles (VLPs) containing HIV-1 Gag and the vesicular stomatitis virus envelope glycoprotein G (VSV-G). cGAMP loading of VLPs augments CD4 and CD8 T-cell responses. It also increases VLP- and VSV-G-specific antibody titres in a STING-dependent manner and enhances virus neutralisation, accompanied by increased numbers of T follicular helper cells. Vaccination with cGAMP-loaded VLPs containing haemagglutinin induces high titres of influenza A virus neutralising antibodies and confers protection upon virus challenge. This requires cGAMP inclusion within VLPs and is achieved at markedly reduced cGAMP doses. Similarly, cGAMP loading of VLPs containing the SARS-CoV-2 Spike protein enhances Spike-specific antibody titres. cGAMP-loaded VLPs are thus an attractive platform for vaccination.
Subject(s)
COVID-19 , Influenza Vaccines , Vaccines, Virus-Like Particle , Animals , Humans , Mice , Nucleotides, Cyclic , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Vaccines, Virus-Like Particle/geneticsABSTRACT
This community-based participatory action research (CBPAR) study describes a method for evaluating an after-school resilience-focused intervention in a low-resource rural area of southern India. Communities Rising, a locally developed resilience and academic program, was evaluated in a cross-continent collaboration between a research team at a U.S. university and the local community. The CBPAR literature highlights the importance of cultural considerations, community considerations, and community participation in the research process. The present case study describes the CBPAR research process and considerations at every phase of the research project, providing a road map of how community engagement can strengthen research, empower the community, and provide valuable knowledge. This study was conducted in three phases that focused on inclusion of local voices in the development both of the resilience program and the evaluation data collection process. Youth surveyors were particularly key to the research process. Data on participant demographics, satisfaction with the program, and qualitative contributions are also provided. Strengths and limitations of this study process in a rural community are discussed.
Subject(s)
Community-Based Participatory Research/organization & administration , Resilience, Psychological , Schools/organization & administration , Stress, Psychological/psychology , Students/psychology , Child , Female , Humans , India , Male , Poverty , Program Development/methods , Program Evaluation , Rural Population , Social Class , Surveys and QuestionnairesABSTRACT
Here, we identify coiled-coil domain-containing protein 13 (Ccdc13) in a genome-wide RNA interference screen for regulators of genome stability. We establish that Ccdc13 is a newly identified centriolar satellite protein that interacts with PCM1, Cep290 and pericentrin and prevents the accumulation of DNA damage during mitotic transit. Depletion of Ccdc13 results in the loss of microtubule organisation in a manner similar to PCM1 and Cep290 depletion, although Ccdc13 is not required for satellite integrity. We show that microtubule regrowth is enhanced in Ccdc13-depleted cells, but slowed in cells that overexpress Ccdc13. Furthermore, in serum-starved cells, Ccdc13 localises to the basal body, is required for primary cilia formation and promotes the localisation of the ciliopathy protein BBS4 to both centriolar satellites and cilia. These data highlight the emerging link between DNA damage response factors, centriolar and peri-centriolar satellites and cilia-associated proteins and implicate Ccdc13 as a centriolar satellite protein that functions to promote both genome stability and cilia formation.
Subject(s)
Cell Cycle Proteins/physiology , Centrioles/metabolism , Cilia/metabolism , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Genomic Instability , HCT116 Cells , HEK293 Cells , HeLa Cells , Humans , TransfectionABSTRACT
The centrosome acts as a centre for microtubule organisation and plays crucial roles in cell polarity, migration, growth and division. Cep131 has recently been described as a basal body component essential for cilium formation, but its function in non-ciliogenic cells is unknown. We identified human Cep131 (also known as AZI1) in a screen for regulators of genome stability. We show that centrosomal localisation of Cep131 is cell-cycle-regulated and requires both an intact microtubule network and a functional dynein-dynactin transport system. Cep131 is recruited to centriolar satellites by PCM1, and localised to the centriolar core region by both pericentrin and Cep290. Depletion of Cep131 results in a reduction in proliferation rate, centriole amplification, an increased frequency of multipolar mitosis, chromosomal instability and an increase in post-mitotic DNA damage. These data therefore highlight the importance of human Cep131 for maintaining genomic integrity.
