Biosynthetic Functions Research Articles

Skeletal and dental tissue mineralization: The potential role of the endoplasmic reticulum/Golgi complex and the endolysosomal and autophagic transport systems.

This paper presents a review of the potential role of the endoplasmic reticulum/Golgi complex and intracellular vesicles in mediating events leading to or associated with vertebrate tissue mineralization. The possible importance of these organelles in this process is suggested by observations that calcium ions accumulate in the tubules and lacunae of the endoplasmic reticulum and Golgi. Similar levels of calcium ions (approaching millimolar) are present in vesicles derived from endosomes, lysosomes and autophagosomes. The cellular level of phosphate ions in these organelles is also high (millimolar). While the source of these ions for mineral formation has not been identified, there are sound reasons for considering that they may be liberated from mitochondria during the utilization of ATP for anabolic purposes, perhaps linked to matrix synthesis. Published studies indicate that calcium and phosphate ions or their clusters contained as cargo within the intracellular organelles noted above lead to formation of extracellular mineral. The mineral sequestered in mitochondria has been documented as an amorphous calcium phosphate. The ion-, ion cluster- or mineral- containing vesicles exit the cell in plasma membrane blebs, secretory lysosomes or possibly intraluminal vesicles. Such a cell-regulated process provides a means for the rapid transport of ions or mineral particles to the mineralization front of skeletal and dental tissues. Within the extracellular matrix, the ions or mineral may associate to form larger aggregates and potential mineral nuclei, and they may bind to collagen and other proteins. How cells of hard tissues perform their housekeeping and other biosynthetic functions while transporting the very large volumes of ions required for mineralization of the extracellular matrix is far from clear. Addressing this and related questions raised in this review suggests guidelines for further investigations of the intracellular processes promoting the mineralization of the skeletal and dental tissues.

Genome-Wide Network Analysis of DRG-Sciatic Nerve Network-Inferred Cellular Senescence and Senescence Phenotype in Peripheral Sensory Neurons.

Accumulation of senescent neurons in the dorsal root ganglion (DRG) is an important tissue phenotype that causes age-related degeneration of peripheral sensory nerves. Senescent neurons are neurons with arrested cell cycle that have undergone cellular senescence but remain in the tissue and play various biological roles. To understand the accumulation of senescent neurons in the DRG during aging, we aimed to elucidate the mechanism that induces cellular senescence in DRG neurons and the role of senescent DRG neurons. We integrated multiple public transcriptome datasets for DRGs, which include cell bodies in neurons, and the sciatic nerve, which includes axons in neurons, using network medicine-based bioinformatics analysis. We thus inferred the molecular mechanisms involved in cellular senescence of DRG neurons, from molecular responses to senescence, in the DRG-sciatic nerve network. Network medicine-based bioinformatics analysis revealed that age-related Mapk3 decline leads to impaired cholesterol metabolism and biosynthetic function in axons, resulting in compensatory upregulation of Srebf1, a transcription factor involved in lipid and cholesterol metabolism. This in turn leads to CDKN2A-mediated cellular senescence. Furthermore, our analysis revealed that senescent DRG neurons develop a senescence phenotype characterized by activation of antigen-presenting cells via upregulation of Ctss as a hub gene. B cells were inferred as antigen-presenting cells activated by Ctss, and CD8-positive T cells were inferred as cells that receive antigen presentation from B cells.

NifEN: a versatile player in nitrogenase assembly, catalysis and evolution.

The Mo-nitrogenase catalyzes the reduction of N2 to NH3 at the cofactor of its catalytic NifDK component. NifEN shares considerable homology with NifDK in primary sequence, tertiary structure and associated metallocenters. Better known for its biosynthetic function to convert an all-iron precursor (L-cluster; [Fe8S9C]) to a mature cofactor (M-cluster; [(R-homocitrate) MoFe7S9C]), NifEN also mimics NifDK in catalyzing substrate reduction at ambient conditions. The recently discovered ability of NifEN to reduce N2 to NH3 is particularly interesting, as it points to NifEN as a plausible, prototype ancient nitrogenase during evolution. Moreover, the dual function of NifEN in assembly and catalysis makes it a great template to reconstruct the functional variants or equivalents of NifDK, which could facilitate the mechanistic investigation and heterologous synthesis of nitrogenase. This perspective provides an overview of our recent studies of NifEN, with a focus on the implications of its functional versatility for nitrogenase assembly, catalysis and evolution.

