Biology subjects

Ayoub, N.

Publications and source records attributed to Ayoub, N..

8 recordsLinked to original sources

bioRxiv2026

LSm4 biomolecular condensates drive XRN2-mediated RNA decay at DNA double-strand breaks to facilitate repair

Maintenance of genome integrity requires accurate repair of DNA double-strand breaks (DSBs), particularly within transcriptionally active regions. Persistent R-loops at DSBs can impede homologous recombination (HR) repair. While factors that resolve R-loops at DSB sites are known, the mechanisms ensuring timely degradation of nascent RNA to prevent pathological R-loop accumulation remain elusive. Here, we identified a critical role for the RNA-binding protein LSm4 in orchestrating localized RNA decay at DSBs to facilitate repair. We demonstrated that among LSm1-8 subunits, only LSm4 undergoes liquid-liquid phase separation (LLPS) and forms biomolecular condensates (BCs) specifically at DSBs in transcriptionally active chromatin. These damage-induced LSm4 BCs function as hubs that promote nuclear RNA decapping and recruit the 5'[->]3' exonuclease XRN2 to degrade nascent transcripts proximal to DSBs. Accordingly, LSm4-XRN2 axis suppresses R-loop hyperaccumulation, thereby enabling efficient RAD51 filament assembly and intact HR repair. Consequently, loss of LSm4 increases translocations and leads to genomic instability. Collectively, our findings define a new regulatory layer in which LSm4 BCs spatially license RNA degradation, preventing R-loop accumulation at DSB microenvironment to facilitate error-free repair.

molecular biology↗

bioRxiv2026

Deciphering the nucleotide driven kinetic and oligomeric dynamics in IMPDH regulation

Inosine monophosphate dehydrogenase (IMPDH) controls guanine nucleotide biosynthesis. Here, using biochemical and integrative structural biology approaches, we characterize the class II bacterial IMPDH from Burkholderia thailandensis. We demonstrate that MgGTP acts as a direct allosteric inhibitor independently of MgATP, promoting tetramer-to-octamer assembly via the Bateman domain. When both nucleotides are present, the enzyme exhibits a biphasic response: low MgGTP concentrations enhance activity, whereas higher concentrations restore inhibition. Structural analyses reveal distinct octameric conformations and capture a pre-catalytic Michaelis complex with substrates and effectors bound. These findings uncover a regulatory mechanism where the Bateman domain integrates opposing nucleotide signals to dynamically control guanine nucleotide biosynthesis, highlighting IMPDH as a potential antimicrobial target in pathogenic Burkholderia species.

biochemistry↗

bioRxiv2025

Exploring the multi-protein assembly of the enzymes of the de novo purine nucleotide biosynthetic pathway from Pseudomonas aeruginosa

Purine nucleotide biosynthesis is a crucial metabolic pathway responsible that produces building blocks essential for a plethora of cellular processes. In bacteria, the de novo purine nucleotide biosynthetic pathway (DNPNB) involves fifteen chemical steps catalysed by fourteen different enzymes. While the mammalian orthologues have been extensively shown to interact and form a metabolon named "purinosome", the possible existence of a prokaryotic equivalent was only recently revealed for the case of Escherichia coli. In this study, we explored the potential conservation of a bacterial purinosome-like complex in Pseudomonas aeruginosa, an opportunistic pathogen known for its high antibiotic resistance. Using a bacterial two-hybrid system, we mapped protein-protein interactions among all tested DNPNB enzymes in P. aeruginosa and revealed a dense interaction network. An in-silico protein-protein docking approach on three core enzymes allowed the structural reconstitution of a complex composed of PurK, PurE and PurC with a 4:8:8 stoichiometry, respectively. Interestingly, a tunnel connecting the different active sites has been revealed, showing a metabolon-like property for possible efficient substrate channelling. These findings support a conserved regulatory organization of purine biosynthesis in bacteria, providing deeper insights into bacterial metabolism and paving the way for potential antibiotic targets.