Subject(s)
Cell Cycle Proteins , Centrioles , Centrosome , Genomic Instability , Microtubule Proteins , Antigens, Neoplasm/metabolism , Autoantigens/metabolism , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Cell Line , Centrioles/genetics , Centrioles/ultrastructure , Centrosome/metabolism , Centrosome/ultrastructure , Chromosomal Instability , Cytoskeletal Proteins , Dynactin Complex , Dyneins/metabolism , Humans , Microtubule Proteins/genetics , Microtubule Proteins/metabolism , Microtubule-Associated Proteins/metabolism , Mitosis/genetics , Neoplasm Proteins/metabolismABSTRACT
Hematopoietic stem cells (HSCs) reconstitute multilineage human hematopoiesis after clinical bone marrow (BM) transplantation and are the cells of origin of some hematological malignancies. Although HSCs provide multilineage engraftment, individual murine HSCs are lineage biased and contribute unequally to blood cell lineages. Here, we performed high-throughput single-cell RNA sequencing in mice after xenograft with molecularly barcoded adult human BM HSCs. We demonstrated that human individual BM HSCs are also functionally and transcriptionally lineage biased. Specifically, we identified platelet-biased and multilineage human HSCs. Quantitative comparison of transcriptomes from single HSCs from young and aged BM showed that both the proportion of platelet-biased HSCs and their level of transcriptional platelet priming increase with age. Therefore, platelet-biased HSCs and their increased prevalence and transcriptional platelet priming during aging are conserved features of mammalian evolution.
Subject(s)
Blood Platelets , Hematopoietic Stem Cells , Animals , Hematopoietic Stem Cells/immunology , Hematopoietic Stem Cells/cytology , Humans , Mice , Blood Platelets/immunology , Aging/immunology , Cell Lineage/immunology , Biological Evolution , Single-Cell AnalysisABSTRACT
Immunomodulatory imide drugs (IMiDs) including thalidomide, lenalidomide, and pomalidomide, can be used to induce degradation of a protein of interest that is fused to a short zinc finger (ZF) degron motif. These IMiDs, however, also induce degradation of endogenous neosubstrates, including IKZF1 and IKZF3. To improve degradation selectivity, we took a bump-and-hole approach to design and screen bumped IMiD analogs against 8380 ZF mutants. This yielded a bumped IMiD analog that induces efficient degradation of a mutant ZF degron, while not affecting other cellular proteins, including IKZF1 and IKZF3. In proof-of-concept studies, this system was applied to induce efficient degradation of TRIM28, a disease-relevant protein with no known small molecule binders. We anticipate that this system will make a valuable addition to the current arsenal of degron systems for use in target validation.
ABSTRACT
Untreated psychosis in adolescents and young adults is associated with significant and progressive impairment. Early intervention to provide support and treatment for those at risk of psychosis is essential. Several early intervention models have been developed for those at-risk and those who are victims of a recent episode - including the Portland Identification and Early Referral model (PIER; McFarlane, 2001). This study extends previous work demonstrating a variety of positive treatment outcomes achieved by PIER in the context of a large-scale implementation across the state of Delaware. The sample included 108 youth and young adults who were either at risk for psychosis or had already experienced a first episode within the past two years. Participants received the PIER treatment model and were followed from baseline to six months after they were discharged from treatment. Researchers predicted that PIER participants would experience an increase in functioning and a decrease in positive psychosis symptoms. Change over time was examined through the lens of two analytic techniques: the Reliable Change Index (RCI) analyses and Growth Curve Modeling (GCM). Results show improvement on a number of outcomes over the course of the intervention as expected. Clinical implications, limitations, and suggestions for further research are discussed.
Subject(s)
Psychotic Disorders , Young Adult , Humans , Adolescent , Psychotic Disorders/therapy , Psychotic Disorders/diagnosis , Treatment Outcome , Patient Discharge , Early Medical Intervention/methodsABSTRACT
Mutations within viral epitopes can result in escape from T cells, but the contribution of mutations in flanking regions of epitopes in SARS-CoV-2 has not been investigated. Focusing on two SARS-CoV-2 nucleoprotein CD8+ epitopes, we investigated the contribution of these flanking mutations to proteasomal processing and T cell activation. We found decreased NP9-17-B*27:05 CD8+ T cell responses to the NP-Q7K mutation, likely due to a lack of efficient epitope production by the proteasome, suggesting immune escape caused by this mutation. In contrast, NP-P6L and NP-D103 N/Y mutations flanking the NP9-17-B*27:05 and NP105-113-B*07:02 epitopes, respectively, increased CD8+ T cell responses associated with enhanced epitope production by the proteasome. Our results provide evidence that SARS-CoV-2 mutations outside the epitope could have a significant impact on proteasomal processing, either contributing to T cell escape or enhancement that may be exploited for future vaccine design.
