Functional investigation of the SAM-dependent methyltransferase RdmB in anthracycline biosynthesis.

A novel sub-class of S-adenosyl-l-methionine (SAM)-dependent methyltransferases catalyze atypical chemical transformations in the biosynthesis of anthracyclines. Exemplified by RdmB from Streptomyces purpurascens, it was found with 10-decarboxylative hydroxylation activity on anthracyclines. We herein investigated the catalytic activities of RdmB and discovered a previously unknown 4-O-methylation activity. The site-directed mutagenesis studies proved that the residue at position R307 and N260 are vital for the decarboxylative hydroxylation and 4-O-methylation, respectively, which define two distinct catalytic centers in RdmB. Furthermore, the multifunctionality of RdmB activity was found as cofactor-dependent and stepwise. Our findings expand the versatility and importance of methyltransferases and should aid studies to enrich the structural diversity and bioactivities of anthracyclines.

Improved methods for genetic manipulation of the alkaliphile Halalkalibacterium halodurans.

An improved approach was developed for the genetic manipulation of the Gram-positive extremophile Halalkalibacterium halodurans (formerly called Bacillus halodurans). We describe an allelic replacement method originally developed for Staphylococcus aureus that allows the deletion, mutation, or insertion of genes without leaving markers or other genetic scars. In addition, a protocol for rapid in vitro plasmid methylation and transformation is presented. The combined methods allow the routine genetic manipulation of H. halodurans from initial transformation to the desired strain in 8 days. These methods improve H. halodurans as a model organism for the study of extremophiles.

Construction of a chemiluminescent biosensor based on enzymatic extension and click chemistry for sensitive measurement of MGMT activity in human breast tissues.

O6-methylguanine methyltransferase (MGMT) is responsible for dealkylation of naturally occurring O6-methylguanines, and it is closely related with DNA replication, transcription, and cancers. Herein, we develop a chemiluminescent biosensor based on enzymatic extension and click chemistry for sensitive measurement of MGMT activity. When MGMT is present, the MGMT-catalyzed demethylation reaction initiates the cleavage of biotinylated dumbbell probes by PvuII restrictive enzyme, releasing two DNA fragments with 3'-OH end. The resultant DNA fragments can trigger terminal transferase (TdT)- and click chemistry-assisted isothermal amplification to obtain abundant G-rich sequences. The G-rich sequences can be captured by magnetic beads to produce a high chemiluminescence signal. This biosensor can greatly amplify the chemiluminescence signal, facilitating label-free and template-free measurement of MGMT. Especially, the introduction of dumbbell probe and PvuII enzyme can efficiently eliminate the false positive and improve the assay specificity. This biosensor possesses high sensitivity with a detection limit of 1.4 × 10-9 ng/µL, and it may accurately quantify the intracellular MGMT. Importantly, this biosensor can be used to screen the MGMT inhibitors and distinguish the MGMT level in breast tumor tissues and normal tissues, with great potential in drug discovery and cancer diagnosis.

Identification of a polyphenol O-methyltransferase with broad substrate flexibility in Streptomyces albidoflavus J1074.

Flavonoids are a large and important group of phytochemicals with a great variety of bioactivities. The addition of methyl groups during biosynthesis of flavonoids and other polyphenols enhances their bioactivities and increases their stability. In a previous study of our research group, we detected a novel flavonoid O-methyltransferase activity in Streptomyces albidoflavus J1074, which led to the heterologous biosynthesis of homohesperetin from hesperetin in feeding cultures. In this study, we identify the O-methyltransferase responsible for the generation of this methylated flavonoid through the construction of a knockout mutant of the gene XNR_0417, which was selected after a blast analysis using the sequence of a caffeic acid 3'-O-methyltransferase from Zea mays against the genome of S. albidoflavus J1074. This mutant strain, S. albidoflavus ∆XNR_0417, was no longer able to produce homohesperetin after hesperetin feeding. Subsequently, we carried out a genetic complementation of the mutant strain in order to confirm that the enzyme encoded by XNR_0417 is responsible for the observed O-methyltransferase activity. This new strain, S. albidoflavus SP43-XNR_0417, was able to produce not only homohesperetin from hesperetin, but also different mono-, di-, tri- and tetra-methylated derivatives on other flavanones, flavones and stilbenes, revealing a broad substrate flexibility. Additionally, in vitro experiments were conducted using the purified enzyme on the substrates previously tested in vivo, demonstrating doubtless the capability of XNR_0417 to generate various methylated derivatives.

