Gene Activation Research Articles

A delayed response in phytohormone signaling and production contributes to pine susceptibility to Fusarium circinatum

BackgroundFusarium circinatum is the causal agent of pine pitch canker disease, which affects Pinus species worldwide, causing significant economic and ecological losses. In Spain, two Pinus species are most affected by the pathogen; Pinus radiata is highly susceptible, while Pinus pinaster has shown moderate resistance. In F. circinatum-Pinus interactions, phytohormones are known to play a crucial role in plant defense. By comparing species with different degrees of susceptibility, we aimed to elucidate the fundamental mechanisms underlying resistance to the pathogen. For this purpose, we used an integrative approach by combining gene expression and metabolomic phytohormone analyses at 5 and 10 days post inoculation.ResultsGene expression and metabolite phytohormone contents suggested that the moderate resistance of P. pinaster to F. circinatum is determined by the induction of phytohormone signaling and hormone rearrangement beginning at 5 dpi, when symptoms are still not visible. Jasmonic acid was the hormone that showed the greatest increase by 5 dpi, together with the active gibberellic acid 4 and the cytokinin dehydrozeatin; there was also an increase in abscisic acid and salicylic acid by 10 dpi. In contrast, P. radiata hormonal changes were delayed until 10 dpi, when symptoms were already visible; however, this increase was not as high as that in P. pinaster. Indeed, in P. radiata, no differences in jasmonic acid or salicylic acid production were found. Gene expression analysis supported the hormonal data, since the activation of genes related to phytohormone synthesis was observed earlier in P. pinaster than in the susceptible P. radiata.ConclusionsWe determine that the moderate resistance of P. pinaster to F. circinatum is in part a result of early and strong activation of plant phytohormone-based defense responses before symptoms become visible. We suggest that jasmonic acid signaling and production are strongly associated with F. circinatum resistance. In contrast, P. radiata susceptibility was attributed to a delayed response to the fungus at the moment when symptoms were visible. Our results contribute to a better understanding of the phytohormone-based defense mechanism involved in the Pinus-F. circinatum interactions and provide insight into the development of new strategies for disease mitigation.

On-Site Inactivation for Disinfection of Antibiotic-Resistant Bacteria in Hospital Effluent by UV and UV-LED.

The problem of antimicrobial resistance (AMR) is not limited to the medical field but is also becoming prevalent on a global scale in the environmental field. Environmental water pollution caused by the discharge of wastewater into aquatic environments has caused concern in the context of the sustainable development of modern society. However, there have been few studies focused on the treatment of hospital wastewater, and the potential consequences of this remain unknown. This study evaluated the efficacy of the inactivation of antimicrobial-resistant bacteria (AMRB) and antimicrobial resistance genes (AMRGs) in model wastewater treatment plant (WWTP) wastewater and hospital effluent based on direct ultraviolet (UV) light irradiation provided by a conventional mercury lamp with a peak wavelength of 254 nm and an ultraviolet light-emitting diode (UV-LED) with a peak emission of 280 nm under test conditions in which the irradiance of both was adjusted to the same intensity. The overall results indicated that both UV- and UV-LED-mediated disinfection effectively inactivated the AMRB in both wastewater types (>99.9% after 1-3 min of UV and 3 min of UV-LED treatment). Additionally, AMRGs were also removed (0.2-1.4 log10 for UV 254 nm and 0.1-1.3 log10 for UV 280 nm), and notably, there was no statistically significant decrease (p < 0.05) in the AMRGs between the UV and UV-LED treatments. The results of this study highlight the importance of utilizing a local inactivation treatment directly for wastewater generated by a hospital prior to its flow into a WWTP as sewage. Although additional disinfection treatment at the WWTP is likely necessary to remove the entire quantity of AMRB and AMRGs, the present study contributes to a significant reduction in the loads of WWTP and urgent prevention of the spread of infectious diseases, thus alleviating the potential threat to the environment and human health risks associated with AMR problems.

Screening of indigenous Bacillus thuringiensis for dipteran active cry gene profiles and potential toxicity against melon fruit fly, Zeugodacus cucurbitae (Coquillett)

