GNAO1-related neurodevelopmental disorders are caused by mutations in the GNAO1 gene encoding the major neuronal G protein, Gαo. GNAO1 encephalopathies manifest in a range of symptoms, including epilepsy, movement disorder, hypotonia, and developmental delay, affecting >400 patients worldwide to date. A growth in the number of diagnosed cases is expected due to the wider availability of whole genome sequencing. One of the most recurrent pathogenic variants causing GNAO1 encephalopathy is an intronic mutation c.724-8G>A, which results in an in-frame insertion of two amino acid residues, Pro-Gln, after Thr241: Gαo[T241_N242insPQ]. We previously performed in-depth profiling of Gαo[insPQ] using structural, biochemical, and cellular studies. Compared with the wild-type protein, Gαo[insPQ] exhibits faster GTP binding and decreased hydrolysis. Importantly, Gαo[insPQ] is deficient in interacting with regulator of G protein signaling (RGS), GTPase-activating proteins that deactivate Gαo. These defects render Gαo[insPQ] a constitutively active mutant loaded with GTP in the G protein signaling. Patients harboring Gαo[insPQ] variant are in urgent need of novel therapy as they are refractory to available medications. In the present study, we performed a high-throughput screening to find molecules that might suppress the constitutive GTP loading by Gαo[insPQ]. We used a high-diversity chemical library of 54080 compounds, identifying a novel compound, N-[5-(2-methylpropyl)-1,3,4-thiadiazol-2-yl]-1H-1,2,3-benzotriazole-5-carboxamide, that decreases the GTP binding rate of Gαo, likely acting as a competitive inhibitor with higher selectivity to the pathogenic protein. This small-molecule inhibitor of Gαo opens new opportunities to drug discovery towards Gαo-dependent pathologies.

Periplasmic binding proteins (PBPs) are attractive scaffolds for fluorescent biosensors because they undergo large ligand-induced conformational changes and exhibit high specificity. Despite their potential, generalizable design strategies have been hindered by an incomplete understanding of how labeling positions influence protein dynamics and signal transduction. Here, we established a systematic experimental framework using the lysine-arginine-ornithine binding protein (LAO) as a model to characterize PBP-based fluorescent biosensors. Seven positions classified as endosteric, peristeric, or allosteric relative to the binding site were labeled with monobromobimane (mBBr). Our characterization revealed position-dependent effects on quantum yields, fluorescence intensity changes, ligand-binding affinities, and thermal stability. Molecular dynamics simulations (MD) of wild-type and mBBr-labeled variants provided a mechanistic context for these experimental observations. Functional biosensor positions (D51C, D53C, K228C, Y230C, and E167C) maintained open-like conformations necessary for signal generation upon ligand encounter. In contrast, peristeric position A89C, located at the hinge region, spontaneously adopted a closed-like conformation even in the absence of ligand, resulting in an inverse fluorescence response and reduced affinity. These results demonstrate that while structural criteria can guide initial site selection, experimental validation remains essential, as static structures alone cannot predict the complex interactions among labeling position, dynamics, and biosensor function. The experimental and computational framework established here with LAO as a model system provides methodological principles that may inform biosensor engineering from other PBP scaffolds, while emphasizing that conformational dynamics must be considered alongside static structural criteria.

Quorum sensing (QS) is a density-dependent communication process that enables bacteria to coordinate group behaviors, including competence, biofilm formation, bacteriocin production, and virulence. Early observations of genetic transformation in Streptococcus pneumoniae and density-dependent bioluminescence in marine bacteria laid the foundation for the eventual unification of these phenomena under the QS framework. In streptococci, QS is mediated primarily by secreted peptide pheromones that are processed, exported, and sensed through either membrane-associated two-component signal transduction systems or intracellular RRNPP-family regulators. These signaling pathways control tightly regulated and often transient physiological states, most notably competence, through interconnected regulatory circuits such as ComABCDE and ComRS. Increasing evidence indicates that, in some streptococci, these systems do not operate in isolation but instead form integrated and, in some cases, noncanonical networks that respond to environmental and metabolic cues. The chemical accessibility of peptide signals has enabled detailed structure-activity relationship studies, revealing key determinants of receptor activation and specificity. In particular, systematic modification of competence-stimulating peptides (CSPs) and sigX-inducing peptides (XIPs) has led to the identification of residues critical for activity and enabled the development of peptide analogs that act as agonists or competitive antagonists of QS pathways. These studies have demonstrated that synthetic peptides can modulate QS-regulated phenotypes, including competence, and in some cases attenuate infection in vivo. Collectively, these advances establish streptococcal QS as a chemically tractable system and highlight peptide-based modulation as a strategy for probing and influencing bacterial communication.

CC2D1A is a multidomain scaffold protein implicated in transcriptional regulation and autosomal recessive non-syndromic intellectual disability (NSID), yet its molecular mechanism is still poorly understood due to a lack of structural information. Here, we present the crystal structure of the human CC2D1A₄₉₁-₈₁₀ fragment, encompassing the fourth DM14 domain, a coiled-coil region, and a C-terminal C2 domain. These elements form a compact, integrated architecture, with the C2 domain mediating symmetric dimerization through conserved electrostatic interactions. In addition, a unique antiparallel β1-β10 sheet connects the coiled-coil and C2 domains stabilizing the tertiary structure. Fluorescence polarization assays reveal micromolar DNA-binding affinity, likely mediated by the basic surface of the DM14 domain. Comparison with the Drosophila homolog Lgd highlights conserved topology with added structural features, offering insights into CC2D1A's vertebrate-specific functions and NSID-related mutations.