VLDL induces EZH2/PRC2 complex-mediated cellular senescence in metabolic syndrome to enhance cardiotoxicity

Abstract Introduction In metabolic syndrome, very-low-density lipoprotein (VLDL) carries cardiotoxic lipids that cannot be completely modified by lipid-lowering medications. VLDL isolated from patients with metabolic syndrome (ms-VLDL) induces cardiac remodeling compared with that isolated from non-metabolic syndrome subjects (n-VLDL). Purpose This study focused on identifying dysregulated molecular pathways of ms-VLDL-associated with senescence. Methods RNA-sequencing, accompanied with IPA and KEGG pathway analysis were used to identify differentially expressed genes (DEGs) in cardiomyocytes after two different VLDL incubations, and the identified DEGs were subjected to functional annotation, including enriched molecular pathway and gene ontology analyses. A Seahorse XFe96 Analyzer was used to determine the mitochondrial biosynthesis oxygen consumption rate (OCR) and extracellular acidification rate (ECAR). Molecules involved in the mitochondrial pathway were validated using Western blotting, qPCR, and immunofluorescence. Results Compared to the n-VLDL group, the ms-VLDL group had a larger number of significant DEGs involved in several critical biological processes and molecular pathways, such as cardiomyocyte injury (ms-VLDL vs. n-VLDL for significant DEGs numbers, 2,642 vs. 1,725), senescence (259 vs. 184), and mitochondrial dysfunction (137 vs. 68). In terms of mitochondrial biosynthetic function, ms-VLDL induced 8.5 folds OCR reduction compared to n-VLDL, along with intra-mitochondrial calcium overload and excessive ROS. The essential genes involved in these pathways were shown to be differentially regulated, and our analysis identified the Ezh2/PRC2 complex, which is strongly linked to ms-VLDL-induced cardiomyocyte senescence and lipotoxicity. Conclusion This study identified the crucial roles of epigenetic modification enzyme Ezh2/PRC2 complex in induction the senescence pathway, which represents a potential mechanism for ms-VLDL-induced adverse cardiac remodeling in metabolic syndrome. Further functional studies are required to validate their roles in the pathogenesis of lipotoxicity and the connection of cardiomyopahty.RNA-seq of VLDL treated cardiomyocyteSenescence induction of VLDL

Transcriptomic Analysis of Hippocampus abdominalis Larvae Under High Temperature Stress.

Acute temperature stress was explored in Hippocampus abdominalis through a comprehensive RNA-seq analysis. RNA-seq was conducted on 20-day-old H. abdominalis after 24 h of temperature stress. Four experimental conditions were established: a control group (18 °C) and three temperature treatment groups (21, 24, and 27 °C). Seahorse larvae were found to be unaffected by 21 °C and 24 °C and were able to survive for short periods of time during 24 h of incubation, whereas mortality approached 50% at 27 °C. The sequencing process produced 75.63 Gb of high-quality clean data, with Q20 and Q30 base percentages surpassing 98% and 96%, respectively. A total of 141, 333, and 1598 differentially expressed genes were identified in the 21, 24, and 27 °C groups vs. a control comparison group, respectively. Notably, the number of up-regulated genes was consistently higher than that of down-regulated genes across all comparisons. Gene Ontology functional annotation revealed that differentially expressed genes were predominantly associated with metabolic processes, redox reactions, and biosynthetic functions. In-depth KEGG pathway enrichment analysis demonstrated that down-regulated genes were significantly enriched in pathways related to steroid biosynthesis, terpenoid backbone biosynthesis, spliceosome function, and DNA replication. Up-regulated genes were enriched in pathways associated with the FoxO signaling pathway and mitophagy (animal). The results indicated that temperature stress induced extensive changes in gene expression in H. abdominalis, involving crucial biological processes such as growth, biosynthesis, and energy metabolism. This study provided key molecular mechanisms in the response of H. abdominalis to temperature stress, offering a strong basis for future research aimed at understanding and mitigating the effects of environmental stressors on marine species.

