Journal of Biological Chemistry最新文献

Prostaglandin H₂ (PGH₂) is formed from arachidonic acid by cyclooxygenases (COX) and metabolized by thromboxane (TXS) and prostacyclin synthases (PGIS), two self-sufficient cytochromes P450 (CYP). The related fungal linoleic acid (LA) biosynthetic route is catalyzed by di-heme proteins of five dioxygenases (DOX) fused to three akin CYP subfamilies of allene oxide (AOS), linoleate diol (LDS), and epoxy alcohol (EAS) synthases. AlphaFold2 predicted the 3D structures of the DOX-CYP domains with very high confidence. Superposition with the COX:LA enzyme complex indicated that the protein fold of central α-helices and the motifs of the substrate recognition sites (SRS) were conserved, which suggest evolution from an ancient peroxidase precursor. TXS, PGIS, and AOS catalyze homolytic scissions of oxygen-oxygen bonds and LDS/EAS heterolytic scissions. The SRS4 of LDS and EAS predicted an Asn residue at close distal axial position of the heme thiolate iron in analogy with PGIS and plant AOS, but a nonpolar in TXS and 8S/9S-AOS, and a polar (Thr) in 8R/9R-AOS. Replacements of amide residues in SRS4 of LDS shifted the position of intramolecular hydroxylation of 8R-hydroperoxy-LA and the heterolytic scission to towards homolytic. The self-sufficient CYP may catalyze homo- and heterolytic cleavage of hydroperoxides and the endoperoxide of PGH₂ by different mechanisms, but the presentation of the oxygen-oxygen bonds to the metal centers might be crucial. The AF2 models illustrate the structural, catalytical, and evolutionary relationships between COX and microbiological DOX with CYP companions in unprecedented details, which reveal multiple amino acids of potential catalytic significance for future research.

Mitochondrial AAA+ proteases, LONP1, ClpXP, YME1L (i-AAA), and the m-AAA complex, maintain protein quality and shape organelle function. Growing interest in these enzymes stems from their association with neurodegeneration, cardiomyopathy, metabolic disease, and cancer. Recent structural and biophysical work clarifies how ATP-driven conformational cycles enable substrate recognition, unfolding, translocation, and proteolysis, and how assembly state, subunit composition, and regulatory inputs tune activity. These insights help interpret patient variants and guide experiments that connect mechanism to phenotype. Here we review shared mechanistic principles across the four proteases, contrast their architectures and regulatory features, and relate these properties to substrate selection and disease mechanisms, with emphasis on evidence from structural, biochemical, and cellular studies. We also survey strategies to modulate function. Small molecules, exemplified by Dordaviprone (ONC201) which activate human ClpP, provide proof of concept, and emerging modalities such as engineered macromolecules, may offer the selectivity and localization required to correct disease mechanisms or exploit disease dependencies. By integrating mechanism, disease links, and modulation strategies, this review provides a framework for translating basic insight on mitochondrial AAA+ proteases into new tools and, ultimately, therapies.

Congenital CMV infection is the most common perinatal infection, affecting up to 0.5% of infants. This elicits long-term disabilities that include neuropsychiatric manifestations, such as intellectual disability, microcephaly. Despite its high prevalence, the underlying mechanism of how congenitally acquired CMV infection causes brain pathology remains unknown. Here we discovered the molecular interplay of key host (DISC1 and PML) and viral (IE1) proteins within the neural progenitor cells, which underlay an attenuated neural progenitor proliferation in congenital CMV infection. Abolishing the viral IE1 protein by delivering IE1-targeting CRISPR/Cas9 to fetal brain rescued this progenitor cell deficit, a key pathology in congenital CMV infection. A selective targeting to a viral-specific protein by the CRISPR/Cas9 system is minimal in off-target effects. We further observed that CMV-encoded IE1 protein interferes with host PML-DISC1 interaction, resulting in disturbance of the Notch pathway in vitro and in embryonic brains. Therefore, we believe that a pivotal role of IE1 in an attenuated neural progenitor proliferation in the developing cortex through its interfering with interaction between host DISC1 and PML proteins.

