Biochemical Journal最新文献

The AMP-activated protein kinase (AMPK) is a sensor of cellular energy status activated by increases in AMP or ADP relative to ATP. Once activated, it phosphorylates targets that promote ATP-generating catabolic pathways or inhibit ATP-consuming anabolic pathways, helping to restore cellular energy balance. Analysis of human cancer genome studies reveals that the PRKAA2 gene (encoding the α2 isoform of the catalytic subunit) is often subject to mis-sense mutations in cancer, particularly in melanoma and non-melanoma skin cancers, where up to 70 mis-sense mutations have been documented, often accompanied by loss of the tumour suppressor NF1. Recently it has been reported that knockout of PRKAA2 in NF1-deficient melanoma cells promoted anchorage-independent growth in vitro, as well as growth as xenografts in immunodeficient mice in vivo, suggesting that AMPK-α2 can act as a tumour suppressor in that context. However, very few of the mis-sense mutations in PRKAA2 that occur in human skin cancer and melanoma have been tested to see whether they cause loss-of-function. We have addressed this by making most of the reported mutations and testing their activity when expressed in AMPK knockout cells. Of 55 different mis-sense mutations (representing 75 cases), 9 (12%) appeared to cause a total loss of activity, 18 (24%) a partial loss, 11 (15%) an increase in phenformin-stimulated kinase activity, while just 37 (49%) had no clear effect on kinase activity. This supports the idea that AMPK-α2 acts as a tumour suppressor in the context of human skin cancer.

Vaccinia-Related Kinase 2 (VRK2) is an anti-apoptotic Ser/Thr kinase that enhances drug sensitivity in cancer cells. This protein exists in two isoforms: VRK2A, the longer variant, and VRK2B, which lacks the C-terminal region and transmembrane domain. While the therapeutic importance of VRK2 family proteins is known, the specific roles of VRK2A and its interplay with apoptotic regulator Bcl-xL (B-cell lymphoma-extra Large) remain elusive. Bcl-xL regulates cell death by interacting with BAX (B-cell lymphoma-2 Associated X-protein), controlling its cellular localization and influencing BAX-associated processes and signaling pathways. As VRK2A interacts with the Bcl-xL-BAX complex, comprehending its regulatory engagement with Bcl-xL presents potential avenues for intervening in diseases. Using a multi-disciplinary approach, this study provides information on the cellular localization of VRK2A and establishes its interaction with Bcl-xL in the cellular milieu, pinpointing the interacting site and elucidating its anti-apoptotic property within the complex. Furthermore, this study also put forth a model that highlights the importance of VRK2A in stabilizing the ternary complex, formed with Bcl-xL and BAX, thereby impeding BAX dissociation and hence apoptosis. Therefore, further investigations associated with this important revelation will provide cues for designing cancer therapeutics in the future.

Anthropogenic emissions have caused atmospheric carbon dioxide (CO2) concentrations to double since the industrial revolution. Although this could benefit plant growth from the 'CO2 fertilisation' effect, recent studies report conflicting impacts of elevated CO2 (eCO2) on plant-pathogen interactions. Fungal pathogens are the leading cause of plant disease. Since climate change has been shown to affect the distribution and virulence of these pathogens, it is important to understand how their plant hosts may also respond. This review assesses existing reports of positive, negative, and neutral effects of eCO2 on plant immune responses to fungal pathogen infection. The interaction between eCO2 and immunity appears specific to individual pathosystems, dependent on environmental context and driven by the interactions between plant defence mechanisms, suggesting no universal effect can be predicted for the future. This research is vital for assessing how plants may become more at risk under climate change and could help to guide biotechnological efforts to enhance resistance in vulnerable species. Despite the importance of understanding the effects of eCO2 on plant immunity for protecting global food security, biodiversity, and forests in a changing climate, many plant-pathogen interactions are yet to be investigated. In addition, further research into the effects of eCO2 in combination with other environmental factors associated with climate change is needed. In this review, we highlight the risks of eCO2 to plants and point to the research required to address current unknowns.

