The FEBS journal最新文献

The 6-kb linear repeat genome of the mitochondrion (mtDNA) of the malaria parasite is among the smallest known in nature, but is well-conserved in comparison with its apicoplast and nuclear genomes. Except for the presence of base excision repair (BER) and two double-strand break repair (DSBR) proteins in mitochondria, the mechanisms for preservation of mtDNA integrity during traversal of the parasite through different cell types and environments in the mosquito vector and mammalian host are not characterized. We identified two putative organellar exonucleases in Plasmodium falciparum, PfExo_mit1 and PfExo_mit2, with homologs present only within certain alveolates. Immunofluorescence localization and chromatin immunoprecipitation experiments using antibodies generated against recombinant proteins showed that they are localized to the mitochondrion. PfExo_mit1 and PfExo_mit2 demonstrated specificity for different DNA substrates; PfExo_mit1 cleaved ssDNA in both polarities, while PfExo_mit2 was a bipolar exonuclease on dsDNA with 3'-5' exonuclease activity on ssDNA. The mismatch repair (MMR) protein PfMutS, which carries an additional endonuclease domain, was localized in the mitochondria and interacted with PfExo_mit2 in pull-down assays. PfExo_mit2 also interacted with the mitochondria-targeted DSBR protein PfRad51, suggesting that it is a component of both MMR and DSBR pathways. When Exo_mit1 expression in the rodent parasite P. berghei was silenced in sporozoites via conditional mutagenesis, PbExo_mit1 conditional knockout sporozoites invaded hepatocytes and developed in the liver, but could not transition to the blood stage. PbExo_mit1 localized to the mitochondria in liver stages as well, indicating that its ssDNA exonuclease function in mtDNA processing in the liver impacted establishment of blood-stage infection.

The hyperthermophile Thermotoga maritima does not possess a typical branched-chain amino acid aminotransferase or aromatic amino acid aminotransferase, leaving the biosynthetic pathways of these amino acids unclear. In this study, we identified and characterized a novel branched-chain and aromatic amino acid aminotransferase (TM1131). We also characterized a histidinol-phosphate aminotransferase (TM1040) with reported aminotransferase activity toward aromatic amino acids. TM1131 exhibited broad substrate specificity and the highest activity toward branched-chain and aromatic amino acids as an amino donor and toward corresponding 2-oxoacids as an amino acceptor. TM1040 also showed broad substrate specificity, with the highest activity toward l-lysine and l-arginine as an amino donor, and toward 2-oxoacids corresponding to l-methionine, l-leucine, and l-phenylalanine. Additionally, we investigated the multifunctionality of these two enzymes to explore other potential amino acid metabolic activities. Intriguingly, TM1131 displayed aspartate 4-decarboxylase activity, albeit with lower catalytic efficiency than measured for aminotransferase activity. TM1131 is involved in the final step of the biosynthetic pathways of branched-chain and aromatic amino acids, to which TM1040 also likely contributes.

Leishmania donovani is a causative agent of the neglected tropical disease known as visceral leishmaniasis or Kala Azar. This disease is lethal when untreated, with more than 90 000 cases annually. Little is known about magnesium regulation in these parasites despite magnesium being the second most abundant intracellular cation and universally required for normal cell function. L. donovani contains two protein tyrosine phosphatase (PTP) proteins (PTP1 and PTP2) and a DUF21 protein (domain of unknown function 21), which are respectively homologous to mammalian PRL [phosphatases of regenerating liver; also known as protein tyrosine phosphatase type IVA (PTP4A)] and mammalian transmembrane protein CNNM (cyclin M family). In mammalian cells, the PRL and CNNM multiprotein complex has been shown to sense and modulate intracellular magnesium levels. Herein, we revealed that L. donovani PTP1 and DUF21 can also form a specific protein complex. Using CRISPR-Cas9 gene editing, four L. donovani knockouts, LdΔPTP1, LdΔPTP2, a double knockout termed LdΔPTP1/2, and LdΔDUF21 have been generated. Magnesium-dependent growth curves demonstrated that the LdΔPTP1/2 mutant could not survive in low magnesium and had a reduced level of survival in infected macrophages. In contrast, LdΔDUF21 is sensitive to high levels of magnesium and has an increased level of intracellular magnesium and an increased survival in macrophages compared to wild-type L. donovani. Taken together, these observations provide evidence that, similar to the PRL and CNNM proteins in mammalian cells, PTP and DUF21 homologs in L. donovani have the ability to complex and respond to environmental changes in magnesium.