Sialic acid (Sia) is essential for human physiology and health, as emphasized by the range of human diseases that is linked to abnormalities in the Sia pathway. Sias are typically found at the outermost part of glycoconjugates that are involved in several biological processes, including cell adhesion and signaling. Sia metabolism is key to the production of CMP-Sia, the building block for sialylation, and is targeted as a therapeutic strategy to ameliorate the effects of abnormal sialylation in disease. Interestingly, patients with different genetic defects in Sia metabolism show contrasting clinical symptoms affecting different tissues. For example, neurological symptoms are dominant in some congenital disorders of glycosylation (CDGs) like NANS-CDG, while the brain is unaffected in GNE myopathy which presents with isolated muscle symptoms. This suggests that more complex tissue-specific regulatory mechanisms may exist. In this review, we discuss the biosynthetic and genetic pathways in Sia metabolism with a specific focus on its role in brain, muscle, and platelets in health and genetic disease. Moreover, this review presents an overview of the clinical symptoms and genetic spectrum for each genetic disease. Overall, the molecular an biochemical profiles are not fully understood in these patients and effective therapies are limited. Therefore, additional research should focus on unravelling metabolic mechanisms that could be targeted to develop novel therapeutic strategies.

Neuropathic pain is a chronic condition characterized by damage to and dysfunction of the peripheral or central nervous system. There are currently no effective treatment options available for neuropathic pain, and existing drugs often provide only temporary relief with potential side effects. Multilineage-differentiating stress-enduring (Muse) cells are characterized by high expansion potential, a stable phenotype and strong immunosuppression. These properties make them attractive candidates for therapeutics for neuropathic pain management. Muse cells from different species demonstrated analgesic potential by reversing chronic constriction injury model (CCI)-induced neuropathic pain. Protein profiling revealed a high degree of similarity between Muse cells and bone marrow stromal cells (BMSCs). The intrathecal injection of Muse cells effectively reduced neuropathic pain in various mouse models, resulting in better analgesic effects than the administration of equivalent low doses of BMSCs. Immunohistochemical analysis and qPCR revealed the ability of Muse cells to inhibit spinal cord neuroinflammation caused by spared nerve injury model (SNI). In addition, Transwell and ELISA revealed that Muse cells migrated through the injured dorsal root ganglion (DRG) via the CCR7-CCL21 chemotactic axis. In addition, the secretion of TGF-β and IL-10 by Muse cells was identified as the mechanism underlying the analgesic effect of Muse cells. The capacity of Muse cells to mitigate neuroinflammation and produce analgesic effects via the modulation of TGF-β and IL-10 underscores their potential as promising therapeutic approaches for the treatment of neuropathic pain.

O-linked N-acetylglucosamine (O-GlcNAc) is a monosaccharide modification occurring on serine or threonine residues of most eukaryotic proteins. Only two enzymes, O-GlcNAc transferase (OGT) and O-GlcNAc hydrolase (OGA), regulate the dynamic flux of O-GlcNAc modification, rendering it extremely responsive to nutrition and stress conditions. O-GlcNAcylation stands at the center of EMT, sensing nutrient and stress signals to direct the transcriptional and signaling programs that enable phenotypic plasticity, thereby establishing its fundamental role in fibrosis and tumor metastasis. EMT is an essential biological event that confers mesenchymal characteristics to epithelial cells, characterized by the suppression of E-cadherin, a key epithelial adhesion molecule, and the overexpression of N-cadherin, a mesenchymal cadherin that promotes motility, or Vimentin, a mesenchymal intermediate filament protein. This review covers recent insights on the multiple canonical and non-canonical roles of O-GlcNAc, presenting O-GlcNAc cycling as a significant post-translational mechanism involved in various aspects of EMT. Furthermore, we systematically examine the functional connections between O-GlcNAcylation and EMT, focusing on identifying key O-GlcNAcylated proteins that regulate EMT and evaluating the relative contributions of transcriptional and post-translational mechanisms mediated by this modification. A comprehensive understanding of the intricate molecular circuitry governing the interplay between O-GlcNAcylation and EMT will deepen our mechanistic insights into cellular plasticity and offer novel therapeutic avenues for combating metastasis and other EMT-associated pathologies.

Glycosylation is an evolutionarily conserved post-translational modification of most proteins that are either secreted from cells or remain embedded within membranes as transmembrane proteins. It controls protein stability, plasma half-life, intracellular trafficking and can contribute to the actual biological function of the protein. Protein glycosylation can be divided into N-linked glycosylation that refers to the linkage of an oligosaccharide to the amide nitrogen of an asparagine residue, O-glycosylation that describes attachment of an oligosaccharide to the hydroxyl oxygen of a serine or threonine residue, and C-mannosylation, a rare modification in which a mannose residue is bound to the indole of a tryptophan residue via a carbon-carbon linkage. In this review, we summarize current knowledge about C-mannosylation. We describe how C-mannosylation was initially discovered and on which types of proteins it usually occurs. We explain the operation of the C-mannosyltransferases, the enzymes that attach the mannose to the substrate proteins, and which conformations the C-mannose adopt. Furthermore, we summarize what is known so far about the influence of the C-mannosylation on the function of the actual protein. Our review highlights an often overlooked post-translational modification as important regulator of protein function.