Exploring the vitamin biosynthesis landscape of the human gut microbiota.

The human gut microbiota possesses the capacity to synthesize vitamins, especially B group vitamins, which are recognized as indispensable for various biological processes both among members of these bacterial communities and host cells. Accordingly, vitamin production by intestinal commensals has attracted significant interest. Nevertheless, our current understanding of bacterial vitamin synthesis is primarily based on individual genomic and monoculture investigations, therefore not providing an overall view of the biosynthetic potential of complex microbial communities. In the current study, we utilized over 100 bacterial genes known to be involved in the biosynthesis of B group and K vitamins to assess the corresponding vitamin biosynthetic potential of approximately 8,000 human gut microbiomes. Our analyses reveal that host-associated factors, such as age and geographical origin, appear to influence the diversity and abundance of vitamin biosynthetic pathways. Furthermore, we identify gut microbiota members that substantially contribute to these biosynthetic functions at each stage of human life. Interestingly, inference of microbial co-associations and network relationships uncovered the apparent key role played by folate and cobalamin in equilibrium establishment of the infant and adult gut microbial communities, respectively.IMPORTANCEOverall, this study expands our understanding of microbe-mediated vitamin biosynthesis in the human gut and may provide potential novel targets to improve availability of these essential micronutrients in the host.

Analysis of the characteristics of intestinal microbiota in patients with different severity of obstructive sleep apnea

Intestinal microbiota imbalance plays an important role in the progression of obstructive sleep apnea (OSA), and is considered to be the main mediator that triggers metabolic comorbidities. Here, we analyzed the changes in intestinal microbiota in patients with different severities of OSA based on apnea hypopnea index (AHI) classification, and explored the role of intestinal microbiota in the severity of OSA. This study included 19 healthy volunteers and 45 patients with OSA [5 ≤ AHI < 15 (n = 14), 15 ≤ AHI < 30 (n = 13), AHI ≥ 30 (n = 18)]. Relevant sleep monitoring data and medical history data were collected, and microbial composition was analyzed using 16S rRNA high-throughput sequencing technology. The diversity analysis of intestinal microbiota among different groups of people was conducted, including alpha diversity, beta diversity, species diversity, and marker species as well as differential functional metabolic pathway prediction analysis. With the increase of AHI classification, the alpha diversity in patients with OSA significantly decreased. The results revealed that the severity of OSA is associated with differences in the structure and composition of the intestinal microbiota. The abundance of bacteria producing short-chain fatty acids (such as Bacteroides, Ruminococcacea, and Faecalibacterium) in severe OSA is significantly reduced and a higher ratio of Firmicutes to Bacteroidetes. Random forest analysis showed that Parabacteroides was a biomarker genus with important discriminatory significance. The differential metabolic pathway prediction function shows that the main function of maintaining intestinal microbiota homeostasis is biosynthetic function. Our results show that the differences in the composition of intestinal microbiota in patients with different severities of OSA are mainly related to short-chain fatty acid-producing bacteria. These changes may play a pathological role in OSA combined with metabolic comorbidities.

CLOCI: unveiling cryptic fungal gene clusters with generalized detection.

