Biology subjects

Beck, D. B.

Publications and source records attributed to Beck, D. B..

5 recordsLinked to original sources

bioRxiv2025

UBA1 Mutations Drive RIPK1-Mediated Cell Death and Monocyte Dysfunction in VEXAS Syndrome

VEXAS syndrome is a severe adult-onset autoinflammatory disease caused by somatic mutations in UBA1 gene, disrupting cytoplasmic ubiquitin-activating enzyme E1 function in hematopoietic progenitors. The pathogenesis remains poorly understood, particularly how UBA1 mutations perturb myeloid function. Here, we combine a genetically engineered THP-1 monocytic model with ex vivo analyses of blood and tissue samples from VEXAS patients to investigate the consequences of the canonical UBA1M41V mutation. We show that UBA1-mutated monocytes exhibit TNF--induced cell death, characterized by RIPK1 phosphorylation, and MLKL-and caspase-8-mediated cell death. This is associated with defective transcriptional induction of NF-{kappa}B target genes and reduced cFLIP(L) expression in response to TNF-. Monocytes also display blunted cytokine responses to multiple Toll-like receptor (TLR) agonists despite preserved TLR expression, linked to an impaired NF-{kappa}B response. UBA1M41V-derived macrophages exhibit an inflammatory transcriptional profile and increased secretion of chemokines that promote monocyte recruitment. We demonstrate that these UBA1M41V macrophages display impaired efferocytosis due to lysosomal dysfunction. Together, these findings reveal a pathogenic axis in VEXAS syndrome linking UBA1 loss of function and defective ubiquitination to RIPK1-mediated inflammatory cell death, impaired antimicrobial signaling, and defective resolution mechanisms. Our study provides novel mechanistic insights into the myeloid dysfunction that drives inflammation and cytopenia in VEXAS and highlights the necroptosis and efferocytosis pathways as potential therapeutic targets. Key points1/ UBA1-mutated monocytes are susceptible to RIPK1 dependent cell death and display impaired NF-{kappa}B-mediated cytokine responses to TLR agonists. 2/ UBA1-mutated macrophages promote inflammation and chemokine-mediated monocyte recruitment, while exhibiting defective efferocytosis.

immunology↗

bioRxiv2024

Single-cell genotype-phenotype mapping identifies therapeutic vulnerabilities in VEXAS syndrome

Somatic evolution leads to the emergence of clonal diversity across tissues with broad implications for human health. A striking example of somatic evolution is the VEXAS (Vacuoles E1 enzyme X-linked Autoinflammatory Somatic) syndrome, caused by somatic UBA1 mutations in hematopoietic stem cells (HSCs), inducing treatment-refractory, systemic inflammation. However, the mechanisms that lead to survival and expansion of mutant HSCs are unknown, limiting the development of effective therapies. The lack of animal or cellular models of UBA1-mutant HSCs has hindered such mechanistic understanding, mandating analysis of primary human VEXAS samples, which harbor admixtures of wild-type and UBA1-mutant HSCs. To address these challenges, we applied single-cell multi-omics to comprehensively define mutant UBA1-induced transcriptome, chromatin accessibility and signaling pathway alterations in VEXAS patients, allowing for the direct comparison of mutant versus wild-type cells within the same environment. We confirmed the expected enrichment of UBA1M41V/T mutations in myeloid cells, and additionally discovered that these mutations were also prevalent in Natural Killer (NK) cells in VEXAS patients, providing new insights into disease phenotypes. Through mapping genotypes to molecular phenotypes, including transcriptome, chromatin accessibility, cell surface protein or intracellular protein profiles, in HSCs, we found that UBA1M41V/T-mutant cells showed an increased inflammation signature (interferon alpha and gamma response pathways), as well as activation of unfolded protein response (UPR) via pro-survival, but not pro-apoptotic, mediators of the PERK pathway, compared to UBA1 wild-type HSCs. Ex vivo validation experiments showed that inhibiting UBA1 in normal CD34+ or using UBA1-mutant HSCs led to PERK pathway up-regulation, increased myeloid differentiation and cell survival, which was reversed by PERK inhibition. Thus, we demonstrated that human VEXAS HSCs show cell-intrinsic inflammatory phenotypes and survive the proteomic stress caused by compromised ubiquitination through PERK-mediated activation of the UPR. Together, these analyses nominate PERK pathway inhibition as a potential new therapeutic strategy for eradicating the VEXAS-inducing clone, demonstrating the power of single-cell multi-omics profiling of primary patient sample to enable genotype-to-phenotype somatic mapping for the discovery of novel candidates for clinical intervention.