BackgroundMelon fruit fly, Zeugodacus cucurbitae Coquillett (Diptera: Tephritidae), is a devastating polyphagous pest attacking large number of fruits and vegetables causing huge economic yield losses across the world. Management of this notorious pest is very challenging as the larvae feed inside the fruit. Hence, the present research study aimed to screen the indigenous Bacillus thuringiensis Berliner (Bacillales: Bacillaceae) strains causing toxicity to larvae and to identify the insecticidal toxicity-related genes present in respective strains. In the present study, 50 indigenous B. thuringiensis (Bt) strains along with one reference strain were screened against second-instar larvae Z. cucurbitae. All the strains were analyzed for presence of 21 dipteran active cry genes.ResultsMortality in Z. cucurbitae larvae due to Bt strains ranged from 16 to 92%. PCR results revealed that each strain tested positive for a minimum of three cry genes to maximum of nine cry genes. Among the cry genes, cry1A, cry2A, cry1C, cry19, cry11 and cry70 were detected in high frequency of 100, 88, 84, 74, 58 and 54%, respectively. Bioassay studies revealed that ten out of fifty strains displayed more than 50% mortality. Hence, these ten strains, along with the reference strain, were further tested for mortality for the calculation of the median lethal concentration (LC50). The LC50 values ranged between 38.48 and 105.18 μg/ml. The lowest LC50 found for the strain NBAIR Bt107 was 38.48 μg/ml and was on par with the reference strain (Bti 4Q1) (31.3 μg/ml).ConclusionIndigenous Bt strains displayed a toxicity against the larvae of Z. cucurbitae. The probable dipteran active cry genes responsible for toxicity were interpreted. Thus, the Cry toxins from Bt can play a very important role in the management of Z. cucurbitae.

Whatever does not kill them makes them stronger: Using quaternary ammonia antimicrobials to alleviate the inhibition of ammonia oxidation under perfluorooctanoic acid stress

Perfluorooctanoic acid (PFOA), benzalkyl dimethylammonium compounds (BAC) and antibiotic resistance genes (ARGs) have negative effects on biological sewage treatment. The performance of nitrification systems under stress of PFOA (0.1–5 mg/L) or/and BAC (0.2–10 mg/L) was explored during 84-day experiments using four sequencing batch reactors, in this study. Low (0.1 mg/L) concentration PFOA had a positive influence on ammonia removal, while medium and high (2 and 5 mg/L) concentrations PFOA caused severe inhibition. Meanwhile, PFOA stress resulted in the enrichment of ARGs in water (w-ARGs). BAC (0–10 mg/L) had no obvious influence on ammonia removal. However, BAC promoted the reduction of ARGs and the bacterial community was the main participator (48.07%) for the spread of ARGs. Interestingly, the joint stress of PFOA and BAC increased the ammonia-oxidizing bacteria (AOB) activity from 5.81 ± 0.19 and 6.05 ± 0.79 mg N/(g MLSS·h) to 7.09 ± 0.87 and 7.23 ± 0.29 mg N/(g MLSS·h) in medium and high concentrations, compared to single stress of PFOA, which was observed for the first time. BAC could reduce bioavailability of PFOA through competitive adsorption and decreasing sludge hydrophobicity by the lower β-Sheet and α-Helix in tightly bound protein. Furthermore, the joint stress of PFOA and BAC was able to intensify the proliferation of w-ARGs and extracellular ARGs in sludge, and developed the most active horizontal gene transfer mediated by intl1 compared to single stress of PFOA or BAC. The batch tests verified the detoxification capacity of BAC on nitrification under 2.5 mg/L PFOA (48 h exposing), and the maximum alleviation of AOB activity was achieved at BAC and PFOA mass ratio of 2:1. In summary, BAC could be used to alleviate the inhibition of PFOA on ammonia oxidation, providing an efficient and sustainable approach in wastewater treatment.

Strong activation of p53 by actinomycin D and nutlin-3a overcomes the resistance of cancer cells to the pro-apoptotic activity of the FAS ligand.

The FAS ligand (FASLG) is expressed on lymphocytes, which employ it to activate death receptors on target cells. Cancer cells are generally resistant to apoptosis triggered by FASLG. In this work, we found a way to circumvent this resistance by treatment with actinomycin D (ActD) and nutlin-3a (Nut3a). We selected this drug combination based on our transcriptomic data showing strong activation of proapoptotic genes, including those for receptor-mediated apoptosis, in cells exposed to actinomycin D and nutlin-3a. To test our hypothesis, we pre-exposed cancer cell lines to this drug combination for 45h and then treated them with recombinant FASLG. This almost instantaneously killed most cells. Actinomycin D and nutlin-3a strongly cooperated in the sensitization because the effect of the drugs acting solo was not as spectacular as the drug combination, which together with FASLG killed more than 99% of cells. Based on the caspase activation pattern (caspase-8, caspase-9, caspase-10), we conclude that both extrinsic and intrinsic pro-apoptotic pathways were engaged. In engineered p53-deficient cells, this pro-apoptotic effect was completely abrogated. Therefore, the combination of ActD + Nut3a activates p53 in an extraordinary way, which overcomes the resistance of cancer cells to apoptosis triggered by FASLG. Interestingly, other combinations of drugs, e.g., etoposide + nutlin-3a, actinomycin D + RG7112, and actinomycin D + idasanutlin had a similar effect. Moreover, normal human fibroblasts are less sensitive to death induced by ActD + Nut3a + FASLG. Our findings create the opportunity to revive the abandoned attempts of cancer immunotherapy employing the recombinant FAS ligand.