This article is being retracted from Bioscience Reports at the request of the Editor-in-Chief and the Editorial Board. This follows the receipt of a notification from a reader, alerting the Editorial Board to irregularities in the Western Blots of Figures 1B and C.Specifically, there are background details in Figure 1B that unexpectedly repeat, and a region of the CC column of Figure 1C has a smudged appearance. The Editorial Board also has concerns surrounding the authorship of the manuscript.The authors have been contacted with regards to the retraction, but they have not responded to the Journal's queries or the concerns raised. Given the extent of the issues raised, the Editorial Board stands by the decision to retract the article.

This article is being retracted from Bioscience Reports at the request of the Editor-in-Chief and the Editorial Board. This follows the receipt of a notification from a reader, alerting the Editorial Board to concerns over the appearance of the apoptosis scatterplots of Figures 4E and 6F. Concerns include a ‘halo’ of regularly spaced dots and central areas with an unexpected texture. The Editorial Board note additional concerns over the article, including a lack of detail into which antibodies were used along with no mention in the author contribution of who performed the experiments. The authors have been contacted with regards to the retraction and have not responded to the Journal's queries or the concerns raised. Given the extent of the issues raised, the Editorial Board stands by the decision to retract the article.

DExD-box (DDX) proteins are essential for RNA metabolism and are targets for treatment of cancers and neurodevelopmental disorders. The biochemical mechanisms of many DDX proteins remain unclear, including human DDX52. DDX52 is essential for cell survival and is an emerging biomarker for the onset of metastatic melanoma. In this work, we identified that human DDX52 is an ATP-dependent translocase with 3’–5’ polarity, which can unwind DNA duplexes and DNA/RNA hybrids. Further, DDX52 is a nucleic acid annealase, an activity that requires an N-terminal intrinsically disordered protein region. DDX52 becomes hyperactive at DNA annealing if DDX52 helicase activity is inactivated by mutagenesis. Using CRISPR-Cas9 genetic editing, we generated U2OS cell lines heterozygous for DDX52 (DDX52+/–), which exhibit growth defects and impaired cell migration, providing direct support for previous suggestions that DDX52 may promote cancer cell metastasis and C-myc regulation.

Myocardial stunning, characterized by transient post-ischemic contractile dysfunction despite the restoration of coronary blood flow, has been a pivotal subject of cardiovascular research. Initially perceived as a consequence of irreversible myocardial damage and fibrosis, the concept evolved in the 1970s when studies revealed that reperfusion could salvage ischemic myocardium, leading to therapies like thrombolysis, percutaneous coronary intervention, and coronary artery bypass grafting. The phenomenon of myocardial stunning was first detailed by Heyndrickx et al. and later termed by Braunwald and Kloner, challenging previous views by demonstrating that reperfusion can cause temporary yet reversible dysfunction without necrosis. Extensive research elucidated mechanisms involving reactive oxygen species (ROS), calcium overload, and impaired excitation–contraction coupling. Recent advances in proteomics and phospho-proteomics identified molecular changes linked to contractile dysfunction, extracellular matrix damage, and apoptosis. The role of epigenetics has also garnered attention for its potential to influence myocardial stunning and offer therapeutic avenues. This review comprehensively explores the historical and mechanistic landscape of myocardial stunning, recent molecular insights, and its clinical relevance. Future research directions emphasize advanced proteomic and phosphor-proteomic analyses, epigenetic mechanisms, clinical translation, non-invasive diagnostics, ROS role clarification, ischemia preconditioning impacts, and integrative systems biology. Addressing these areas will enhance our understanding and lead to improved therapeutic strategies for ischemic heart disease.

Targeting AXL receptor kinase with a highly selective antibody presents a promising approach for inhibiting AXL and potentially improving cancer treatment. An essential step in antibody optimisation is the mapping of paratope residues to epitope residues. In the present study, we identify the residues of tilvestamab, a function-blocking anti-AXL monoclonal antibody, that are essential for its binding to the extracellular domain of AXL. A single-chain variable fragment (scFv) fused to osmotically inducible protein Y (osmY) was designed to enable the secretion of soluble scFv-osmY mutants, which could be directly subjected to high-throughput biolayer interferometry screening for binding to the AXL Ig1 domain. Each complementarity-determining region residue of scFv was mutated to Ala, while additional mutations were made on the basis of predicted contribution to binding. We generated AlphaFold3 predictions for the scFv (tilvestamab)-AXL Ig1 complex to gain insights into the molecular interactions of the essential residues, as determined by the experimental data. Our study reveals that tilvestamab binds to the Ig1 domain of AXL, with twelve residues on scFv (tilvestamab) contributing most to binding, likely being situated at the binding interface. Glu2 near the N-terminus of AXL is essential for binding. The data give a structural view into the AXL-tilvestamab complex and allow for further optimisation of the binding interface.