biochemistry↗

bioRxiv2024

SLAYER: Synthetic Lethality Analysis for Enhanced Targeted Therapy Implicates AhR inhibitor as a Target in RB1-Mutant Bladder Tumors

Synthetic lethality represents a promising therapeutic approach in precision oncology, yet systematic identification of clinically relevant synthetic lethal interactions remains challenging. Here we present SLAYER (Synthetic Lethality AnalYsis for Enhanced taRgeted therapy), a computational framework that integrates cancer genomic data and genome-wide CRISPR knockout screens to identify potential synthetic lethal interactions. SLAYER employs parallel analytical approaches examining both direct mutation-dependency associations and pathway-mediated relationships across 808 cancer cell lines. Our integrative method identified 4,332 statistically significant interactions, which were refined to 142 high-confidence candidates through stringent filtering for effect size, druggability, and clinical prevalence. Systematic validation against protein interaction databases revealed a 15-fold enrichment of known associations among SLAYER predictions compared to random gene pairs. Through pathway-level analysis, we identified inhibition of the aryl hydrocarbon receptor (AhR) as potentially synthetically lethal with RB1 mutations in bladder cancer. Experimental studies demonstrated selective sensitivity to AhR inhibition in RB1-mutant versus wild-type bladder cancer cells, which probably operates through indirect pathway-mediated mechanisms rather than direct genetic interaction. In summary, by integrating mutation profiles, gene dependencies, and pathway relationships, our approach provides a resource for investigating genetic vulnerabilities across cancer types.

bioinformatics↗

bioRxiv2023

Targeting RBM10 deficiency in lung adenocarcinoma

The splicing factor RBM10 is frequently mutated in lung adenocarcinoma (LUAD) (9-25%). Most RBM10 cancer mutations are loss-of-function, correlating with increased tumorigenesis and limiting targeted therapy efficacy in EGFR-mutated lung cancer. Notably, therapeutic strategies leveraging RBM10 deficiency remain unexplored. Hence, we conducted RBM10 CRISPR-Cas9 synthetic lethality (SL) screen and identified [~]250 RBM10 SL genes, including WEE1 kinase. WEE1 inhibition sensitized RBM10-deficient LUAD cells in-vitro and in-vivo. Mechanistically, we identified a splicing-independent role of RBM10 in promoting replication fork progression that underpins RBM10-WEE1 SL. Also, we revealed that RBM10 is associated with active replication forks, which is reliant on PRIM1, an enzyme synthesizing RNA primers for Okazaki fragments. Functionally, we demonstrated that RBM10 serves as an anchor for recruiting HDAC1 and facilitates H4K16 deacetylation to maintain replication fork stability. Collectively, our data revealed a hitherto unrecognized function of RBM10 in fine-tuning DNA replication, and provide therapeutic arsenal for targeting RBM10-deficient tumors.

molecular biology↗

bioRxiv2021

Frizzled-4 regulates β-catenin in endothelial cells exposed to disturbed flow via an atypical Wnt pathway leading to proinflammatory activation and increased permeability

ObjectiveEndothelial cells are regulated by hemodynamic wall shear stress and multidirectional shear stress is known to promote endothelial dysfunction, although the molecular mechanisms are poorly defined. Wnt pathways play an important role in non-vascular mechanoresponsive cells. Here we investigated their role in endothelial mechanosignalling and endothelial dysfunction. Approach & ResultsHuman aortic endothelial cells were exposed to shear stress using an orbital shaker. The expression of Frizzled-4 receptors was significantly increased in endothelial cells exposed to low magnitude multidirectional flow (LMMF) relative to high magnitude uniaxial flow (HMUF). Increased expression was also detected in regions of the murine aortic arch exposed to LMMF. The increased Frizzled-4 expression in cultured cells was abrogated following knockdown of R-spondin-3 (RSPO-3) using RNA interference. LMMF also increased the stabilisation and nuclear localisation of {beta}-catenin, an effect that was dependent on Frizzled-4 and RSPO-3. Inhibition of {beta}-catenin using a small molecule inhibitor (iCRT5), or knockdown of Frizzled-4 or R-spondin-3 resulted in a significant reduction of pro-inflammatory gene expression in endothelial cells exposed to LMMF. Stabilisation of the {beta}-catenin destruction complex using IWR-1 under LMMF also reduced pro-inflammatory gene expression, as did inhibition of Wnt5a signalling. Interestingly, inhibition of the canonical Wnt pathway had no effect. Inhibition of {beta}-catenin signalling also reduced endothelial permeability; this was associated with altered junctional and focal adhesion organisation and cytoskeletal remodelling. ConclusionsThese data suggest the presence of an atypical Wnt-{beta}-catenin pathway in endothelial cells that promotes inflammatory activation and barrier disruption in response to LMMF.