Gene body DNA hydroxymethylation restricts the magnitude of transcriptional changes during aging

DNA hydroxymethylation (5hmC), the most abundant oxidative derivative of DNA methylation, is typically enriched at enhancers and gene bodies of transcriptionally active and tissue-specific genes. Although aberrant genomic 5hmC has been implicated in age-related diseases, its functional role in aging remains unknown. Here, using mouse liver and cerebellum as model organs, we show that 5hmC accumulates in gene bodies associated with tissue-specific function and restricts the magnitude of gene expression changes with age. Mechanistically, 5hmC decreases the binding of splicing associated factors and correlates with age-related alternative splicing events. We found that various age-related contexts, such as prolonged quiescence and senescence, drive the accumulation of 5hmC with age. We provide evidence that this age-related transcriptionally restrictive function is conserved in mouse and human tissues. Our findings reveal that 5hmC regulates tissue-specific function and may play a role in longevity.

CpAGL18 is involved in the cross-talk of ethylene and auxin signaling in the regulation of fruit ripening in papaya

1-methylcyclopropene (1-MCP) effectively delays the ripening of papaya fruit, but inappropriate 1-MCP treatments induce ripening disorders. Thus, this study aimed to clarify the roles of CpAGL18 in regulating papaya fruit ripening and the possible mechanism underlying the ripening disorder. The expression of CpAGL18, a MADS-box transcription factor, is closely related to papaya fruit ripening. Transient and ectopic overexpression of CpAGL18 promotes papaya and tomato fruit ripening. However, the transient silencing of CpAGL18 repressed fruit ripening. CpAGL18 bound and activated the promoter activities of genes in the ethylene and auxin signaling pathways, including CpERS1, CpERF224.4, CpEIN3, CpWAT1, CpIAA11 and CpIAA16-like, which were validated by the DLR, Y1H, EMSA, and ChIP-qPCR assays. CpAGL18 interacts with CpEBF1/2, and the interaction promoted the activities of CpERS1, CpERF22.4.4, CpIAA16 and CpWAT1. Additionally, it was demonstrated that CpEBF1/2 had no ubiquitination effect on CpAGL18. These data showed that CpAGL18 might act as a regulator in integrating ethylene and auxin signaling to regulate fruit ripening. The unsuitable 1-MCP severely impaired the ethylene signaling and regeneration of the new ethylene receptor of fruit by inhibiting CpAGL18 expression, possibly accounting for the ripening disorder formation.

Identification of quantitative trait loci and candidate genes for grain superoxide dismutase activity in wheat

BackgroundSuperoxide dismutase (SOD) can greatly scavenge reactive oxygen species (ROS) in plants. SOD activity is highly related to plant stress tolerance that can be improved by overexpression of SOD genes. Identification of SOD activity-related loci and potential candidate genes is essential for improvement of grain quality in wheat breeding. However, the loci and candidate genes for relating SOD in wheat grains are largely unknown. In the present study, grain SOD activities of 309 recombinant inbred lines (RILs) derived from the ‘Berkut’ × ‘Worrakatta’ cross were assayed by photoreduction method with nitro-blue tetrazolium (NBT) in four environments. Quantitative trait loci (QTL) of SOD activity were identified using inclusive composite interval mapping (ICIM) with the genotypic data of 50 K single nucleotide polymorphism (SNP) array.ResultsSix QTL for SOD activity were mapped on chromosomes 1BL, 4DS, 5AL (2), and 5DL (2), respectively, explaining 2.2 ~ 7.4% of the phenotypic variances. Moreover, QSOD.xjau-1BL, QSOD.xjau-4DS, QSOD.xjau-5 A.1, QSOD.xjau-5 A.2, and QSOD.xjau-5DL.2 identified are likely to be new loci for SOD activity. Four candidate genes TraesCS4D01G059500, TraesCS5A01G371600, TraesCS5D01G299900, TraesCS5D01G343100LC, were identified for QSOD.xjau-4DS, QSOD.xjau-5AL.1, and QSOD.xjau-5DL.1 (2), respectively, including three SOD genes and a gene associated with SOD activity. Based on genetic effect analysis, this can be used to identify desirable alleles and excellent allele variations in wheat cultivars.ConclusionThese candidate genes are annotated for promoting SOD production and inhibiting the accumulation of ROS during plant growth. Therefore, lines with high SOD activity identified in this study may be preferred for future wheat breeding.