Protein ubiquitylation typically involves isopeptide bond formation between the C-terminus of ubiquitin to the side-chain amino group on Lys residues. However, several ubiquitin ligases (E3s) have recently been identified that ubiquitylate proteins on non-Lys residues. For instance, HOIL-1 belongs to the RING-in-between RING (RBR) class of E3s and has an established role in Ser ubiquitylation. Given the homology between HOIL-1 and ARIH1, an RBR E3 that functions with the large superfamily of cullin-RING E3 ligases (CRLs), a biochemical investigation was undertaken, showing ARIH1 catalyzes Ser ubiquitylation to CRL-bound substrates. However, the efficiency of ubiquitylation was exquisitely dependent on the location and chemical environment of the Ser residue within the primary structure of the substrate. Comprehensive mutagenesis of the ARIH1 Rcat domain identified residues whose mutation severely impacted both oxyester and isopeptide bond formation at the preferred site for Ser ubiquitylation while only modestly affecting Lys ubiquitylation at the physiological site. The results reveal dual isopeptide and oxyester protein ubiquitylation activities of ARIH1 and set the stage for physiological investigations into this function of emerging importance.

Polycomb repressive complex 2 (PRC2) is central to polycomb repression as it trimethylates lysine 27 on histone H3 (H3K27me3). How PRC2 is recruited to its targets to deposit H3K27me3 remains an open question. Polycomb-like (PCL) proteins, a group of conserved PRC2 accessory proteins, can direct PRC2 to its targets. In this report, we demonstrate that a PCL protein named PHF1 forms phase-separated condensates at H3K27me3 loci that recruit PRC2. Combining cellular observation and biochemical reconstitution, we show that the N-terminal domains of PHF1 cooperatively mediate target recognition, the chromo-like domain recruits PRC2, and the intrinsically disordered region (IDR) drives phase separation. Moreover, we reveal that the condensates compartmentalize PRC2, DNA, and nucleosome arrays by phase separation. Luciferase reporter assays confirm that PHF1 phase separation promotes transcription repression, further supporting a role of the condensates in polycomb repression. Based on our findings, we propose that these condensates create favorable microenvironments at the target loci for PRC2 to function.

Enzymes have been shaped by evolution over billions of years to catalyse the chemical reactions that support life on earth. Dispersed in the literature, or organised in online databases, knowledge about enzymes can be structured in distinct dimensions, either related to their quality as biological macromolecules, such as their sequence and structure, or related to their chemical functions, such as the catalytic site, kinetics, mechanism, and overall reaction. The evolution of enzymes can only be understood when each of these dimensions is considered. In addition, many of the properties of enzymes only make sense in the light of evolution. We start this review by outlining the main paradigms of enzyme evolution, including gene duplication and divergence, convergent evolution, and evolution by recombination of domains. In the second part, we overview the current collective knowledge about enzymes, as organised by different types of data and collected in several databases. We also highlight some increasingly powerful computational tools that can be used to close gaps in understanding, in particular for types of data that require laborious experimental protocols. We believe that recent advances in protein structure prediction will be a powerful catalyst for the prediction of binding, mechanism, and ultimately, chemical reactions. A comprehensive mapping of enzyme function and evolution may be attainable in the near future.