Subcellular compartmentalization may be an effective way of controlling the abundance and activity of miRNAs in mammalian cells. Exploring the regulatory processes that control miRNA activity, we found that specific miRNAs are reversibly localized to the mitochondrial matrix in a context-dependent manner. Our data suggest a de novo role of mitochondria as miRNA storage site in mammalian cells. miR-122 is a key hepatic miRNA regulating metabolic processes in the mammalian liver. In this study, we observed increased mitochondrial targeting of miR-122 in amino acid-starved hepatic cells. Interestingly, when cells are refed with amino acids, mitochondrial miR-122 is relocalized to the cytosol and reused for translational repression. Moreover, this phenomenon is not limited to miR-122, as other mitochondrial miRNAs (mito-miRs) follow similar transient storage inside mitochondria in stressed cells. Bioinformatic analysis revealed that mitochondria-localized mito-miRs preferentially target mRNAs encoding crucial mitochondrial components related to apoptosis. Hence, hepatic cells regulate apoptosis pathways during the starvation-refeeding cycle by shuttling a specific set of miRNAs to and from mitochondria, thereby balancing cytosolic miRNA content. Stress response miRNA binder ELAVL1 or HuR protein was found to be both necessary and sufficient for transporting the mito-miRs to the mitochondrial matrix - a process also controlled by the interaction between mitochondria and the endoplasmic reticulum.

EphA2, a receptor tyrosine kinase, is overexpressed in various cancers. Its ligand-independent non-canonical signaling is pro-tumorigenic, and elevated EphA2 expression is associated with poor prognosis in patients. Although preclinical and clinical studies targeting EphA2 have been conducted as cancer therapeutics, its role in the DNA damage response remains elusive. This study examined the role of EphA2 in cell cycle progression in Adriamycin (ADR)-treated cells. ADR treatment transcriptionally upregulated EphA2 expression in a p53-independent manner. Suppression of EphA2 upregulation abrogated G2 arrest, as evidenced by reductions in both cyclin B1 accumulation and Wee1 inhibition-driven cell division. However, the 2N-G1 cell population remained low, with increased tetraploid cells. Time-lapse imaging revealed that tetraploid formation resulted from mitotic bypass rather than mitotic slippage or cytokinesis failure. EphA2 knockdown upregulated p21 expression together with p53, and p21 knockdown suppressed EphA2 knockdown-induced mitotic bypass. Monitoring fluorescence from a GFP fusion with the cyclin B1 destruction box demonstrated degradation in interphase without cell division, suggesting premature activation of APC/C^Cdh1 in interphase. Notably, p21 upregulation following EphA2 knockdown was observed specifically in cervical cancer cell lines. Finally, ADR-induced suppression of cell proliferation was further enhanced by EphA2 knockdown and partially reversed by p21 knockdown. In conclusion, EphA2 suppression induces p21-dependent mitotic bypass and tetraploidization, leading to reduced cell proliferation. EphA2 upregulation following DNA damage may be pro-tumorigenic by maintaining G2 arrest to keep DNA damage at tolerable levels. These findings provide a rationale for combining EphA2 inhibition with DNA-damaging agents in certain cancer types.

TET2 is an epigenetic modifier whose canonical activity leads to the removal of cytosine methylation in the genome, which in essence results in the activation of gene expression. This function is particularly well described in the context of hematopoiesis and its alterations that lead to leukemia. However, in recent years, it has become evident that the non-canonical functions of TET2 also play a vital role in its activity. Rather than depending on its catalytic activity, these functions arise from TET2 interactions with other epigenetic modifiers. This review summarizes the structure, regulation, and functions of TET2 in immune cells. We describe how TET2 controls gene expression at both the DNA and RNA levels. In addition, we discuss the role of TET2 in hematopoietic stem cell fate and in clonal hematopoiesis of indeterminate potential (CHIP). Finally, we highlight the impact of TET2 mutations on age-related inflammatory diseases, including cardiovascular and neurodegenerative disorders. Collectively, available evidence positions TET2 as a key integrator of epigenetic state and immune signaling, with context-dependent effects on inflammation and tissue homeostasis, and underscores the therapeutic potential of targeting TET2-dependent pathways in clonal hematopoiesis and inflammatory diseases.