cell biology↗

bioRxiv2021

Systematic identification of a CDYL1-dependent decrease in lysine crotonylation at DNA double-strand break sites functionally uncouples transcriptional silencing and repair

Previously, we showed that CDYL1 is recruited to DNA double-strand breaks (DSBs) to promote homology-directed repair (HDR) and foster transcriptional silencing. Yet, how CDYL1 elicits DSB-induced silencing is not fully understood. Here, we systematically identify a CDYL1-dependent local decrease in the transcriptionally active marks lysine crotonylation (PanKcr) and crotonylated histone residue H3K9cr at AsiSI-induced DSBs, which correlates with transcriptional silencing. Mechanistically, we reveal that CDYL1 crotonyl-CoA hydratase activity counteracts PanKcr and H3K9cr at AsiSI sites, which triggers the eviction of the transcriptional elongation factor ENL and foster transcriptional silencing. Furthermore, genetic inhibition of CDYL1 hydratase activity blocks the reduction in H3K9cr and alleviates DSB-induced silencing, while HDR efficiency unexpectedly remains intact. Therefore, our results functionally uncouple the repair and silencing activity of CDYL1 at DSBs. In a broader context, we address a long-standing question concerning the functional relationship between HDR and DSB-induced transcriptional silencing, suggesting that they may occur independently. HighlightsO_LISystematic identification of a local decrease in lysine crotonylation PanKcr and H3K9cr at AsiSI-induced DSBs that correlates with transcriptional silencing. C_LIO_LICDYL1 crotonyl-CoA hydratase activity downregulates Kcr at DSBs. C_LIO_LIKcr reduction at DSBs promotes ENL eviction and fosters transcriptional silencing. C_LIO_LICDYL1 roles in DSB-induced transcriptional silencing and HDR are functionally uncoupled. C_LI

molecular biology↗

bioRxiv2020

The common house spider, Parasteatoda tepidariorum, maintains silk gene expression on sub-optimal diet

Orb-web weaving spiders and their relatives spin multiple task-specific fiber types. The unique material properties of each silk type result from differences in amino acid sequence and structure of their component proteins, primarily spidroins (spider fibrous proteins). Amino acid content and gene expression measurements of spider silks suggest some spiders change expression patterns of individual protein components in response to environmental cues. We quantified mRNA abundance of three spidroin encoding genes involved in prey capture in the common house spider, Parasteatoda tepidariorum (Theridiidae), fed different diets. After 10 days of acclimation to the lab on a diet of mealworms, spiders were split into three groups: (1) individuals were immediately dissected, (2) spiders were fed high-energy crickets, or (3) spiders were fed low-energy flies, for 1 month. All spiders gained mass during the acclimation period and cricket-fed spiders continued to gain mass, while fly fed spiders either maintained or lost mass. Using quantitative PCR, we found no significant differences in the absolute or relative abundance of dragline gene transcripts, major ampullate spidroin 1 (MaSp1) and major ampullate spidroin 2 (MaSp2), among groups. In contrast, prey-wrapping minor ampullate spidroin (MiSp) gene transcripts were significantly less abundant in fly-fed than lab-acclimated spiders. However, when measured relative to actin, cricket-fed spiders showed the lowest expression of MiSp. Our results suggest that house spiders are able to maintain silk production, even in the face of a low-quality diet.

genetics↗