ABSTRACT
Most existing studies characterizing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific T cell responses are peptide based. This does not allow evaluation of whether tested peptides are processed and presented canonically. In this study, we use recombinant vaccinia virus (rVACV)-mediated expression of SARS-CoV-2 spike protein and SARS-CoV-2 infection of angiotensin-converting enzyme (ACE)-2-transduced B cell lines to evaluate overall T cell responses in a small cohort of recovered COVID-19 patients and uninfected donors vaccinated with ChAdOx1 nCoV-19. We show that rVACV expression of SARS-CoV-2 antigen can be used as an alternative to SARS-CoV-2 infection to evaluate T cell responses to naturally processed spike antigens. In addition, the rVACV system can be used to evaluate the cross-reactivity of memory T cells to variants of concern (VOCs) and to identify epitope escape mutants. Finally, our data show that both natural infection and vaccination could induce multi-functional T cell responses with overall T cell responses remaining despite the identification of escape mutations.
Subject(s)
COVID-19 , SARS-CoV-2 , Humans , ChAdOx1 nCoV-19 , Vaccination , Antibodies, ViralABSTRACT
Unlike most cell types, many cancer cells survive at low extracellular pH (pHe), a chemical signature of tumors. Genes that facilitate survival under acid stress are therefore potential targets for cancer therapies. We performed a genome-wide CRISPR-Cas9 cell viability screen at physiological and acidic conditions to systematically identify gene knockouts associated with pH-related fitness defects in colorectal cancer cells. Knockouts of genes involved in oxidative phosphorylation (NDUFS1) and iron-sulfur cluster biogenesis (IBA57, NFU1) grew well at physiological pHe, but underwent profound cell death under acidic conditions. We identified several small-molecule inhibitors of mitochondrial metabolism that can kill cancer cells at low pHe only. Xenografts established from NDUFS1-/- cells grew considerably slower than their wild-type controls, but growth could be stimulated with systemic bicarbonate therapy that lessens the tumoral acid stress. These findings raise the possibility of therapeutically targeting mitochondrial metabolism in combination with acid stress as a cancer treatment option.
Subject(s)
Neoplasms , Oxidative Phosphorylation , CRISPR-Cas Systems/genetics , Cell Survival/genetics , Humans , Hydrogen-Ion Concentration , Neoplasms/geneticsABSTRACT
OBJECTIVE: To examine behavioral observations of affiliation (ie, warmth versus hostility) and control (ie, dominance versus submissiveness) and prior divorce as predictors of coronary artery calcification (CAC) in older couples. In some but not all studies, marital disruption and low marital quality have been shown to confer risk of coronary artery disease (CAD). Inconsistencies might reflect limitations of self-reports of marital quality compared with behavioral observations. Also, aspects of marital quality related to CAD might differ for men and women. METHODS: Couples underwent computed tomography scans for CAC and marital assessments, including observations of laboratory-based disagreement. Participants were 154 couples (mean age, 63.5 years; mean length of marriage, 36.4 years) free of prior diagnosis of CAD. RESULTS: Controlling traditional risk factors, we found behavioral measures of affiliation (low warmth) accounted for 6.2% of variance in CAC for women, p < .01, but not for men. Controlling behavior (dominance) accounted for 6.0% of variance in CAC for men, p < .02, but not for women. Behavioral measures were related to self-reports of marital quality, but the latter were unrelated to CAC. History of divorce predicted CAC for men and women. CONCLUSIONS: History of divorce and behavioral--but not self-report--measures of marital quality were related to CAD, such that low warmth and high dominance conferred risk for women and men, respectively. Prior research might underestimate the role of marital quality in CAD by relying on global self-reports of this risk factor.