The urgent need for sustainable agriculture places biological nitrogen fixation at the forefront of current biotechnological research. Plant growth-promoting rhizobacteria play crucial roles in agriculture by enhancing nutrient absorption, regulating hormonal balance, and providing reduced nitrogen to plants. Among these, diazotrophic bacteria, such as Azotobacter vinelandii, stand out for their ability to fix atmospheric nitrogen and release it in bioavailable forms. In this issue of The FEBS Journal, scientists mapped the interaction between NifL and NifA proteins, which regulate nitrogen fixation in A. vinelandii and many other Proteobacteria. This understanding will allow for engineering bacteria to enhance nitrogen delivery to plants by improving nitrogen fixation.

Escherichia coli HTH-type transcriptional dual regulator CecR belongs to TetR family regulators (TFRs), which regulate the expression of genes enabling bacteria to survive under stress conditions. Previous studies (Yamanaka et al., Microbiology 2016; 162: 1253-1264) showed that CecR senses the presence of antibiotics, cephalosporins and chloramphenicol, in the cell and activates the expression of a putative drug efflux pump. Although CecR is present in many pathogenic strains of Escherichia and Salmonella genera, this regulator is poorly characterized. Here, we report the first crystal structure of E. coli CecR. Each protomer of the CecR homodimer is composed of an N-terminal DNA-binding and a C-terminal ligand-binding domain. In addition to nine canonical TetR α-helices, CecR contains structural elements characteristic of TetR subfamily D. The ligand-binding cavity of CecR has a tunnel-like shape, not common in TFRs. Unexpectedly, the CecR-ligand-binding cavity contained polyethylene glycol (PEG) fragments, originating from crystallization solution, and suggesting a potential site for effector binding. Additionally, the affinity of CecR to various antibiotics was determined. The strongest interactions were observed for CecR and cefepime, a representative of the fourth-generation cephalosporins. Molecular docking of the analyzed antibiotics into the ligand-binding tunnel of CecR indicated the amino acid residues important for ligand recognition. The CecR structure reported here provides the first structural information on the ligand-binding cavity and ligand recognition by CecR. As CecR is an important regulator, widespread among pathogenic bacteria belonging to the Enterobacteriales order, the results of our study are an important contribution to the understanding of the CecR-related mechanisms underlying antimicrobial resistance.

Human spliceosome-associated factor 3, SART3, is a key factor in spliceosome recycling and engages with U6 small nuclear RNA (snRNA) to promote the formation of the U4/U6 small nuclear ribonucleoprotein complex. Unlike its counterpart U4/U6 snRNA-associated-splicing factor PRP24 (Prp24) from Saccharomyces cerevisiae, which uses four RNA recognition motifs (RRMs) for the U6 snRNA interaction, SART3 has two RRMs at its C terminus. Here, we demonstrate that SART3 binds U6 snRNA as a dimer, and four RRM subunits recognize the asymmetric bulge of U6 snRNA. SART3 RRMs adopt a tandem βαββαβ motif of the canonical RRM fold to interact with the U6 bulge region via a conserved electropositive surface. We identified the cognate U6 elements that specifically bind SART3 RRM1, which is distinct from the Prp24-U6 interactions in yeast. Our findings suggest a divergent RRM binding mechanism for U6 snRNA recognition during spliceosome assembly and recycling.

Increasing evidence supports a mechanistic role for B cells in the pathogenesis of the autoimmune disorder multiple sclerosis (MS). We previously documented that the MS risk gene ataxin-1 (ATXN1) modulates key B-cell functions and the severity of the MS disease model experimental autoimmune encephalomyelitis (EAE). ATXN1 encodes the polyglutamine protein ataxin-1, which works in the cell nucleus as a corepressor of gene transcription. However, considering the ubiquitous expression of the ataxin-1 protein and the limitations of global Atxn1-null mouse models, the exact contribution of ataxin-1 to B-cell functioning and the overall EAE phenotype is not completely understood. To fill this gap, here we employed CRISPR-mediated genomic editing to develop the first conditional-knockout mouse line lacking ataxin-1 in the B-cell compartment. Using this novel in vivo model, we demonstrated that ataxin-1 regulates B-cell proliferation and activation in a cell-autonomous fashion, and decreases the activation of T cells and monocytes through indirect mechanisms. We also found that depleting ataxin-1 in B cells affects cytokine and immunoglobulin release in response to encephalitogenic challenges, but it is insufficient to modify the trajectory and neuropathology of the EAE model. Altogether, these results pinpoint a complex regulatory role for ataxin-1 in autoimmune demyelination involving multiple cellular targets.