Gene clusters are genomic loci that contain multiple genes that are functionally and genetically linked. Gene clusters collectively encode diverse functions, including small molecule biosynthesis, nutrient assimilation, metabolite degradation, and production of proteins essential for growth and development. Identifying gene clusters is a powerful tool for small molecule discovery and provides insight into the ecology and evolution of organisms. Current detection algorithms focus on canonical 'core' biosynthetic functions many gene clusters encode, while overlooking uncommon or unknown cluster classes. These overlooked clusters are a potential source of novel natural products and comprise an untold portion of overall gene cluster repertoires. Unbiased, function-agnostic detection algorithms therefore provide an opportunity to reveal novel classes of gene clusters and more precisely define genome organization. We present CLOCI (Co-occurrence Locus and Orthologous Cluster Identifier), an algorithm that identifies gene clusters using multiple proxies of selection for coordinated gene evolution. Our approach generalizes gene cluster detection and gene cluster family circumscription, improves detection of multiple known functional classes, and unveils non-canonical gene clusters. CLOCI is suitable for genome-enabled small molecule mining, and presents an easily tunable approach for delineating gene cluster families and homologous loci.

Pleiotropic Regulation of PGC-1α in Tumor Initiation and Progression.

Significance: Mitochondria are recognized as a central metabolic hub with bioenergetic, biosynthetic, and signaling functions that tightly control key cellular processes. As a crucial component of mitochondrial biogenesis, peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α) is involved in regulating various metabolic pathways, including energy metabolism and reactive oxygen species homeostasis. Recent Advances: Recent studies have highlighted the significant role of PGC-1α in tumorigenesis, cancer progression, and treatment resistance. However, PGC-1α exhibits pleiotropic effects in different cancer types, necessitating a more comprehensive and thorough understanding. Critical Issues: In this review, we discuss the structure and regulatory mechanisms of PGC-1α, analyze its cellular and metabolic functions, explore its impact on tumorigenesis, and propose potential strategies for targeting PGC-1α. Future Directions: The targeted adjustment of PGC-1α based on the metabolic preferences of different cancer types could offer a hopeful therapeutic approach for both preventing and treating tumors. Antioxid. Redox Signal. 41, 557-572.

Revision of the sophorolipid biosynthetic pathway in Starmerella bombicola based on new insights in the substrate profile of its lactone esterase

BackgroundSophorolipids (SLs) are a class of natural, biodegradable surfactants that found their way as ingredients for environment friendly cleaning products, cosmetics and nanotechnological applications. Large-scale production relies on fermentations using the yeast Starmerella bombicola that naturally produces high titers of SLs from renewable resources. The resulting product is typically an extracellular mixture of acidic and lactonic congeners. Previously, we identified an esterase, termed Starmerella bombicola lactone esterase (SBLE), believed to act as an extracellular reverse lactonase to directly use acidic SLs as substrate.ResultsWe here show based on newly available pure substrates, HPLC and mass spectrometric analysis, that the actual substrates of SBLE are in fact bola SLs, revealing that SBLE actually catalyzes an intramolecular transesterification reaction. Bola SLs contain a second sophorose attached to the fatty acyl group that acts as a leaving group during lactonization.ConclusionsThe biosynthetic function by which the Starmerella bombicola ‘lactone esterase’ converts acidic SLs into lactonic SLs should be revised to a ‘transesterase’ where bola SL are the true intermediate. This insights paves the way for alternative engineering strategies to develop designer surfactants.

Features of HPV+ OPSCC predictive of recurrence identified by transcriptional profiling of a case-control cohort.

