Biology subjects

Carroll, L.

Publications and source records attributed to Carroll, L..

9 recordsLinked to original sources

bioRxiv2026

BAF complex-independent gene activation by SS18::SSX

In synovial sarcoma, the BAF subunit SS18 is fused to SSX, a transcriptional repressor, generating the oncogenic SS18::SSX fusion protein. Incorporation of SS18::SSX into BAF complexes leads to their aberrant retargeting to Polycomb-repressed genes via SSX, while simultaneously altering their composition and activity. The presence of BAF at Polycomb target sites is widely assumed to be essential for gene activation. Here, we directly tested the requirement for BAF activity in synovial sarcoma cell survival and SS18::SSX-driven transcription. Using targeted degradation of BAF ATPase subunits and deletion of core components, we show that BAF loss has modest effects on sarcoma cell viability and does not impede SS18::SSX target gene expression. Consistently, deletion of the BAF ATPase subunit Smarca4 does not impair SS18::SSX-driven tumor formation in vivo. Using domain-specific SS18::SSX mutants, we further demonstrate that the fusion can activate oncogenic transcription independently of BAF interaction, and that this activity depends on the C-terminal QPGY-rich domain of SS18. Mechanistically, SS18::SSX promotes transcription by engaging the histone acetyltransferase EP300, independently of BAF. Accordingly, pharmacologic degradation of EP300/CREBBP suppresses SS18::SSX-driven transcriptional programs and impairs synovial sarcoma cell survival. Together, these findings challenge the view that BAF activity is required for SS18::SSX-mediated transcriptional activation and demonstrate that aberrant Polycomb target gene expression is sustained through recruitment of transcriptional coactivators in the absence of BAF. Our work reveals new therapeutic vulnerabilities in synovial sarcoma and suggests broader relevance for targeting coactivator-dependent transcription in fusion-driven cancers. HighlightsO_LIBAF degradation does not alter SS18::SSX-activated transcriptional programs C_LIO_LIDirect SS18::SSX transcriptional activation is independent of BAF interaction C_LIO_LIThe SS18 C-terminus engages the co-activator EP300 to promote gene expression C_LIO_LISmall molecule degraders of EP300/CREBBP abolish SS18::SSX-mediated transcription C_LI

cancer biology↗

bioRxiv2024

Therapeutic potential of red blood cell-derived extracellular vesicles in reducing neuroinflammation and protecting against retinal degeneration.

Neuroinflammation is a pathological process mediated through immune cell activation and pro-inflammatory cytokine release, resulting in neuronal cell death. In the central nervous system (CNS), neuroinflammation is a characteristic feature underlying the onset and progression of retinal and neurodegenerative diseases. Targeting neuroinflammation to reduce neuronal cell death and protect against visual and cognitive declines is therefore a key therapeutic strategy. However, due to the complex and multi-faceted nature of these diseases, to date there has been little therapeutic success with single target approaches insufficient to tackle widespread and multi-pathway inflammatory cascades. Furthermore, as the retina and brain reside within immune-privileged environments, a major challenge in treating these diseases is producing and delivering a therapeutic that, in itself, does not exacerbate inflammation. Extracellular vesicles (EV), derived from red blood cells (RBC EV), present a promising solution to overcome these hurdles, due to their innate ability to cross blood-tissue barriers, biocompatible nature, and their broad anti-inflammatory properties to modulate complex neuroinflammatory pathways. This study therefore investigated the therapeutic potential of RBC EV in mediating neuroinflammation using an in-vivo photo-oxidative damage model of retinal degeneration as a model for CNS neuroinflammation. In this work, we developed a novel incubation pipeline using N1 medium supplement and superoxide dismutase (SOD) supplementation to promote the production of safe, neuroprotective, and anti-inflammatory RBC EV. Delivery of RBC EV in vivo, was shown to be safe with strong penetration across all retinal layers. Further, therapeutic administration of RBC EV via local intravitreal injection significantly reduced inflammation and cell death and preserved retinal function. Notably, strong safety and therapeutic efficacy was also demonstrated in the retina following systemic (intraperitoneal) administration, highlighting a potential game-changing approach for less-invasive therapeutic delivery to the CNS. Finally, multi-omic analyses and in vitro findings supported an anti-inflammatory mechanism-of-action, with RBC EV modulating pro-inflammatory cytokine release, including those known to be involved in the pathogenesis of retinal and neurodegenerative diseases. Taken together, these findings highlight the broad applicability of RBC EV in treating neuroinflammation in the CNS, presenting a scalable and effective treatment approach for these currently untreatable diseases.

neuroscience↗

bioRxiv2024

SS18::SSX redistributes BAF chromatin remodelers selectively to activate and repress transcription