genomics↗

bioRxiv2023

Shared and Distinct Mechanisms of UBA1 Inactivation Across Different Diseases

Most cellular ubiquitin signaling is initiated by UBA1, which activates and transfers ubiquitin to tens of E2 enzymes. Clonally acquired UBA1 missense mutations cause an inflammatory-hematologic overlap disease called VEXAS (vacuoles, E1, X-linked, autoinflammatory, somatic) syndrome. Despite extensive clinical investigation into this lethal disease, little is known about the underlying molecular mechanisms. Here, by dissecting VEXAS-causing UBA1 mutations, we discovered that p.Met41 mutations alter cytoplasmic isoform expression, whereas other mutations reduce catalytic activity of nuclear and cytoplasmic isoforms by diverse mechanisms, including aberrant oxyester formation. Strikingly, non-p.Met41 mutations most prominently affect transthioesterification, revealing ubiquitin transfer to cytoplasmic E2 enzymes as a shared property of pathogenesis amongst different VEXAS syndrome genotypes. A similar E2 charging bottleneck exists in some lung cancer-associated UBA1 mutations, but not in spinal muscular atrophy-causing UBA1 mutations, which instead, render UBA1 thermolabile. Collectively, our results highlight the precision of conformational changes required for faithful ubiquitin transfer, define distinct and shared mechanisms of UBA1 inactivation in diverse diseases, and suggest that specific E1-E2 modules control different aspects of tissue differentiation and maintenance.

biochemistry↗

bioRxiv2022

A tissue-specific ubiquitin switch coordinates brain, craniofacial, and skin development

The molecular mechanisms that coordinate patterning of the embryonic ectoderm into spatially distinct lineages to form the nervous system, epidermis, and craniofacial structures are unclear. Here, biochemical disease-variant profiling reveals a posttranslational pathway that drives early ectodermal differentiation in the vertebrate head. The anteriorly expressed ubiquitin ligase CRL3-KLHL4 restricts signaling of the ubiquitous cytoskeletal regulator CDC42. The major substrate of CRL3-KLHL4 is the canonical CDC42 effector kinase PAK1 that monoubiquitylation switches into a CDC42 inhibitor. Loss of CRL3-KLHL4 or a disease-associated KLHL4 variant reduce PAK1 ubiquitylation causing overactivation of CDC42 signaling and defective ectodermal patterning and neurulation. Thus, tissue-specific, ubiquitin-dependent restriction of CDC42 signaling is essential for face, brain, and skin formation, demonstrating how cell-fate and morphometric changes are coordinated for faithful organ development.

developmental biology↗

bioRxiv2019

Delineation of the First Human Mendelian Disorder of the DNA Demethylation Machinery: TET3 Deficiency

Germline pathogenic variants in chromatin-modifying enzymes are a common cause of pediatric developmental disorders. These enzymes catalyze reactions that regulate epigenetic inheritance via histone post-translational modifications and DNA methylation. Cytosine methylation of DNA (5mC) is the quintessential epigenetic mark, yet no human Mendelian disorder of DNA demethylation has been delineated. Here, we describe in detail the first Mendelian disorder caused by disruption of DNA demethylation. TET3 is a methylcytosine dioxygenase that initiates DNA demethylation during early zygote formation, embryogenesis, and neuronal differentiation and is intolerant to haploinsufficiency in mice and humans. Here we identify and characterize 11 cases of human TET3 deficiency in 8 families with the common phenotypic features of intellectual disability/global developmental delay, hypotonia, autistic traits, movement disorders, growth abnormalities, and facial dysmorphism. Mono-allelic frameshift and nonsense variants in TET3 occur throughout the coding region. Mono-allelic and bi-allelic missense variants localize to conserved residues with all but one occurring within the catalytic domain and most displaying hypomorphic function in a catalytic activity assay. TET3 deficiency shows substantial phenotypic overlap with other Mendelian disorders of the epigenetic machinery, including intellectual disability and growth abnormalities, underscoring shared disease mechanisms.

genetics↗