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Insulin action on the skeleton is essential for bone development and whole-body energy metabolism, however a global view of signaling in this tissue is lacking. Furthermore, whether there are signaling differences that drive the gene-specific activation under insulin-resistant (IR) or ageing conditions is unknown. Here, we perform a phosphoproteomic analysis of insulin signaling in the bones of young, lean, insulin-sensitive versus old, obese, IR mice revealing a rewiring of phosphorylation. We target dysregulated phosphoproteins in a zebrafish functional genomic screen of bone development and mineralization revealing candidates important for skeletal formation. One of these is ALF Transcription Elongation Factor 4 (AFF4), the core scaffold of the Super Elongation Complex and we show that phosphorylation of S831 on AFF4 is an insulin-dependent substrate of P70S6K and attenuated in aged, IR bone. Phosphorylation of S831 is defective in IR osteoblasts and associated with reduced transcriptional elongation at discrete locations in the genome. Mechanistically, we show phosphorylation of S831 increases recruitment of chromatin remodelers, ENL/AF9 to crotonylated histone via the YEATS domain, and promotes gene-specific activation. Our analysis identifies regulators of insulin action on the skeleton, further uncovering a mechanism of IR via locus-specific changes in transcriptional elongation and gene activation. Insulin signaling plays a crucial role in coordinating skeletal development with whole‑body energy metabolism. Here, the authors use phosphoproteomics to show insulin-signaling rewiring in aged, insulin-resistant bone and identify defective phosphorylation of AFF4 as a key mechanism for regulating gene-specific transcriptional activation.

Many machine learning applications in bioinformatics currently rely on matching gene identities when analyzing input gene signatures and fail to take advantage of preexisting knowledge about gene functions. To further enable comparative analysis of OMICS datasets, including target deconvolution and mechanism of action studies, we develop an approach that represents gene signatures projected onto their biological functions, instead of their identities, similar to how the word2vec technique works in natural language processing. We develop the Functional Representation of Gene Signatures (FRoGS) approach by training a deep learning model and demonstrate that its application to the Broad Institute’s L1000 datasets results in more effective compound-target predictions than models based on gene identities alone. By integrating additional pharmacological activity data sources, FRoGS significantly increases the number of high-quality compound-target predictions relative to existing approaches, many of which are supported by in silico and/or experimental evidence. These results underscore the general utility of FRoGS in machine learning-based bioinformatics applications. Prediction networks pre-equipped with the knowledge of gene functions may help uncover new relationships among gene signatures acquired by large-scale OMICs studies on compounds, cell types, disease models, and patient cohorts. Large-scale OMICs investigations of biological systems can be used to predict functional relationships between compounds, genes and proteins. Here, the authors develop a deep learning-based approach that significantly increases the number of high-quality compound-target predictions relative to existing methods.

The human brain has enormously complex cellular diversity and connectivities fundamental to our neural functions, yet difficulties in interrogating individual neurons has impeded understanding of the underlying transcriptional landscape. We developed a scalable approach to sequence and quantify RNA molecules in isolated neuronal nuclei from a postmortem brain, generating 3227 sets of single-neuron data from six distinct regions of the cerebral cortex. Using an iterative clustering and classification approach, we identified 16 neuronal subtypes that were further annotated on the basis of known markers and cortical cytoarchitecture. These data demonstrate a robust and scalable method for identifying and categorizing single nuclear transcriptomes, revealing shared genes sufficient to distinguish previously unknown and orthologous neuronal subtypes as well as regional identity and transcriptomic heterogeneity within the human brain.

Skeletal muscle accounts for 30–40% of body weight and plays an indispensable role in maintaining movement and is also a central regulator of whole-body metabolism. As such, understanding the molecular mechanisms of skeletal muscle health and disease is vital. Proteomics has been revolutionized in recent years and provided new insights into skeletal muscle. In this review, we first highlight important considerations unique to the field which make skeletal muscle one of the most challenging tissues to analyse by mass spectrometry. We then highlight recent advances using the latest case studies and how this has allowed coverage of the skeletal muscle temporal, fibre type and stem cells proteome. We also discuss how exercise and metabolic dysfunction can remodel the muscle proteome. Finally, we discuss the future directions of the field and how they can be best leveraged to increase understanding of human biology.