Subject(s)
Calcinosis/epidemiology , Coronary Artery Disease/epidemiology , Divorce , Interpersonal Relations , Marriage/psychology , Spouses/psychology , Aged , Calcinosis/diagnostic imaging , Calcinosis/psychology , Coronary Artery Disease/diagnostic imaging , Coronary Artery Disease/psychology , Cross-Sectional Studies , Dominance-Subordination , Female , Health Status , Hostility , Humans , Male , Middle Aged , Risk Factors , Self Report , Sex Factors , Tomography, X-Ray ComputedABSTRACT
Acute erythroid leukemia (AEL) commonly involves both myeloid and erythroid lineage transformation. However, the mutations that cause AEL and the cell(s) that sustain the bilineage leukemia phenotype remain unknown. We here show that combined biallelic Cebpa and Gata2 zinc finger-1 (ZnF1) mutations cooperatively induce bilineage AEL, and that the major leukemia-initiating cell (LIC) population has a neutrophil-monocyte progenitor (NMP) phenotype. In pre-leukemic NMPs Cebpa and Gata2 mutations synergize by increasing erythroid transcription factor (TF) expression and erythroid TF chromatin access, respectively, thereby installing ectopic erythroid potential. This erythroid-permissive chromatin conformation is retained in bilineage LICs. These results demonstrate that synergistic transcriptional and epigenetic reprogramming by leukemia-initiating mutations can generate neomorphic pre-leukemic progenitors, defining the lineage identity of the resulting leukemia.
Subject(s)
CCAAT-Enhancer-Binding Protein-alpha/genetics , Cell Lineage , Cell Transformation, Neoplastic/pathology , Erythroid Precursor Cells/pathology , GATA2 Transcription Factor/genetics , Leukemia, Erythroblastic, Acute/pathology , Mutation , Neutrophils/pathology , Aged , Alleles , Animals , Cell Differentiation , Cell Transformation, Neoplastic/genetics , Disease Models, Animal , Erythroid Precursor Cells/metabolism , Female , GATA1 Transcription Factor/genetics , Humans , Leukemia, Erythroblastic, Acute/genetics , Male , Mice , Mice, Inbred C57BL , Middle Aged , Neutrophils/metabolism , Zinc FingersABSTRACT
Marital strain confers risk of cardiovascular disease (CVD), perhaps though cardiovascular reactivity (CVR) to stressful marital interactions. CVR to marital stressors may differ between middle-age and older adults, and types of marital interactions that evoke CVR may also differ across these age groups, as relationship contexts and stressors differ with age. The authors examined cardiovascular responses to a marital conflict discussion and collaborative problem solving in 300 middle-aged and older married couples. Marital conflict evoked greater increases in blood pressure, cardiac output, and cardiac sympathetic activation than did collaboration. Older couples displayed smaller heart rate responses to conflict than did middle-aged couples but larger blood pressure responses to collaboration-especially in older men. These effects were maintained during a posttask recovery period. Women did not display greater CVR than men on any measure or in either interaction context, though they did display greater parasympathetic withdrawal. CVR to marital conflict could contribute to the association of marital strain with CVD for middle-aged and older men and women, but other age-related marital contexts (e.g., collaboration among older couples) may also contribute to this mechanism.
Subject(s)
Aging/physiology , Blood Pressure/physiology , Conflict, Psychological , Cooperative Behavior , Heart Rate/physiology , Marriage/psychology , Age Factors , Aged , Aging/psychology , Cross-Sectional Studies , Female , Humans , Interpersonal Relations , Male , Middle Aged , Problem Solving , Sex FactorsABSTRACT
Prior theory and research regarding age differences in marital interaction suggest that older couples display and experience more positivity and less negativity than middle-aged couples. However, studies of overt behavior in older couples are relatively rare and have emphasized disagreement, neglecting other important contexts for older couples such as collaboration during everyday problem solving. Further, the affiliation or communion dimension of social interaction (i.e., warmth vs. hostility) is commonly assessed but not the control or agency dimension (e.g., dominance vs. submissiveness). The present study examined affect, cognitive appraisals, and overt behavior during disagreement (i.e., discussing a current conflict) and collaboration (i.e., planning errands) in 300 middle-aged and older married couples. Older couples reported less negative affect during disagreement and rated spouses as warmer than did middle-aged couples. However, these effects were eliminated when older couples' greater marital satisfaction was controlled. For observed behavior, older couples displayed little evidence of greater positivity and reduced negativity-especially women. During collaboration, older couples displayed a unique blend of warmth and control, suggesting a greater focus on emotional and social concerns during problem solving.