Cofactor Binding Triggers Rapid Conformational Remodelling of the Active Site of a Methyltransferase Ribozyme.

The methyltransferase ribozyme SMRZ-1 utilizes S-adenosyl-methionine (SAM) and Cu (II) ions to methylate RNA. Comparison of the SAM bound and unbound RNA structures has shown a conformational change in the RNA. However, the contribution of specific interactions and the role of a pseudo-triplex motif in the catalytic centre on the methylation reaction is not completely understood. In this study, we have used atomic substitutions and mutational analysis to investigate the reaction specificity and the key interactions required for catalysis. Substitution of the fluorescent nucleotide 2-aminopurine within the active ribozyme enabled the conformational dynamics of the RNA upon co-factor binding to be explored using fluorescence spectroscopy. We show that fast co-factor binding (t1/2 ∼ 0.7 seconds) drives a conformational change in the RNA to facilitate methyl group transfer. The importance of stacking interactions at the pseudo-triplex motif and chelation of the Cu (II) ion were shown to be essential for SAM binding.

Diversity and transcription of genes involved in respiratory As(V) reduction and As(III) methylation in Japanese paddy soils.

BACKGROUND: Arsenic (As) metabolism by soil microorganisms has an impact on As geochemical cycling in paddy soils, which in turn affects As uptake in rice. However, little is known about the key microorganisms involved in this process in Japanese paddy soil. RESULTS: Total RNA was extracted from Japanese paddy soils with different levels of dissolved As under flooded conditions, and the transcription of As metabolic genes (arrA, ttrA and arsM) was analyzed via a metatranscriptomic approach. The results showed that ttrA was the predominant respiratory arsenate reductase gene transcribed in these soils rather than arrA, suggesting that ttrA contributes to the reductive dissolution of As. The predominant taxa expressing ttrA differed among soils but were mostly associated with genera known for their iron- and/or sulfate-reduction activity. In addition, a wide variety of microorganisms expressed and upregulated arsM approximately 5.0- to 13.2-fold at 9 d compared with 3 d of incubation under flooded conditions in flasks. CONCLUSIONS: Our results support the involvement of microbial activity in the geochemical cycling of As in Japanese paddy soils and suggest that ttrA may be one of the key genes involved in the formation of arsenite, an inorganic species taken up by rice.

A regio-specific 4'-O-methyltransferase from Epimedium pseudowushanense regiospecifically catalyzing 8-prenylkeampferol to icaritin.

Epimedium is widely used in traditional Chinese medicine and contains rich bioactive compounds. These compounds often have a methyl group at their 4'-OH position catalyzed by methyltransferases. Therefore, studying methyltransferases in Epimedium plants is of great significance. In this study, a flavonol methyltransferase, EpOMT4, was isolated from Epimedium pseudowushanense B.L. Guo. The recombinant enzyme regiospecifically transferred a methyl group to the 4'-OH position of 8-prenylkaempferol forming icaritin. The study demonstrates that enzymatic methylation of flavonoids in Epimedium plants holds significant potential and could provide a promising alternative method for the biosynthetic production of bioactive methylated prenylflavonoids.

Nicotinamide N-methyltransferase (NNMT): A key enzyme in cancer metabolism and therapeutic target.

Emerging research has positioned Nicotinamide N-methyltransferase (NNMT) as a key player in oncology, with its heightened expression frequently observed across diverse cancers. This increased presence is tightly linked to tumor initiation, proliferation, and metastasis. The enzymatic function of NNMT is centered on the methylation of nicotinamide (NAM), utilizing S-adenosylmethionine (SAM) as the methyl donor, which results in the generation of S-adenosyl-L-homocysteine (SAH) and methyl nicotinamide (MNAM). This metabolic process reduces the availability of NAM, necessary for Nicotinamide adenine dinucleotide (NAD+) synthesis, and generates SAH, precursor to homocysteine (Hcy). These alterations are theorized to foster the resilience, expansion, and invasiveness of cancer cells. Furthermore, NNMT is implicated in enhancing cancer malignancy by affecting multiple signaling pathways, such as phosphatidylinositol 3-kinase (PI3K)-protein kinase B (AKT), cancer-associated fibroblasts (CAFs) and 5-Methyladenosine (5-MA), epithelial-mesenchymal transition (EMT), and epigenetic mechanisms. Upregulation of NNMT metabolism plays a key role in the formation and maintenance of the tumour microenvironment. While the use of small molecule inhibitors and RNA interference (RNAi) to target NNMT has shown therapeutic promise, the full extent of NNMT's influence on cancer is not yet fully understood, and clinical evidence is limited. This article systematically describes the relationship between the functional metabolism of NNMT enzymes and the cancer and tumour microenvironments, describing the mechanisms by which NNMT contributes to cancer initiation, proliferation, and metastasis, as well as targeted therapies. Additionally, we discuss the future opportunities and challenges of NNMT in targeted anti-cancer treatments.