6055 Background: De-escalating therapy for HPV+ oropharyngeal cancers (OPSCCs) is hampered by poor ability to predict recurrence, and limited insight into the biologic traits predisposing to recurrence impedes personalizing therapy. We aimed to (1) define biologic features underlying therapy resistance by transcriptional profiling and (2) evaluate the features for prognostic utility. Methods: A single institution cohort of 851 HPV+ OPSCC patients undergoing transoral robotic surgery during 2007-2020 was used to identify 50 cases that recurred locoregionally in the adjuvant RT field (n=14) and/or at distant sites (n=43). As controls, we used 49 recurrence-free patients with long-term follow up and similar stage, smoking history, and adjuvant therapy. RNAseq of pretreatment tumors was used to compare individual gene expression and Hallmark/Kegg pathway activity between cases and controls using unpaired t-test (p&lt;.05) and Gene Set Enrichment Analysis (p-adj&lt;.05), respectively. Activity of the significant pathways was quantified in individual tumors using Gene Set Variation Analysis (GSVA), and a regression-based composite of GSVA scores was developed to distinguish cases from controls by ROC analysis. The composite score was used to test for stratification of recurrence free survival (RFS) in external cohorts using Youden Index and logrank test. Results: The21 Hallmark/Kegg pathways downregulated in cases indicated suppression of anti-tumor immunity, and the 20 upregulated ones revealed increased biosynthetic function and tumor cell proliferation. The 1472 upregulated mRNAs contained several components of the ATR-chk1 DNA repair pathway and trans-lesion synthesis polymerases, suggesting that mitigation of DNA replication stress enhanced tumor growth. The 958 downregulated mRNAs suggested reductions in the cytoplasmic dsDNA sensing and downstream NF-kB signals that lead to replication stress-driven anti-tumor immunity. Two GSVA scores were created to quantify the key tumor-intrinsic and immune suppressive features separately in each tumor. The scores correlated with each other (R=0.6, p&lt;.001), and a regression-based combination of them distinguished cases from controls (AUC=.76, p&lt;.001). This composite score also stratified RFS in three external cohorts: TCGA (n=52, p&lt;0.001) and two single institution cohorts containing both surgical and nonsurgical cases (n=46, p=.002; n=81, p&lt;0.001). Conclusions: HPV+ OPSCCs failing primary surgical therapy show evidence of reduced replication stress mediating tumor progression and low anti-tumor immunity. These features appear generalizable to heterogeneously treated external cohorts, where they predicted recurrence. Our results provide a new basis for creating transcriptomic predictors of treatment response and suggest targetable molecular mechanisms to overcome therapy resistance.

Advances of Bacterial Biomaterials for Disease Therapy.

Bacteria have immense potential as biological therapeutic agents that can be used to treat diseases, owing to their inherent immunomodulatory activity, targeting capabilities, and biosynthetic functions. The integration of synthetic biomaterials with natural bacteria has led to the construction of bacterial biomaterials with enhanced functionality and exceptional safety features. In this review, recent progress in the field of bacterial biomaterials, including bacterial drug delivery systems, bacterial drug-producing factories, bacterial biomaterials for metabolic engineering, bacterial biomaterials that can be remotely controlled, and living bacteria hydrogel formulations, is described and summarized. Furthermore, future trends in advancing next-generation bacterial biomaterials for enhanced clinical applications are proposed in the conclusion.

Abstract 6608: Therapeutic enzyme depletion of L-serine for the treatment of serine auxotrophic tumors

Abstract The non-essential amino acid L-serine (serine) plays an important role in cancer cell growth due to its diverse biosynthetic functions. In many cancer cells, serine can be taken up from the circulation or can be synthesized through the serine synthesis pathway (SSP). We have recently discovered that luminal breast tumors are auxotrophic for serine due to lineage-specific silencing of the SSP gene PSAT1, rendering luminal breast cancer cells sensitive to serine deprivation both in vitro and in vivo. At present the only way to reduce serine availability for tumors in vivo is through dietary serine starvation, which in mice reduces circulating serine levels by ~50% and has been shown to inhibit the growth of numerous mouse models of cancer. Importantly, dietary serine is achieved in mice using a synthetic protein-free diet that may be difficult for humans to sustain. Nevertheless, dietary serine starvation is currently being explored in a clinical trial for pancreatic cancer. To circumvent some of the challenges associated with dietary serine starvation, we have developed a therapeutic serine degrading enzyme that we are exploring as a potential treatment for serine auxotrophic tumors. Our enzyme, eSDH, is based on the human serine dehydratase enzyme that facilitates the pyridoxal phosphate-dependent irreversible deamination of serine to pyruvate and ammonia. Relative to wild-type serine dehydratase, eSDH has significantly improved stability in serum as well increased activity towards serine. Injection of eSDH into mice is well-tolerated and leads to long-term, near-complete (&amp;gt;95%) reduction in circulating serine levels without the need for dietary changes. Importantly, eSDH treatment inhibits the growth of serine auxotrophic cancer cells in vitro and in vivo. We are currently assessing the efficacy of eSDH treatment in advanced models of luminal/ER+ breast cancer and when utilized in combination with endocrine therapy and other standard-of-care treatments for luminal/ER+ cancer patients. In addition to the degradation of serine, eSDH also depletes circulating L-threonine (threonine), which could contribute to the in vivo activity of eSDH. However, we have recently developed new eSDH variants that are highly selective for serine over threonine and show enhanced specificity for serine auxotrophic cancer cells. In conclusion, we have developed a prototypical therapeutic serine degrading enzyme that may provide an opportunity to clinically achieve more complete serine starvation in patients without the need for restrictive dietary changes. Citation Format: Vipin Rawat, Huiping Zhao, Ladan Mashouri, Prarthana Prasanth, Kelly Conger, Sarita Bhetawal, Alessandra Araujo, Everett Stone, Jonathan Coloff. Therapeutic enzyme depletion of L-serine for the treatment of serine auxotrophic tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6608.