Subject(s)
Affect , Aging/psychology , Conflict, Psychological , Cooperative Behavior , Marriage/psychology , Adult , Age Distribution , Age Factors , Aged , Cohort Studies , Female , Hostility , Humans , Interpersonal Relations , Male , Middle Aged , Models, Psychological , Personal Satisfaction , Problem Solving , Spouses/psychology , Task Performance and AnalysisABSTRACT
Motion-onset visually evoked potentials (mVEPs) are neural potentials that are time-locked to the onset of motion of evoking stimuli. Due to their visually elegant properties, mVEP stimuli may be suited to video game control given gaming's inherent demand on the users' visual attention and the requirement to process rapidly changing visual information. Here, we investigate mVEPs associated with five different stimuli to control the position of a car in a visually rich 3D racing game in a group of 15 BCI naïve teenagers and compared with 19 BCI naive adults. Results from an additional 14 BCI experienced adults were compared with BCI naïve adults. Our results demonstrate that the game control accuracy is related to the number of trials used to make a decision on the users' chosen button/stimulus (76%, 62%, and 35% for 5, 3, and 1 trials, respectively) and information transfer rate (ITR) (13.4, 13.9, and 6.6 bits per minute (BPM)), although, even though accuracy decreases when using three compared to the commonly used five trial repetitions, ITR is maintained. A Kruskal-Wallis test suggests that BCI naïve adults do not outperform BCI naïve teenagers in the 3D racing game in the first and seconds laps (p > 0.05), but do outperform in the third lap (p < 0.05). A comparison between BCI naïve and BCI experienced adults indicates BCI experienced adults do not perform better than BCI naïve adults (p > 0.05).
Subject(s)
Aging/psychology , Brain-Computer Interfaces , Evoked Potentials, Visual/physiology , Adolescent , Adult , Algorithms , Child , Computer Graphics , Electroencephalography , Female , Humans , Male , Motion Perception/physiology , Photic Stimulation , Psychomotor Performance , Video Games , Young AdultABSTRACT
OBJECTIVE: Aspects of negative affect and social behavior studied as risk factors for coronary heart disease are usually examined separately and through self-reports. Using structural models of these personality domains, we tested associations of self-reports and spouse ratings of anxiety, depressive symptoms, anger, affiliation and dominance with coronary artery disease (CAD). DESIGN: In 154 healthy older couples, the authors tested cross-sectional associations with CAD of three facets of negative affectivity and two dimensions of the Interpersonal Circumplex, (IPC) using scales derived from the NEO-PI-R. MAIN OUTCOME MEASURES: CAD was quantified as Agatston scores from CT scans of coronary artery calcification (CAC). RESULTS: Self-reports were generally unrelated to CAC, whereas spouse ratings were consistently associated, largely independent of potential confounds. When considered simultaneously, anxiety and anger were related to CAC but depression was not. When considered together, both dominance and (low) affiliation were related to CAC. CONCLUSIONS: Structural models of negative affectivity and social behavior can facilitate integrative study of psychosocial risk factors. Further, self-report measures of these traits might under-estimate related CHD risk.
Subject(s)
Affect , Coronary Artery Disease/epidemiology , Coronary Artery Disease/psychology , Health Status , Personality Assessment , Personality , Social Dominance , Spouses , Surveys and Questionnaires , Aged , Anger , Anxiety Disorders/epidemiology , Anxiety Disorders/psychology , Calcinosis/epidemiology , Calcinosis/pathology , Coronary Artery Disease/pathology , Depressive Disorder/epidemiology , Depressive Disorder/psychology , Female , Humans , Male , Marriage/psychology , Middle Aged , Observer Variation , Risk FactorsABSTRACT
Genomic damage can feature DNA-protein crosslinks whereby their acute accumulation is utilized to treat cancer and progressive accumulation causes neurodegeneration. This is typified by tyrosyl DNA phosphodiesterase 1 (TDP1), which repairs topoisomerase-mediated chromosomal breaks. Although TDP1 levels vary in multiple clinical settings, the mechanism underpinning this variation is unknown. We reveal that TDP1 is controlled by ubiquitylation and identify UCHL3 as the deubiquitylase that controls TDP1 proteostasis. Depletion of UCHL3 increases TDP1 ubiquitylation and turnover rate and sensitizes cells to TOP1 poisons. Overexpression of UCHL3, but not a catalytically inactive mutant, suppresses TDP1 ubiquitylation and turnover rate. TDP1 overexpression in the topoisomerase therapy-resistant rhabdomyosarcoma is driven by UCHL3 overexpression. In contrast, UCHL3 is downregulated in spinocerebellar ataxia with axonal neuropathy (SCAN1), causing elevated levels of TDP1 ubiquitylation and faster turnover rate. These data establish UCHL3 as a regulator of TDP1 proteostasis and, consequently, a fine-tuner of protein-linked DNA break repair.