FEBS Letters最新文献

The GPN-loop GTPase Npa3 plays a critical role in RNA polymerase II (RNAPII) assembly and nuclear import. We employed here the npa3ΔC mutant, which supports normal RNAPII localization and function, to investigate potential links between Npa3 and target of rapamycin complex I (TORC1) signaling. The npa3ΔC cells exhibited increased sensitivity to rapamycin, a synthetic sickness interaction with tor1Δ, and a delayed growth recovery rate from rapamycin-induced G1 arrest. Co-expression analysis identified LTV1, a gene involved in TORC1 signaling and ribosome nuclear export, as one of the top genes co-expressed with NPA3. Furthermore, overexpression of eukaryotic translation initiation factor 1A (eIF1A, TIF11) or regulator of heterotrimeric G-protein signaling (RGS2) restored growth in npa3ΔC cells under rapamycin treatment. Interestingly, RGS2 also rescued growth under hygromycin B stress. Our findings suggest a genetic interplay between Npa3 and TORC1.

Gastric cancer research has rapidly progressed due to interdisciplinary advances in stem cell biology and bioengineering. Gastric organoid models, particularly those derived from adult stem cells, have emerged as powerful tools that recapitulate the cellular complexity of the human stomach. This review highlights the development of various gastric organoid platforms, with a specific focus on the convergence of engineering strategies to overcome the limitations of conventional organoid systems. We explore how CRISPR-based functional genomics, matrix innovations, co-culture systems, microphysiological systems (MPS), and big data integration are collectively enhancing organoid models. Furthermore, we examine how artificial intelligence may refine the clinical relevance and precision of gastric organoid models. By assessing both current capabilities and future directions, this review offers a perspective on how gastric organoid systems may reflect human physiology more accurately and improve therapeutic outcomes.

The t(10;11)(p13;q14-21) PICALM::MLLT10 chromosomal translocation results in the production of the CALM-AF10 fusion oncoprotein and is a driver mutation in both acute myeloid and T-lymphoblastic leukemia. PICALM::MLLT10 translocated leukemia is primarily an epigenetically driven disease. Global hypomethylation results in genomic instability, while focal H3K79 hypermethylation at target genes induces cell proliferation and blocks differentiation. Nucleocytoplasmic shuttling of CALM-AF10 and its protein partners and impaired endocytosis at the plasma membrane further influence the leukemic phenotype. Leukemias characterized by PICALM::MLLT10 have historically been recognized to portend a poor prognosis; however, insights from larger patient cohorts provide refinement to the prognostic relevance of this chromosomal translocation, highlighting chemotherapy resistance in this leukemic subtype. In addition, a deeper biological understanding of the disease hints at potential therapeutic targets. This approach is demonstrated in the recent promising results achieved utilizing venetoclax, a BCL2 inhibitor, in patients with PICALM::MLLT10 acute leukemia. Herein, we provide updates on the pathophysiology, clinical presentation, prognosis, and treatment of PICALM::MLLT10 acute leukemia.

Fragment-based inhibitor design is an established and widely used approach in drug discovery pipelines. Despite several examples of drugs originating from this approach, the identification of fragments still suffers from issues with solubility, reactivity, cost and worldwide accessibility. Here, we design a low-cost minimal fragment library (LoCoFrag100) for crystallographic screening, with an average cLogP of 0.03 (median 0.23) and an average of £20/g for each compound, facilitating assembly in any laboratory. Formatted in a 10 × 10 matrix to minimize Tanimoto similarity in the 20 cocktails, we demonstrate its applicability on three structurally distinct enzymes involved in microbial cell wall synthesis. Hit rates range from 1 to 6% among these enzymes, with three fragments suggesting avenues for inhibitor exploration. Impact Statement LoCoFrag100 is a low-cost, easily accessible fragment library that enables rapid survey of target ligandability in any laboratory, providing evidence to prioritise targets for follow-up research.

Glycosaminoglycan assembly on proteoglycans involves a common tetrasaccharide linker that starts with xylose attached to a serine on the protein. Defective linker biosynthesis caused by a missense mutation of human UDP-xylose synthase (hUXS1) is associated with connective tissue disorders characterized by skeletal abnormality and short stature. The Ile181Asn variant of hUXS1 was reported as inactive in releasing UDP-xylose from UDP-glucuronic acid. Here, we show that Ile181Asn-hUXS1 exhibited catalytic properties similar to the wild-type enzyme but featured a significant decrease in stability, expressed in melting temperature lowered from 48.2 °C to 35.2 °C. At 37 °C, Ile181Asn-hUXS1 was ~10-fold less stable and more prone to precipitation than wild-type hUXS1. The loss of function in Ile181Asn-hUXS1 is thus explained by instability, consistent with molecular dynamics simulations predicting structural destabilization. Impact statement The Ile181Asn variant of human UDP-xylose synthase (hUXS1), associated with a short-stature genetic syndrome, has previously been reported as inactive. We show here with experiments and molecular simulations that hUXS1 malfunction arises from structural instability rather than from a catalytic defect.