Qualitative and quantitative changes in mitochondrial DNA associated with cervical cancer: A comprehensive review.

Cervical cancer is the fourth most commonly diagnosed cancer in women and is considered a preventable disease, as vaccination and screening programs effectively reduce its incidence and mortality rates. Disease physiopathology and malignant cell transformation is a complex process, but it is widely known that high-risk HPV (hrHPV) infection is a necessary risk factor for cancer development. Mitochondria, cell organelles with important bioenergetic and biosynthetic functions, are important for cell energy production, cell growth, and apoptosis. Mitochondrial DNA is a structure that is particularly susceptible to quantitative (mtDNA copy number variation) and qualitative (sequence variations) alterations that are associated with various types of cancer. Novel biomarkers with diagnostic and prognostic value in cervical cancer can be evaluated to provide higher specificity and complement hrHPV molecular testing, which is the most recommended method for primary screening. In accordance with this, this review aimed to assess mitochondrial alterations associated with cervical cancer in clinical cervicovaginal samples, in order to unravel their possible role as specific diagnostic and prognostic biomarkers for cervical malignancy, and also to guide the understanding of their involvement in carcinogenesis, HPV infection, and disease progression.

Development of a mouse model expressing a bifunctional glutathione-synthesizing enzyme to study glutathione limitation in vivo

Glutathione (GSH) is a highly abundant tripeptide thiol that performs diverse protective and biosynthetic functions in cells. While changes in GSH availability are associated with inborn errors of metabolism, cancer and neurodegenerative disorders, studying the limiting role of GSH in physiology and disease has been challenging due to its tight regulation. To address this, we generated cell and mouse models that express a bifunctional glutathione-synthesizing enzyme from Streptococcus thermophilus (GshF), which possesses both glutamate-cysteine ligase and glutathione synthase activities. GshF expression allows efficient production of GSH in the cytosol and mitochondria and prevents cell death in response to GSH depletion, but not ferroptosis induction, indicating that GSH is not a limiting factor under lipid peroxidation. CRISPR screens using engineered enzymes further revealed genes required for cell proliferation under cellular and mitochondrial GSH depletion. Among these, we identified the glutamate-cysteine ligase modifier subunit, Gclm, as a requirement for cellular sensitivity to buthionine sulfoximne, a glutathione synthesis inhibitor. Finally, GshF expression in mice is embryonically lethal but sustains postnatal viability when restricted to adulthood. Overall, our work identifies a conditional mouse model to investigate the limiting role of GSH in physiology and disease.