Proteins have been used as building blocks to provide various supramolecular structures in efforts to develop nano-biomaterials possessing broad biological functionalities. A series of unique structures have been obtained from the engineering of hemoproteins which contain the iron porphyrin known as heme, as a prosthetic group. This work in developing assembling systems is extended using cytochrome b562, a small electron transfer hemoprotein engineered to include an externally-attached heme moiety. The engineered units, which form a one-dimensional assembly via interprotein heme–heme pocket interactions, are conjugated to an apo-form of hexameric tyrosine-coordinated hemoprotein (apoHTHP) to provide a branching unit promoting the assembly of a star-shaped structure. The incorporation of the heme moiety attached to the protein surface of cytochrome b562 into apoHTHP can be accelerated by elevating the reaction temperature to generate a new assembly. The formation of a new larger assembly structure was confirmed by size exclusion chromatography. The ratio of the heme-containing units in the assemblies was analyzed by UV-Vis spectroscopy and the population of protein units estimated from SDS PAGE suggests the presence of plausible star-shaped structures, which are supported by hydrodynamic diameter data obtained by dynamic light scattering.

The incorporation of disease-relevant targets into ternary complexes in a compound-dependent manner by utilizing an assisting chaperone has become a common modality as far as bifunctional ternary complex-forming compounds are concerned. In contrast, examples of ternary complexes formed by molecular glues are much rarer. Due to their lack of significant binary (independent) target affinity, their identification cannot yet be achieved by rational methods and is, therefore, much more challenging. However, it is precisely for that reason (given the associated advantages) that their systematic identification and application in drug discovery has recently attracted particular interest. In contrast to bifunctional ternary complex-forming compounds, molecular glues retrieve a significant part of their thermodynamic stability through newly induced chaperone–target or glue–target interactions that occur only in the ternary complex. These interactions lead to enhanced ligand binding—termed intrinsic cooperativity α—which can be retrieved via the apparent cooperativity either by monitoring ligand binding through the chaperone or through the target protein. In this publication, the advantage of measuring the apparent cooperativity (to determine the cooperativity α) by the weaker binding protein is discussed and illustrated using the example of ternary complexes between FKBP12, MAPRE1 and macrocyclic molecular glues derived from the rapamycin binding motif for FKBP12. Furthermore, the impact of the following three parameters on the apparent cooperativity is illustrated: (1) the concentration of the monitoring protein, (2) the excess of the counter protein, and (3) the affinity of the glue to the weaker binding protein in combination with the degree of intrinsic cooperativity α. From this, experimental conditions to determine the intrinsic cooperativity α with only one binding assay and without the need for a comprehensive mathematical model covering all simultaneous events under non-saturating conditions are highlighted. However, this framework requires a binding assay capable of measuring or at least estimating very weak binary affinities. If this is not possible for experimental reasons, but binding assays for both proteins are available within a normal bandwidth and the affinity to the stronger binding protein is not too high, it is discussed how the binding curve for the weaker binding protein in the presence of an excess of the weaker binding protein can be used to overcome the missing binary Kd for the weakly binding protein.

HAP2(GCS1) is a deeply conserved sperm protein that is essential for gamete fusion. Here we use complementation assays to define major functional regions of the Arabidopsis thaliana ortholog using HAP2(GCS1) variants with modifications to regions amino(N) and carboxy(C) to its single transmembrane domain. These quantitative in vivo complementation studies show that the N-terminal region tolerates exchange with a closely related sequence, but not with a more distantly related plant sequence. In contrast, a distantly related C-terminus is functional in Arabidopsis, indicating that the primary sequence of the C-terminus is not critical. However, mutations that neutralized the charge of the C-terminus impair HAP2(GCS1)-dependent gamete fusion. Our results provide data identifying the essential functional features of this highly conserved sperm fusion protein. They suggest that the N-terminus functions by interacting with female gamete-expressed proteins and that the positively charged C-terminus may function through electrostatic interactions with the sperm plasma membrane.