Oral decitabine/cedazuridine versus intravenous decitabine for acute myeloid leukaemia: A randomised, crossover, registration, pharmacokinetics study.

This study compared decitabine exposure when administered IV (DEC-IV) at a dose of 20 mg/m2 for 5-days with orally administered decitabine with cedazuridine (DEC-C), as well as the clinical efficacy and safety of DEC-C in patients with acute myeloid leukaemia (AML) who were ineligible for intensive induction chemotherapy. In all, 89 patients were randomised 1:1 to DEC-IV or oral DEC-C (days 1-5 in a 28-day treatment cycle), followed by 5 days of the other formulation in the next treatment cycle. All patients received oral DEC-C for subsequent treatment cycles until treatment discontinuation. Equivalent systemic decitabine exposures were demonstrated (5-day area under the curve ratio between the two decitabine formulations of 99.64 [90% confidence interval 91.23%, 108.80%]). Demethylation rates also were similar (≤1.1% difference). Median overall survival (OS), clinical response and safety profile with oral DEC-C were consistent with those previously observed with DEC-IV. Next-generation sequencing was performed to identify molecular abnormalities that impact OS and TP53 mutations were associated with a poor outcome. These findings support the use of oral DEC-C in patients with AML.

Unravelling DNA methylation dynamics during developmental stages in Quercus ilex subsp. ballota [Desf.] Samp.

BACKGROUND: DNA methylation is a critical factor influencing plant growth, adaptability, and phenotypic plasticity. While extensively studied in model and crop species, it remains relatively unexplored in holm oak and other non-domesticated forest trees. This study conducts a comprehensive in-silico mining of DNA methyltransferase and demethylase genes within the holm oak genome to enhance our understanding of this essential process in these understudied species. The expression levels of these genes in adult and seedling leaves, as well as embryos, were analysed using quantitative real-time PCR (qRT-PCR). Global DNA methylation patterns were assessed through methylation-sensitive amplified polymorphism (MSAP) techniques. Furthermore, specific methylated genomic sequences were identified via MSAP sequencing (MSAP-Seq). RESULT: A total of 13 DNA methyltransferase and three demethylase genes were revealed in the holm oak genome. Expression levels of these genes varied significantly between organs and developmental stages. MSAP analyses revealed a predominance of epigenetic over genetic variation among organs and developmental stages, with significantly higher global DNA methylation levels observed in adult leaves. Embryos exhibited frequent demethylation events, while de novo methylation was prevalent in seedling leaves. Approximately 35% of the genomic sequences identified by MSAP-Seq were methylated, predominantly affecting nuclear genes and intergenic regions, as opposed to repetitive sequences and chloroplast genes. Methylation was found to be more pronounced in the exonic regions of nuclear genes compared to their promoter and intronic regions. The methylated genes were predominantly associated with crucial biological processes such as photosynthesis, ATP synthesis-coupled electron transport, and defence response. CONCLUSION: This study opens a new research direction in analysing variability in holm oak by evaluating the epigenetic events and mechanisms based on DNA methylation. It sheds light on the enzymatic machinery governing DNA (de)methylation, and the changes in the expression levels of methylases and demethylases in different organs along the developmental stages. The expression level was correlated with the DNA methylation pattern observed, showing the prevalence of de novo methylation and demethylation events in seedlings and embryos, respectively. Several methylated genes involved in the regulation of transposable element silencing, lipid biosynthesis, growth and development, and response to biotic and abiotic stresses are highlighted. MSAP-seq integrated with whole genome bisulphite sequencing and advanced sequencing technologies, such as PacBio or Nanopore, will bring light on epigenetic mechanisms regulating the expression of specific genes and its correlation with the phenotypic variability and the differences in the response to environmental cues, especially those related to climate change.