AGE-RELATED MECHANISMS OF ALTERED TENDON STRUCTURE AND FUNCTION

During aging, tendons demonstrate substantial disruptions in homeostasis, leading to impairments in structure-function. Impaired tendon function contributes to substantial declines quality of life during aging. Aged tendons are more likely to undergo spontaneous rupture, and the healing response following injury is impaired in aged tendons. Thus, there is a need to develop strategies to maintain tendon homeostasis and healing capacity through the lifespan. Tendon cell density sharply declines by ∼12 months of age in mice, and this low cell density is retained in geriatric tendons. Our data suggests that this decline in cellularity initiates a degenerative cascade due to insufficient production of the extracellular matrix (ECM) components needed to maintain tendon homeostasis. Thus, preventing this decline in tendon cellularity has great potential for maintaining tendon health. Single cell RNA sequencing analysis identifies two changes in the aged tendon cell environment. First, aged tendons primarily lose tenocytes that are associated with ECM biosynthesis functions. Second, the tenocytes that remain in aged tendons have disruptions in proteostasis and an increased pro-inflammatory phenotype, with these changes collectively termed ‘programmatic skewing'. To determine which of these changes drives homeostatic disruption, we developed a model of tenocyte depletion in young animals. This model decreases tendon cellularity to that of an aged tendon, including decreased biosynthetic tenocyte function, while age-related programmatic skewing is absent. Loss of biosynthetic tenocyte function in young tendons was sufficient to induce homeostatic disruption comparable to natural aging, including deficits in ECM organization, composition, and material quality, suggesting loss biosynthetic tenocytes as an initiator of tendon degeneration. In contrast, our data suggest that programmatic skewing underpins impaired healing in aged tendons. Indeed, despite similar declines in the tenocyte environment, middle-aged and young-depleted tendons mount a physiological healing response characterized by robust ECM synthesis and remodeling, while aged tendons heal with insufficient ECM.

Citrus limon (L.) Osbeck Fruit Peel Extract Attenuates Carbon Tetrachloride-Induced Hepatocarcinogenesis in Sprague-Dawley Rats.

Traditional herbal medicine practitioners in the Ashanti region of Ghana use the fruit peels of Citrus limon (L.) Osbeck (C. limon) in preventive and curative treatment of many cancers including liver cancer. This ethnobotanical claim remains to be verified scientifically. Aim of the Study. This study investigated prophylactic hepatoprotective and anti-HCC effects of C. limon peel extract (LPE) in CCl4/olive oil-induced HCC-like rats. After preparation of LPE, it was subjected to phytochemical screening using standard phytochemical methods. A total of 30 healthy adult male Sprague-Dawley rats (weighing 150-200 g) were randomly assigned into six groups of 5 rats each. Rats in the control group received olive oil (5 mL/kg ip) twice weekly for 16 weeks. Rats in the model group received CCl4/olive oil (2 mL/kg, ip) twice weekly for 16 weeks. Rats in capecitabine (10 mg/kg po) and LPE (50, 100, and 200 mg/kg po) groups received CCl4/olive oil (2 mL/kg, i.p) in the morning and their respective treatments in the afternoon twice a week for 16 weeks. Rats in all groups had free access to food and water ad libitum. Body weight and survival rates were monitored. Rats were sacrificed under deep anesthesia, blood was collected, and liver and other organs were isolated. Aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP), gamma-glutamyltransferase (GGT), prothrombin time, bilirubin, C-reactive protein (CRP), alpha- (α-) fetoprotein (AFP), and liver histology were assessed. Alkaloids, tannins, flavonoids, terpenoids, and saponins were detected in LPE. Model rats demonstrated increased serum levels of AFP, CRP, ALP, GGT, ALT, and AST, prothrombin time, total bilirubin, direct bilirubin, blood lymphocyte, and monocyte counts, but decreased serum albumin and total protein compared to control rats. Unlike the control, model rats demonstrated fat accumulation in periportal and centrilobular hepatocytes and neoplastic transformation. Semiquantitation of periodic acid Schiff- (PAS-) stained liver sections showed decreased glycogen storage in hepatocytes of model rats compared to control rats. Compared to the model, LPE treatment protected against CCl4-induced hepatocarcinogenesis, which was evidenced by decreased AFP, CRP, liver enzymes, total and direct bilirubin, prothrombin time, and blood lymphocyte and monocyte counts; attenuation of fat accumulation; and increased glycogen storage, albumin, and total protein. LPE abates CCl4-induced hepatocarcinogenesis by attenuating liver inflammation and improving metabolic, biosynthetic, and detoxification functions of the liver. The prophylactic hepatoprotective and anti-hepatocarcinogenic effects of LPE are attributable to its phytochemical composition raising hopes of finding potential anticancer bioactive compounds from C. limon fruit peels.