Background Transplant renal artery aneurysm (TRAA) is rare. TRAA that develops post transplantation consists of 0.10% of the vascular complications post renal transplant (Transplant Proc 41:1609-1614, 2009; Indian J Urol 29:42-47, 2013). Case presentation We report a case of TRAA in an asymptomatic young female. CT angiogram with detailed 3D reconstruction showed a 2.6 × 2.2 cm wide neck saccular TRAA arising from the anterior segmental branch of the graft renal artery (Figs. 2 and 3). A multidisciplinary team of interventional radiologists, vascular and urologist was involved for preoperative surgical planning and unique repair methods. Endovascular and percutaneous approaches were deemed not feasible, and an open in vivo approach with a saphenous vein graft was taken. Conclusion TRAA, albeit rare, is a complication that can occur post renal transplant. In-vivo surgical repair of TRAA is feasible with a multidisciplinary approach and careful preoperative planning. Saphenous vein graft is still a versatile graft and can be used as a conduit successfully.

Background and Objectives: Depression is a leading cause of disability globally, with treatment challenges including limited access, stigma, and poor adherence. Gamification, which applies game elements such as points, levels, and storytelling into non-game contexts, offers a promising strategy to enhance engagement and augment traditional treatments. Our research is the first study designed to explore the implementation of gamification within the Malaysian context. The objective was to explore the feasibility of implementation of gamification as an adjunctive treatment for adults with depression. Materials and Methods: Focus group discussions were held with five mental health professionals and ten patients diagnosed with moderate depression. The qualitative component assessed perceptions of gamified interventions, while quantitative measures evaluated participants’ depressive and anxiety symptomatology. Results: Three key themes were identified: (1) understanding of gamification as a treatment option, (2) factors influencing its acceptance, and (3) characteristics of a practical and feasible intervention. Clinicians saw potential in gamification to boost motivation, support psychoeducation, and encourage self-paced learning, but they expressed concerns about possible addiction, stigma, and the complexity of gameplay for some patients. Patients spoke of gaming as a source of comfort, escapism, and social connection. Acceptance was shaped by engaging storylines, intuitive design, balanced difficulty, therapist guidance, and clear safety measures. Both groups agreed that gamification should be used in conjunction with standard treatments, be culturally sensitive, and be presented as a meaningful therapeutic approach rather than merely as entertainment. Conclusions: Gamification emerges as an acceptable and feasible supplementary approach for managing depression in Malaysia. Its success depends on culturally sensitive design, robust clinical oversight, and seamless integration with existing care pathways. Future studies should investigate long-term outcomes and establish guidelines for the safe and effective implementation of this approach. We recommend targeted investment into culturally adapted gamified tools, including training, policy development, and collaboration with key stakeholders to realistically implement gamification as a mental health intervention in Malaysia.

Local markets are still an integral part of the food system in developing economies of tropical regions including Samoa. This small South Pacific country is largely dependent on the production of crops in village agriculture, where traditional markets play an important role in sustainability of food supply. Similarly as many small island developing economies, Samoa is currently facing several challenges such as food security and high dependence on food imports. Therefore, we decided to monitor the diversity of plant foods on Samoan local markets and their economic and geographic indicators through interviews with the vendors. Our results suggest that assortment and economical value of plant food products have potential to increase sustainable food security of the local population and support economic growth of the region. For example, underutilized crops available at local markets are prospective species for development of new food products with beneficial nutritional and health properties. Moreover, certain commodities (e.g., papaya, kava and Samoan cocoa) seem to be promising for export. In addition, our findings suggest that development of appropriate processing technologies and the optimization of the logistics of crop products sold at local markets can contribute to an increase in efficiency of the regional agricultural sector.