Targeting the Tumor Epigenetic Regulator SETDB1 for Tumor Therapy.

Epigenetic alterations are implicated in the early stages of tumorigenesis and are widely recognized as a ubiquitous phenomenon in cancer development. Aberrant epigenetic modifications can alter the expression of target genes, induce heterochromatin formation, and gradually drive normal cells towards immortalized tumor cells with significant consequences. SETDB1 (SET domain bifurcated histone lysine methyltransferase 1), a typical histone me-thyltransferase, promotes the formation of heterochromatin and inhibits the transcription of genes by modifying the methylation of lysine 9 of histone 3. SETDB1 is usually highly ex-pressed in tumors with high copy numbers, accompanied by poor prognosis and low patient survival rates, which is a typical case of abnormal epigenetic modification. We discuss the mechanism of SETDB1 in a variety of cancers and review the epigenetic inhibitors that have been reported in recent years, along with their anti-tumor effects. In addition, we summarize the role of SETDB1 in a variety of diseases and cell functions.

Overview of the PRMT6 modulators in cancer treatment: Current progress and emerged opportunity.

Protein Arginine Methyltransferase 6 (PRMT6) is a Type I PRMT enzyme that plays a role in the epigenetic regulation of gene expression by methylating histone and non-histone proteins. It is also involved in various cellular processes, including alternative splicing, DNA repair, and cell signaling. Furthermore, PRMT6 exerts multiple effects on cellular processes such as growth, migration, invasion, apoptosis, and drug resistance in various cancers, positioning it as a promising target for anti-tumor therapeutics. In this review, we initially provide an overview of the structure and biological functions of PRMT6, along with its association with cancer. Subsequently, we focus on recent progress in the design and development of modulators targeting PRMT6. This includes a comprehensive review of PRMT6 inhibitors (isoform-selective and non-selective), dual-target inhibitors based on PRMT6, PRMT6 covalent inhibitors, and PRMT6-targeting hydrophobic tagging (HyT) degraders, from the perspectives of rational design, pharmacodynamics, pharmacokinetics, and the clinical status of these modulators. Finally, we also provided the challenges and prospective directions for PRMT6 targeting drug discovery in cancer therapy.

The SET-Domain-Containing Protein MdSDG26 Negatively Regulates Alternaria alternata Resistance in Apple.

Apple leaf spot is one of the most devastating diseases in the apple industry, caused by Alternaria alternata f. sp mali (A. alternata). SET-domain group (SDG) proteins function as the histone methyltransferases and participate in plant development and stress responses. However, whether SDG proteins are associated with A. alternata resistance is largely unclear. Here, we describe the pathogen-inducible MdSDG26 gene in apple (Malus × domestica). MdSDG26 has two transcript variants that function similarly in catalyzing histone methylation and A. alternata resistance. Transient overexpression of MdSDG26 increased the global levels of H3K4me3 and H3K36me3, whereas knockdown of MdSDG26 only reduced the H3K36me3 level. Transcriptome analysis revealed that MdSDG26 affected the genome-wide transcriptome changes in response to A. alternata infection. ChIP-qPCR analysis demonstrated that MdSDG26 modulates the levels of H3K36me3 and H3K4me3 at both the promoter and exon regions of MdNTL9. As a negative regulator of A. alternata resistance in apples, MdNTL9 plays a pivotal role in MdSDG26-mediated resistance to A. alternata. Therefore, our findings provide compelling evidence for the regulatory function of MdSDG26 in histone methylation and its molecular role in conferring resistance to A. alternata.

Structural basis for inhibition of the SARS-CoV-2 nsp16 by substrate-based dual site inhibitors.

Coronaviruses, including SARS-CoV-2, possess an mRNA 5' capping apparatus capable of mimicking the natural eukaryotic capping signature. Two SAM-dependent methylating enzymes play important roles in this process: nsp14 methylates the N7 of the guanosine cap, and nsp16-nsp10 methylates the 2'-O- of subsequent nucleotides of viral mRNA. The 2'-O-methylation performed by nsp16-nsp10 is crucial for the escape of the viral RNA from innate immunity. Inhibition of this enzymatic activity has been proposed as a way to combat coronaviruses. In this study, we employed X-ray crystallography to analyze the binding of the SAM analogues to the active site of nsp16-nsp10. We obtained eleven 3D crystal structures of the nsp16-nsp10 complexes with SAM-derived inhibitors, demonstrated different conformations of the methionine substituting part of the molecules, and confirmed that simultaneous dual-site targeting of both SAM and RNA sites correlates with higher inhibitory potential.