Synovial sarcoma (SyS) is driven by the expression of a chromosomal translocation-generated SS18::SSX fusion oncoprotein1,2. This oncoprotein fuses the SSX carboxy terminal tail that binds ubiquitylated histone H2A (H2AK119ub)3-6 to the SS18 component of both canonical (CBAF) and non-canonical (GBAF, GLTSCR1-containing) BAF-family chromatin remodeling complexes7-11, wherein SS18::SSX reprograms the epigenetic landscape. Mice that express SS18::SSX develop tumors that faithfully recapitulate human SyS histopathologically and molecularly12-14, allowing dissection of transcriptional reprogramming in vivo15,16. Here, we show that SS18::SSX redistributes GBAF complexes broadly to promoters and distal enhancers decorated by H2AK119ub, which uncharacteristically lack the transcription-silencing trimethylation of H3K27 that otherwise accompanies H2AK119ub. Fusion-GBAF instead associates with H2AK119ub and transcription-enabling H3K4 trimethylation (promoters), monomethylation (distal enhancers), and H3K27 acetylation (both). CBAF with the fusion redistributes away from typical loci, avoids H2AK119ub-bearing regions, and instead narrowly flanks transcription start sites (TSSs), co-distributed with polybromo BAF (PBAF). Accelerated tumorigenesis in our SyS mouse model followed deletion of Smarcb1 (PBAF and CBAF), Pbrm1 (PBAF-specific), and Arid1a or Arid1b (CBAF-specific). While tumors lacking Arid1a or Arid1b retained SyS character, loss of Smarcb1 or Pbrm1 resulted in tumors lacking SyS features. These findings suggest that SyS transcriptional reprogramming includes both improper GBAF localization and gene activation at H2AK119ub-bearing regulatory regions and improper gene silencing through loss of CBAF.

cancer biology↗

bioRxiv2023

Mitochondrial electron transport chain, ceramide and Coenzyme Q are linked in a pathway that drives insulin resistance in skeletal muscle

Insulin resistance (IR) is a complex metabolic disorder that underlies several human diseases, including type 2 diabetes and cardiovascular disease. Despite extensive research, the precise mechanisms underlying IR development remain poorly understood. Here, we provide new insights into the mechanistic connections between cellular alterations associated with IR, including increased ceramides, deficiency of coenzyme Q (CoQ), mitochondrial dysfunction, and oxidative stress. We demonstrate that elevated levels of ceramide in the mitochondria of skeletal muscle cells results in CoQ depletion and loss of mitochondrial respiratory chain components, leading to mitochondrial dysfunction and IR. Further, decreasing mitochondrial ceramide levels in vitro and in animal models (under chow and high fat diet) increased CoQ levels and was protective against IR. CoQ supplementation also rescued ceramide-associated IR. Examination of the mitochondrial proteome from human muscle biopsies revealed a strong correlation between the respirasome system and mitochondrial ceramide as key determinants of insulin sensitivity. Our findings highlight the mitochondrial Ceramide-CoQ-respiratory chain nexus as a potential foundation of an IR pathway that may also play a critical role in other conditions associated with ceramide accumulation and mitochondrial dysfunction, such as heart failure, cancer, and aging. These insights may have important clinical implications for the development of novel therapeutic strategies for the treatment of IR and related metabolic disorders.

cell biology↗

bioRxiv2023

Leveraging genetic diversity to identify small molecules that reverse mouse skeletal muscle insulin resistance

Systems genetics has begun to tackle the complexity of insulin resistance by capitalising on computational advances to study high-diversity populations. "Diversity Outbred in Australia (DOz)" is a population of genetically unique mice with profound metabolic heterogeneity. We leveraged this variance to explore skeletal muscles contribution to whole-body insulin action through metabolic phenotyping and skeletal muscle proteomics of 215 DOz mice. Linear modelling identified 553 proteins that associated with whole-body insulin sensitivity (Matsuda Index) including regulators of endocytosis and muscle proteostasis. To enrich for causality, we refined this network by focussing on negatively associated, genetically regulated proteins, resulting in a 76-protein fingerprint of insulin resistance. We sought to perturb this network and restore insulin action with small molecules by integrating the Broad Institute Connectivity Map platform and in vitro assays of insulin action using the Prestwick chemical library. These complimentary approaches identified the antibiotic thiostrepton as an insulin resistance reversal agent. Subsequent validation in ex vivo insulin resistant mouse muscle, and palmitate induced insulin resistant myotubes demonstrated potent insulin action restoration, potentially via up-regulation of glycolysis. This work demonstrates the value of a drug-centric framework to validate systems level analysis by identifying potential therapeutics for insulin resistance.

systems biology↗

bioRxiv2022

Genetic Architecture of Heart Mitochondrial Proteome influencing Cardiac Hypertrophy

