Centro Nacional de Biotecnología

Magnetic hyperthermia therapy (MHT) is a promising cancer treatment that has demonstrated efficacy in phase I and II clinical trials for glioblastoma and prostate cancer. MHT relies on heat generated by magnetic nanoparticles (MNPs) when exposed to alternating magnetic fields (AMFs). The heat output depends not only on the intrinsic properties of MNPs but also on extrinsic factors such as the extracellular and intracellular environments. Aggregation of MNPs under certain conditions can significantly reduce therapeutic efficiency. To overcome this limitation, we present a strategy to enhance MHT by modulating MNP-cell interactions. We functionalized dimercaptosuccinic acid (DMSA)-coated MNPs with the 92R antibody (DMSA-MNPs@92R), which selectively binds to the low-internalization chemokine receptor CCR9, overexpressed in certain tumors. Exposure of CCR9+ MOLT-4 cells to DMSA-MNPs@92R under AMFs resulted in enhanced tumor cell death. Our approach enables spatially controlled binding, maintaining MNPs in a less-aggregated state and at an optimal distance from the cell membrane to maximize heat generation. Mechanistic analysis confirmed that cytotoxicity is driven by localized hyperthermia at the subcellular level rather than a macroscopic temperature increase. These findings underscore the potential of controlled MNPs-cell interactions to improve in vitro MHT performance and open an interesting avenue for enhancing therapeutic efficacy.

Progenitor proliferation during neurodevelopment requires tight coordination of epigenetic regulation and metabolism. However, the crosstalk between these processes remains poorly understood. To investigate this, we examine in neural stem cells the role of PHF8, a histone demethylase whose mutations are linked to Siderius-Hamel syndrome, a rare neurodevelopmental disorder. Through an integrated multi-omics approach - combining transcriptomics, epigenomics, and metabolomics - we identify PHF8 as a key driver of the serine biosynthesis pathway, safeguarding the intracellular serine pool essential for neural progenitor proliferation. PHF8 fine-tunes chromatin accessibility at promoters of metabolic genes, ensuring their activation during development. Loss of PHF8 disrupts amino acid metabolism, blocks autophagy, and hinders vesicle formation. Ultimately PHF8 depletion leads to replication defects, DNA damage, and proliferation arrest. In vivo, PHF8 deficiency in mouse embryos halts neurogenesis, progenitor expansion, and neuron generation in the developing brain. These findings identify PHF8 as a key molecular link between chromatin regulation, metabolic control, and neural development, offering new insights into the epigenetic basis of neurodevelopmental and metabolic disorders.

The engineering of genetic circuits to perform predefined computations is central to synthetic biology, enabling living cells with new functionalities applicable across various domains. However, these circuits are often specifically tailored to particular cellular hosts, with Escherichia coli being the most popular. Consequently, their intended functions may not translate well to other organisms, limiting their scope. Understanding circuit dynamics in less familiar organisms is crucial, especially for niche-specific applications requiring cellular chassis different from model organisms generally used in synthetic biology. Here, we develop a combined experimental and theoretical pipeline to evaluate the performance of NOT logic circuits, also called inverters, in the soil bacterium Pseudomonas protegens Pf-5—a host renowned for its unique environmental functions and a newcomer to genetic circuitry. Inverters were experimentally tested to characterize input-output functionality, and mathematical modelling was used to infer the dynamic principles of circuit modules. The model quantified the individual impacts of key parameters—such as translation efficiency, repressor performance, and promoter activity—on output levels, enabling predictions about inter-host circuit portability. This parameter calibration revealed unique properties of the chassis, including steeper transitions between on and off circuit states compared to the synthetic biology workhorse Pseudomonas putida. Moreover, model parameters guided the successful serial connection of two NOT gates, enabling YES logic function. As a result, our work provides DNA parts, circuits and mathematical characterizations to establish P. protegens Pf-5 as a viable chassis for environmental synthetic biology.

Coupling between the chaperone and degradation systems, particularly under stress, is essential for eliminating unfolded proteins. The co-chaperone Bag1 links Hsp70 to the 26S proteasome, recruiting Hsp70-bound clients for proteasomal degradation. Here, we present cryo-electron microscopy structures of the Bag1-bound 26S proteasome, revealing unprecedented conformational rearrangements within the 19S regulatory particle. Bag1 binding to the Rpn1 induces a marked reconfiguration of AAA+ adenosine triphosphatase (ATPase) ring, disrupting its canonical spiral staircase and remodeling the central channel architecture. This reconfiguration generates a large cavity above the substrate entry gate of the 20S core particle. The conserved pore-2 loops of ATPases Rpt2 and Rpt5 play critical roles in opening of the 20S gate, enabling substrate entry into proteolytic chamber independently of ubiquitination. These findings suggest a previously unknown mechanism of the proteasomal degradation, by which remodeling the central cavity and 20S gate in the presence of Bag1, possibly bypassing the need for ubiquitination.

Cyclic-minipeptide candidates were computationally co-generated by deep-learning BoltzGen using home-designed Colab A100 notebooks against nonenveloped Nervous Necrosis Virus (NNV), an important worldwide expanded fish-farming pathogen. The Alphafold2-modeled capsid protein of grouper NNV was targeted by 6-, 13-and 21-amino acid sequences to co-generate CP // cyclic-minipeptide-conformers. The co-generated candidates facilitate recombinant/chemical synthesis, reduce physiological exoprotease degradation and conformer stability, increase environmental safety and would require improbable multiple-mutations to induce resistances. Despite exploring minimal percentages of the natural minipeptide wide-space, individually or in combination, these candidates constitute a novel proof-of-concept for experimentally-guided further explorations to control betanodavirus infections in real fish farming.

Background: Transmissible gastroenteritis virus (TGEV), a coronavirus (CoV) infecting pigs, uses its spike (S) glycoprotein to bind porcine aminopeptidase N (pAPN) for cell entry. Although structural studies have identified receptor-binding motifs (RBMs) within the receptor-binding domain (RBD) of the S protein, the functional relevance of individual residues for TGEV receptor recognition, cell entry, and infection remain unclear. Methods: In this study, we performed structure-guided mutagenesis of the TGEV RBD to evaluate the contribution of specific residues to receptor binding and viral infectivity. Results: Using soluble RBD proteins, we found that most of the RBD residues within the pAPN-binding interface contribute to the binding interaction. Nonetheless, TGEV reverse genetics experiments revealed that just three RBD residues (Gly527, Tyr528, and Trp571) were indispensable for viral cell entry. Mutations at these positions, which are conserved among group 1 alpha-CoVs abolished infectivity, highlighting their central role in the virus–receptor interface. Conclusions: Our findings provide a detailed functional map of the TGEV RBD and offer insights into the evolution of receptor recognition across CoV.