Heterologous Expression of Sunflower HaHPT and HaTMT Genes Enhances Rice-Grain Vitamin E Content.

Insufficient dietary vitamin intake can lead to severe health conditions in humans. Improving the vitamin E (VE) content of food crops such as rice through breeding is an economical and effective means to alleviate this problem. In this study, Homogentisate phytyltransferase (HPT) and γ-tocopherol methyltransferase (γ-TMT), two genes derived from sunflower (Helianthus annuus L., a high VE species), were introduced into an elite rice (Oryza sativa L.) cultivar "Ningjing 7" for biofortification. We verified the successful expression of the two genes in multiple transformation events. High-performance liquid chromatography revealed that transgenic plants expressing either HaHPT alone or HaHPT and HaTMT accumulate more VE compared with the wild type. We also revealed that the level of α-tocopherol, the form of VE with the highest biological activity, had increased to 2.33 times in transgenic HaTMT plants compared with the wild type. Transcriptome analysis revealed that the expression levels of some chlorophyll synthesis pathway genes related to VE precursor synthesis significantly increased during grain filling in transgenic rice grains. No difference in agronomic traits was observed between the transgenic plants and their wild type except for a slightly reduced plant height associated with the transgenic plants. These data demonstrate that the heterologous expression of HaHPT gene is effective in increasing the total VE content, while HaTMT plays an important role in the relative abundance of α-tocopherol in rice grains. This study demonstrates a promising strategy for breeding rice with elevated VE content via metabolic engineering.

SETD3 functions beyond histidine methylation.

SETD3 is a member of SET domain-containing proteins. It has been discovered as the first metazoan protein (actin) histidine methyltransferase. In addition to this well-characterized molecular function of SETD3, it has been clearly shown to be involved in multiple biological processes, such as cell differentiation, tumorigenesis and viral infection. Here, we summarize the current knowledge on the roles of SETD3 beyond its histidine methyltransferase activity, and outline its cellular and molecular modes of action, as well as the upstream regulation on SETD3, therefore providing insights for the molecular basis of how SETD3 fine regulates multiple physiological and pathological processes.

Doping In Vivo Alkylation in E. coli by Introducing the Direct Sulfurylation Pathway of S. cerevisiae.

Methylation and alkylation are important techniques used for the synthesis and derivatisation of small molecules and natural products. Application of S-adenosylmethionine (SAM)-dependent methyltransferases (MTs) in biotechnological hosts such as Escherichia coli lowers the environmental impact of alkylations compared to chemical synthesis and facilitates regio- and chemoselective alkyl chain transfer. Here, we address the limiting factor for SAM synthesis, methionine supply, to accelerate in vivo methylation activity. Introduction of the direct sulfurylation pathway, consisting of O-acetylhomoserine sulfhydrolase (ScOAHS) and O-acetyltransferase (ScMET2), from S. cerevisiae into E. coli and supplementation with methanethiol or the corresponding disulfide improves atom-economic methylation activity in three different MT reactions. Up to 17-fold increase of conversion compared to the sole expression of the MT and incorporation of up to 79% of the thiol compound added were achieved. Promiscuity of ScOAHS allowed in vivo production of methionine analogues from organic thiols. Further co-overproduction of a methionine adenosyltransferase yielded SAM analogues which were further transferred by MTs onto different substrates. For methylation of non-physiological substrates, conversion rates up to 73% were achieved, with an isolated yield of 41% for N-methyl-2,5-aminonitrophenol. Our here described technique enables E. coli to become a biotechnological host for improved methylation and selective alkylation reactions.

Generation and characterization of two acid-resistant macrocin O-methyltransferase variants with a higher enzyme activity at 30 °C from Streptomyces fradiae.