Mitochondria play a key role in the normal function of the heart as well as in the pathogenesis of diseases. We report analysis of common genetic variations contributing to mitochondrial and heart functions using an integrative proteomics approach in a panel of inbred mouse strains called the Hybrid Mouse Diversity Panel (HMDP). We performed a whole heart proteomic analysis in the HMDP (72 strains, n=2-3 mice) and retrieved 840 mitochondrial proteins (quantified in [≥]50 strains). High-resolution association mapping on their respective abundance levels identified three trans-acting genetic loci, located on chromosome (chr) 7, chr13 and chr17, that control distinct classes of mitochondrial proteins as well as heart hypertrophy. Follow-up high resolution regional mapping identified NDUFS4, LRPPRC and COQ7 as the candidate genes for chr13, chr17 and chr7 loci, respectively, and both experimental and statistical analyses supported their causal roles. Variations of all three were associated with heart mass in two independent heart stress models, namely, isoproterenol (ISO)-induced heart failure and diet-induced obesity (DIO) models. To identify the aspects of mitochondrial metabolism regulated by these loci, we constructed co-expression protein networks using weighted gene co-expression network analysis (WGCNA). DAVID enrichment analyses of genes regulated by each of the loci revealed that the chr13 locus was highly enriched for complex-I proteins (24 proteins, P = 2.2E-61), the chr17 locus for mitochondrial ribonucleoprotein complex (17 proteins, P = 3.1E-25) and the chr7 locus for ubiquinone biosynthesis (3 proteins, P = 6.9E-05). These results indicate that common variations of certain mitochondrial proteins can act in trans to influence mitochondrial functions and contribute to heart hypertrophy, elucidating mechanisms that may underlie genetic susceptibility to heart failure in human populations.

genetics↗

bioRxiv2021

Uncharted biosynthetic potential of the ocean microbiome

Microbes are phylogenetically and metabolically diverse. Yet capturing this diversity, assigning functions to host organisms and exploring the biosynthetic potential in natural environments remains challenging. We reconstructed >25,000 draft genomes, including from >2,500 uncharacterized species, from globally-distributed ocean microbial communities, and combined them with [~]10,000 genomes from cultivated and single cells. Mining this resource revealed [~]40,000 putative biosynthetic gene clusters (BGCs), many from unknown phylogenetic groups. Among these, we discovered Candidatus Eudoremicrobiaceae as one of the most biosynthetically diverse microbes detected to date. Discrete transcriptional states structuring natural populations were associated with a potentially niche-partitioning role for BGC products. Together with the characterization of the first Eudoremicrobiaceae natural product, this study demonstrates how microbiomics enables prospecting for candidate bioactive compounds in underexplored microbes and environments.

microbiology↗

bioRxiv2020

Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy

A missense mutation of collagen type VIII alpha 2 chain (COL8A2) gene leads to early onset Fuchs endothelial corneal dystrophy (FECD), which progressively impairs vision through loss of corneal endothelial cells. We demonstrate that CRISPR/Cas9-based postnatal gene editing achieves structural and functional rescue in a mouse model of FECD. A single intraocular injection of an adenovirus encoding both the Cas9 gene and guide RNA (Ad-Cas9-Col8a2gRNA), efficiently knocked down mutant COL8A2 expression in corneal endothelial cells, prevented endothelial cell loss, and rescued corneal endothelium pumping function in adult Col8a2 mutant mice. There were no adverse sequelae on histology or electroretinography. Col8a2 start codon disruption represents a non-surgical strategy to prevent vision loss in early-onset FECD. As this demonstrates the ability of Ad-Cas9-gRNA to restore phenotype in adult post-mitotic cells, this method may be widely applicable to adult-onset diseases, even in tissues affected with disorders of non-reproducing cells.

genomics↗

bioRxiv2020

Loss of Corneal Raver2 Expression in Aniridia Associated Keratopathy

PurposeThe aim of this study was to evaluate downstream factors of Pax6 associated with the aniridic corneal phenotype. We characterize the ocular expression of Raver2, a novel heterogeneous ribonucleoprotein (hnRNP). MethodsGenome wide microarrays of corneal RNA from non-aniridic mice (C57BL/6J and MRL/MpJ) and corneas from aniridia model, Pax6+/- (SEY) mice were performed. Raver2 expression profiles were validated using qPCR and western blot. Raver2 immunohistochemistry on corneas of Pax6+/- mice and aniridic humans were compared to control specimens, and insitu-hybridization was used to analyze developmental expression of Raver2. ResultsMicroarray analysis of corneal RNA identified Raver2 as the gene with the strongest differential expression profile across non-aniridic and aniridic mouse strains. Consistent with mouse data, we found that Raver2 is expressed at high levels in healthy human corneal epithelium, and is downregulated in the corneas of human aniridia patients. ConclusionsThese findings suggest that Raver2 has an evolutionarily conserved developmental role downstream of Pax6 in corneal development and maintenance.

developmental biology↗