Staphylococcal nuclease Tudor domain containing 1 (SND1) protein is an oncogene that 'reads' methylarginine marks through its Tudor domain. Specifically, it recognizes methylation marks deposited by protein arginine methyltransferase 5 (PRMT5), which is also known to promote tumorigenesis. Although SND1 can drive hepatocellular carcinoma (HCC), it is unclear whether the SND1 Tudor domain is needed to promote HCC. We sought to identify the biological role of the SND1 Tudor domain in normal and tumorigenic settings by developing two genetically engineered SND1 mouse models, an Snd1 knockout (Snd1 KO) and an Snd1 Tudor domain-mutated (Snd1 KI) mouse, whose mutant SND1 can no longer recognize PRMT5-catalyzed methylarginine marks. Quantitative PCR analysis of normal, KO, and KI liver samples revealed a role for the SND1 Tudor domain in regulating the expression of genes encoding major acute phase proteins, which could provide mechanistic insight into SND1 function in a tumor setting. Prior studies indicated that ectopic overexpression of SND1 in the mouse liver dramatically accelerates the development of diethylnitrosamine (DEN)-induced HCC. Thus, we tested the combined effects of DEN and SND1 loss or mutation on the development of HCC. We found that both Snd1 KO and Snd1 KI mice were partially protected against malignant tumor development following exposure to DEN. These results support the development of small molecule inhibitors that target the SND1 Tudor domain or the use of upstream PRMT5 inhibitors, as novel treatments for HCC.

Plants are vital to human health and well-being, as well as helping to protect the environment against the negative impacts of climate change. They are an essential part of the 'One Health' strategy that seeks to balance and optimize the health of people, animals and the environment. Crucially, plants are central to nature-based solutions to climate mitigation, not least because soil carbon storage is an attractive strategy for mitigating greenhouse gas emissions and the associated climate change. Agriculture depends on genetically pure, high-quality seeds that are free from pests and pathogens and contain a required degree of genetic purity. This themed collection addresses key questions in the field encompassing the biochemical mechanisms that underlie plant responses and adaptations to a changing climate. This collection encompasses an analysis of the biochemistry and molecular mechanisms underpinning crop and forest resilience, together with considerations of plant adaptations to climate change-associated stresses, including drought, floods and heatwaves, and the increased threats posed by pathogens and pests.

MASH is a prevalent liver disease that can progress to fibrosis, cirrhosis, hepatocellular carcinoma (HCC), and ultimately death, but there are no approved therapies. Leukotriene B4 (LTB4) is a potent pro-inflammatory chemoattractant that drives macrophage and neutrophil chemotaxis, and genetic loss or inhibition of its high affinity receptor, leukotriene B4 receptor 1 (BLT1), results in improved insulin sensitivity and decreased hepatic steatosis. To validate the therapeutic efficacy of BLT1 inhibition in an inflammatory and pro-fibrotic mouse model of MASH and fibrosis, mice were challenged with a choline-deficient, L-amino acid defined high fat diet and treated with a BLT1 antagonist at 30 or 90 mg/kg for 8 weeks. Liver function, histology, and gene expression were evaluated at the end of the study. Treatment with the BLT1 antagonist significantly reduced plasma lipids and liver steatosis but had no impact on liver injury biomarkers or histological endpoints such as inflammation, ballooning, or fibrosis compared to control. Artificial intelligence-powered digital pathology analysis revealed a significant reduction in steatosis co-localized fibrosis in livers treated with the BLT1 antagonist. Liver RNA-seq and pathway analyses revealed significant changes in fatty acid, arachidonic acid, and eicosanoid metabolic pathways with BLT1 antagonist treatment, however, these changes were not sufficient to impact inflammation and fibrosis endpoints. Targeting this LTB4-BLT1 axis with a small molecule inhibitor in animal models of chronic liver disease should be considered with caution, and additional studies are warranted to understand the mechanistic nuances of BLT1 inhibition in the context of MASH and liver fibrosis.

The exploitation of a cell's natural degradation machinery for therapeutic purposes is an exciting research area in its infancy with respect to bacteria. Here, we review current strategies targeting the ClpCP system, which is a proteolytic degradation complex essential in the biology of many bacterial species of scientific interest. Strategies include using natural product antibiotics or acyldepsipeptides to initiate the up- or down-regulation of ClpCP activity. We also examine exciting recent forays into BacPROTACs to trigger the degradation of specific proteins of interest through the hijacking of the ClpCP machinery. These strategies represent an important emerging avenue for combatting antimicrobial resistance.