Tylosin is an important macrolide antibiotic produced by Streptomyces fradiae. In the biosynthesis of tylosin, macrocin O-methyltransferase TylF catalyzes the conversion of the side-product tylosin C (macrocin) to the primary component tylosin A (C/A conversion). This conversion is the rate-limiting step in the biosynthesis of tylosin, and affects the quality of the end product. To find a high activity and environment-adapted TylF enzyme, a TylF variant pool has been constructed via protein evolution approach in our previous study (Fan et al., 2023 [41]). In this study, the TylF variants with higher C/A conversion rates were expressed in E. coli and purified. The variants TylFY139F, TylFQ138H, F232Y and TylFT36S, V54A were shown to have a higher C/A conversion rate at 30 °C than that of TylF at 38 °C. Moreover, they had a greater acid resistance and showed more adaptable to the pH change during fermentation. Further protein structural and substrate-binding affinity analyses revealed that the T36S, V54A, Q138H, Y139F, and F232Y mutations enlarged the volume of the substrate-binding pocket, thereby increasing the affinity of enzyme variants for their substrates of SAM and macrocin, and decreasing the inhibition of SAH. Three of the TylF variants were overexpressed in the industrial tylosin-producing S. fradiae strain, and the recombinant strains showed the highest C/A conversion at 30 °C without heating up to 38 °C during the last 24 h of fermentation. This is of great energy-saving significance for tylosin industrial production.

Parallel evolution of methyltransferases leads to vobasine biosynthesis in Tabernaemontana elegans and Catharanthus roseus.

Monoterpenoid indole alkaloids (MIA) are one of the largest and most complex alkaloid class in nature, boasting many clinically significant drugs such as anticancer vinblastine and antiarrhythmic ajmaline. Many MIAs undergo nitrogen N-methylation, altering their reactivity and affinity to the biological targets through a straightforward reaction. Remarkably, all known MIA N-methyltransferases (NMT) originate from the neofunctionalization of ancestral γ-tocopherol C-methyltransferases (γTMTs), a phenomenon seemingly unique to the Apocynaceae family. In this study, we unveil and characterize a new γTMT-like enzyme from the plant Tabernaemontana elegans (toad tree): perivine Nß-methyltransferase (TePeNMT). TePeNMT and other homologs form a distinct clade in our phylogenetic study, setting them apart from other γTMTs and γTMT-like NMTs discovered to date. Enzyme kinetic experiments and enzyme homology modeling studies reveal the significant differences in enzyme active sites between TePeNMT and CrPeNMT, a previously characterized perivine Nß-methyltransferase from Catharanthus roseus (Madagascar periwinkle). Collectively, our findings suggest that parallel evolution of ancestral γTMTs may be responsible for the occurrence of perivine N-methylation in T. elegans and C. roseus.

LONP1 alleviates ageing-related renal fibrosis by maintaining mitochondrial homeostasis.

Mitochondrial dysfunction is a pivotal event contributing to the development of ageing-related kidney disorders. Lon protease 1 (LONP1) has been reported to be responsible for ageing-related renal fibrosis; however, the underlying mechanism(s) of LONP1-driven kidney ageing with respect to mitochondrial disturbances remains to be further explored. The level of LONP1 was tested in the kidneys of aged humans and mice. Renal fibrosis and mitochondrial quality control were confirmed in the kidneys of aged mice. Effects of LONP1 silencing or overexpression on renal fibrosis and mitochondrial quality control were explored. In addition, N6-methyladenosine (m6A) modification and methyltransferase like 3 (METTL3) levels, the relationship between LONP1 and METTL3, and the impacts of METTL3 overexpression on mitochondrial functions were confirmed. Furthermore, the expression of insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) and the regulatory effects of IGF2BP2 on LONP1 were confirmed in vitro. LONP1 expression was reduced in the kidneys of aged humans and mice, accompanied by renal fibrosis and mitochondrial dysregulation. Overexpression of LONP1 alleviated renal fibrosis and maintained mitochondrial homeostasis, while silencing of LONP1 had the opposite effect. Impaired METTL3-m6A signalling contributed at least in part to ageing-induced LONP1 modification, reducing subsequent degradation in an IGF2BP2-dependent manner. Moreover, METTL3 overexpression alleviated proximal tubule cell injury, preserved mitochondrial stability, inhibited LONP1 degradation, and protected mitochondrial functions. LONP1 mediates mitochondrial function in kidney ageing and that targeting LONP1 may be a potential therapeutic strategy for improving ageing-related renal fibrosis.