The folate biosynthesis activity of the human microbiome provides reduced folate metabolites that are readily absorbed from the gastrointestinal (GI) tract. The bacterial folate biosynthesis enzyme dihydropteroate synthase (DHPS), which adds p-aminobenzoate (pABA) to an activated pterin precursor, is an important antibiotic target. Both the broad-spectrum p-aminobenzenesulfonamide antibiotics, and the drug p-aminosalicylate (PAS, 2-hydroxy-pABA) with high selectivity for Mycobacterium tuberculosis, are competitive DHPS substrates. The adducts formed from these drugs, DHP-sulfonamides (sulfapterins) and 2'-hydroxyfolate metabolites, respectively, have been reported to exhibit antifolate activity in studies of microorganisms. The presence of these DHP-adducts and their effects on the host organism are largely undetermined; however, their close structural relationship to dihydrofolate (DHF) suggests that they are likely to mediate some side effects reported for these antibiotics. Naturally occurring pABA analogs that probably function similar to DHPS-targeted antibiotics have been identified in carrots and bacteria. Impact statement pABA analogs represent an important class of antibiotics, that are converted into dihydrofolate analogs by organisms present in the human microbiome. These analogs may mediate reported side-effects associated with these antibiotics. Several naturally occurring pABA mimics have been identified that are likely to exhibit antibiotic activity.

The tumor suppressor p53 is normally maintained at low levels through MDM2-mediated degradation; however, this regulation becomes ineffective upon DNA damage, leading to p53 phosphorylation and accumulation. This study shows that E6-associated protein (E6AP) provides an alternative regulatory pathway during genotoxic stress. Unlike MDM2, E6AP can effectively decrease p53 levels in HepG2 cells exposed to DNA-damaging agents, such as etoposide. Additionally, E6AP specifically targets p53 phosphorylated at serine-15, promoting its proteasomal degradation, whereas MDM2 cannot. This phosphorylation-dependent regulation by E6AP helps maintain p53 at appropriate levels during mild DNA damage, preventing excessive accumulation that could threaten cell survival, while still allowing for necessary stress responses. Impact statement The mechanism by which p53 is negatively regulated under genotoxic stress is largely unknown. E6-associated protein (E6AP), unlike MDM2, downregulates p53 levels following exposure to etoposide. E6AP specifically targets p53 phosphorylated at Ser-15. This mechanism prevents excessive accumulation of p53 that could otherwise reach lethal levels.

RNA sequences with the potential to form G-quadruplexes (rGQs) are widespread but largely unfolded in cells by unknown mechanisms. rGQ folding status is a critical regulator of RNA splicing and translation. We show that rGQs can be unfolded by SR proteins, SR-related proteins, and other Arg-rich proteins, including SRSF1, SRSF3, SRSF9, U1-70K, and U2AF1. The length and composition of Arg-rich regions are key determinants of this activity: Arg residues are the primary drivers, while acidic residues attenuate the unfolding activity. To unfold ARPC2 rGQ, at least 13 Arg residues are required. Our findings identify Arg-rich proteins as previously unrecognized, helicase-independent regulators of rGQ structures, with potential broad impacts on RNA processing that merit further investigation.

The Dot/Icm type IV secretion system (T4SS) is essential for Legionella pneumophila infection, but its in situ architecture and mechanism remain incompletely understood. Using cryo-electron tomography, we performed subtomogram averaging and 3D classification to resolve structural heterogeneity within the complex. We identified multiple assembly states of the inner membrane complex, including a fully assembled form with a hexamer-of-dimers DotO ATPase and symmetry mismatches between subcomplexes. A composite in situ model revealed a central channel above the inner membrane, likely used for substrate secretion. Imaging of infected macrophages showed T4SSs tethered to host vacuoles and extracellular vesicle release, suggesting additional effector delivery routes. These findings provide insight into Dot/Icm T4SS structure and infection-related dynamics.

A newly developed noninvasive Oncomagnetic device (OMD) causes selective cytotoxicity in glioblastoma and diffuse intrinsic pontine glioma cells, providing a novel, non-toxic approach to anticancer therapy. Here, we report results in cultured glioma cells and in a syngeneic mouse model indicating that the immediate intracellular target mechanism of action of the spinning oscillating magnetic field (sOMF) produced by this device is reactive oxygen species-dependent persistent inhibition of mitochondrial complex I. Steps downstream of this mechanism involve the production of oxidative stress, DNA damage, G1 phase cell cycle arrest, and caspase-dependent apoptosis. We also show that sOMF does not produce these effects in normal human astrocytes and astroglial cells. These data provide a rationale for safe clinical use of OMD.