Tyrosine kinase inhibitors (TKIs), targeted therapeutic agents, have significantly improved survival outcomes in patients with chronic myeloid leukemia (CML). However, the emergence of drug resistance remains a challenge, with the underlying mechanisms still largely unknown. Deubiquitinating enzymes (DUBs) have been reported as potential targets in many human cancers. In this study, we identified ubiquitin-specific protease 8 (USP8) as a potential prognostic target for CML drug resistance through bioinformatics analysis and validation in clinical samples. Functionally, knockdown of USP8 inhibited proliferation and increased apoptosis and TKI sensitivity in various CML cell lines. Mechanistically, immunoprecipitation-mass spectrometry analysis and molecular docking demonstrated an interaction between USP8 and eukaryotic translation initiation factor 2 subunit alpha (EIF2S1). USP8 stabilizes EIF2S1 protein by inhibiting its K48-linked ubiquitination, thereby preventing its degradation via the proteasome pathway. In other words, USP8 knockdown suppressed EIF2S1 protein expression and inhibited tumor growth both in vitro and in vivo, further suppressing TKI resistance. In summary, our results suggest that USP8 is overexpressed in CML and linked to resistance to TKIs. We have unveiled a previously unknown mechanism of CML drug resistance, which may provide novel perspectives on the advancement of targeted therapeutic strategies and clinical interventions.

Karen Vousden is an internationally renowned scientist who has made seminal contributions to p53 biology and cancer metabolism. She is currently a group leader at the Francis Crick Institute in London, where she heads the 'Tumour and host metabolism' laboratory. Her group's work investigates how metabolic changes impact cancer development and progression. Karen has had an illustrious and dynamic career. After completing postdoctoral fellowships at the Institute of Cancer Research and the National Cancer Institute (NCI) with Chris Marshall and Douglas Lowy, working on Ras and papillomaviruses respectively, she established her own research group at the Ludwig Institute in London. In 1995, she returned to the NCI, where she held prestigious leadership roles including Chief of the Regulation of Cell Growth Laboratory, before moving to Glasgow to take up the role of Director of the Cancer Research UK (CRUK) Beatson Institute in 2003. From 2016 to 2022, Karen also served as the Chief Scientist for CRUK. She was appointed Commander of the Order of the British Empire in 2010, Fellow of the Royal Society, and Foreign Associate of the US National Academy of Sciences in May 2018. Karen was awarded the Sir Hans Krebs medal at the 47th FEBS Congress in 2023 for her outstanding contributions to Biochemistry and Molecular Biology, and delivered a lecture on 'Diet, metabolism and cancer progression'. You can read her follow-up Review discussing the complex relationship between obesity, adipose tissue dysfunction, and tumour growth here [Solsona-Vilarrasa E & Vousden KH (2025) FEBS J 292, 2189-2207]. In this interview, we discuss Karen's research and her views on how scientists can help combat misinformation in science. We also talk about the innovative nutrition-based therapy currently being trialled by Faeth Therapeutics, a company founded by Karen and her colleagues.

Excessive dietary fructose consumption is a significant contributor to the development of metabolic dysfunction-associated steatotic liver disease (MASLD). However, the role of the intestinal fructose transporter, glucose transporter 5 (GLUT5), in this process remains poorly understood. This study aimed to investigate the potential use of GLUT5 as a novel therapeutic target for MASLD. Eight-week-old male C57BL/6J wild-type (WT) mice were fed a high-fat diet (HFD) or a high-fat diet supplemented with 10% or 25% fructose in drinking water (HFF) for 8 weeks to investigate the effects of high-fructose intake on MASLD development. To further elucidate the role of GLUT5, we utilized an intestine-specific GLUT5 knockout (Slc2a5-IKO) mouse model with HFF diet intervention to evaluate the protective effects of GLUT5 inhibition against high-fructose-induced liver injury. Additionally, the GLUT5 transporter inhibitor, 2,5-anhydro-D-mannitol (2,5-AM), was administered to HFF-fed WT mice to validate the alleviating effect on MASLD. The intestinal expression of GLUT5, liver and intestinal histology, and serum biochemical parameters were analyzed. Intestinal GLUT5 expression was significantly upregulated in WT mice fed HFF for 8 weeks, which developed impaired liver function and metabolic abnormalities. Compared with WT mice, liver damage was significantly attenuated in Slc2a5-IKO mice with the 8-week HFF diet plan. Furthermore, we demonstrated that the GLUT5 inhibitor, 2,5-AM, effectively suppressed intestinal GLUT5 expression and ameliorated the progression of MASLD in WT mice. Intestinal GLUT5 represents a potential therapeutic target for mitigating high-fructose-induced MASLD.