Genome mining and biosynthetic pathways of marine-derived fungal bioactive natural products.

Marine fungal natural products (MFNPs) are a vital source of pharmaceuticals, primarily synthesized by relevant biosynthetic gene clusters (BGCs). However, many of these BGCs remain silent under standard laboratory culture conditions, delaying the development of novel drugs from MFNPs to some extent. This review highlights recent efforts in genome mining and biosynthetic pathways of bioactive natural products from marine fungi, focusing on methods such as bioinformatics analysis, gene knockout, and heterologous expression to identify relevant BGCs and elucidate the biosynthetic pathways and enzyme functions of MFNPs. The research efforts presented in this review provide essential insights for future gene-guided mining and biosynthetic pathway analysis in MFNPs.

Synthesis and application of a compound microbial inoculant for effective soil remediation.

Currently, there is a noticeable scarcity of applications that harness composite microbial inoculants to stimulate straw decomposition, nitrogen fixation, and crop growth. This study addresses this gap by selecting and coculturing three bacterial strains to create a composite microbial inoculant named HY-1. This innovative inoculant exhibits multifunctional capabilities, including nitrogen fixation, straw decomposition, and crop growth promotion. Furthermore, we aimed to explore its impact on soil microbial communities. The results showed that the optimal preparation conditions for the compound microbial inoculant HY-1 were 28.5 ± 0.6°C, pH = 7.34 ± 0.40, and bacteriophage ratio 1:2:1 (Microbacterium: Streptomyces fasciatus: Bacillus amyloliquefaciens). Compared to single strains, the combination exhibited higher levels of cellulose-degrading and nitrogen-fixing enzyme activity, increased the straw degradation rate by 37.91% within 180days, and significantly promoted the growth of corn seedlings. Under the condition of straw return, the compound bio-fungicide HY-1 effectively improved the soil microbial diversity. At that time, the soil had the highest number of unique bacterial operational taxonomic units (166), and the abundance of Proteobacteria in the soil increased by 7.24%, while that of Acidobacteriota decreased by 2.27%. The biosynthetic function of the cell wall/membrane/periplasm and the metabolic function of transporting inorganic ions were significantly enhanced. In this study, we discovered that employing coculturing techniques to produce the composite microbial inoculant HY-1 and applying it in the field effectively compensates for the limitations of single-strain inoculants, which often exhibit fewer functions and less pronounced effects. This approach demonstrates significant potential for enhancing the quality of agricultural soils.

Autoregulatory control of mitochondrial glutathione homeostasis.

Mitochondria must maintain adequate amounts of metabolites for protective and biosynthetic functions. However, how mitochondria sense the abundance of metabolites and regulate metabolic homeostasis is not well understood. In this work, we focused on glutathione (GSH), a critical redox metabolite in mitochondria, and identified a feedback mechanism that controls its abundance through the mitochondrial GSH transporter, SLC25A39. Under physiological conditions, SLC25A39 is rapidly degraded by mitochondrial protease AFG3L2. Depletion of GSH dissociates AFG3L2 from SLC25A39, causing a compensatory increase in mitochondrial GSH uptake. Genetic and proteomic analyses identified a putative iron-sulfur cluster in the matrix-facing loop of SLC25A39 as essential for this regulation, coupling mitochondrial iron homeostasis to GSH import. Altogether, our work revealed a paradigm for the autoregulatory control of metabolic homeostasis in organelles.