Small Interfering RNAs (siRNAs) Negatively Impact Growth and Gene Expression of Environmentally Relevant Bacteria in In Vitro Conditions.

Rising global populations have amplified food scarcity and ushered in the development of genetically modified (GM) crops containing small interference RNAs (siRNAs) that control gene expression to overcome these challenges. The use of RNA interference (RNAi) in agriculture remains controversial due to uncertainty regarding the unintended release of genetic material and downstream nontarget effects, which have not been assessed in environmental bacteria to date. To evaluate the impacts of siRNAs used in agriculture on environmental bacteria, this study assessed microbial growth and viability as well as transcription activity with and without the presence of environmental stressors. Results showed a statistically significant reduction in growth capacity and maximum biomass achieved when bacteria are exposed to siRNAs alone and with additional external stress (p < 0.05). Further transcriptomic analysis demonstrated that nutrient cycling gene activities were found to be consistently and significantly altered following siRNA exposure, particularly among carbon (xylA, FBPase, limEH, Chitinase, rgl, rgh, rgaE, mannanase, ara) and nitrogen (ureC, nasA, narB, narG, nirK) cycling genes (p < 0.05). Decreases in carbon cycling gene transcription profiles were generally significantly enhanced when siRNA exposure was coupled with nutrient or antimicrobial stress. Collectively, findings suggest that certain conditions facilitate the uptake of siRNAs from their surrounding environments that can negatively affect bacterial growth and gene expression activity, with uncertain downstream impacts on ecosystem homeostasis.

Generation and characterization of a depigmented variant of Chlorella pyrenoidosa for application in food products

Microalgae are promising natural resources for an array of essential nutrients and health-promoting compounds. The past decade has witnessed their increasing popularity as a value-added ingredient in the food industry. However, some of their intrinsic properties, especially their typical green color and off-flavor, may compromise consumer acceptability and remain a key challenge for food scientists. This study aimed to generate and characterize a chemically induced pigment-deficient mutant (C49) of Chlorella pyrenoidosa. In contrast to the wild type (WT), C49 could not grow photoautotrophically or mixotrophically. However, when grown heterotrophically, it exhibited a growth curve similar to that of WT. Pigment analysis verified the presence of only trace amounts of chlorophylls (0.27 ± 0.02 mg/g) and carotenoids (0.036 ± 0.003 mg/g) in the C49 which were only ∼1.7% and ∼1% of those in the WT, respectively. C49 also had higher carbohydrate (62.75% ± 2.42%) and lower protein (26.54% ± 1.00%) content than the WT (37.41% ± 2.25% and 36.21% ± 1.97%, respectively). Genetic assays revealed inactivation of the phytoene desaturase gene as a key mechanism underlying the reduced xanthophyll-synthesizing capability of C49, thus causing its failure to accumulate chlorophyll and develop chloroplasts. Transcriptomic and RT-PCR analyses indicated that the high carbohydrate content was likely due to reduced starch degradation. Furthermore, C49 demonstrated an emulsifying capacity comparable to that of WT over a wide pH range (3–8) in oil-in-water emulsions consisting of 5% cell-disrupted homogenate in deionized water and rapeseed oil (1:1, v/v). The results of this study support that depigmented Chlorella pyrenoidosa possesses promising potential for application in the food industry.

Inhibition of DNMT1 attenuates experimental food allergy

BackgroundThe treatment of food allergy (FA) needs improvement. The treatment of immune disorders can be improved by regulating epigenetic marks, which is a promising method. The objective of this research is to alleviate experimental FA by employing an inhibitor of DNA methyltransferase-1 (DNMT1). MethodsOvalbumin was used as the specific antigen to establish a mouse model of FA. Intestinal IL-35+ regulatory B cells (Breg cells) were isolated from FA mice, and characterized using immunological approaches. ResultsFA mice had a lower frequency of IL-35+ Breg cells, which was inversely correlated with their FA response. The quantity of IL-35 was lower in intestinal Breg cells from FA mice. Hypermethylation status was detected in the Il35 promoter, which was accompanied with high levels of H3K9me3. Enforced expression of DNMT1 hindered the promoter activity of the IL35 gene. Administration of an inhibitor of DNMT1 (RG108) restored the immune regulatory capacity of FA intestinal Bregs, and effectively suppressed the expression of DNMT1, and attenuated experimental FA. ConclusionsThe elevated quantity of DNMT1 in intestinal Breg cells compromises the expression of IL-35 and affects the immune regulatory functions, which facilitates the development of FA. The immune regulatory functions of intestinal Breg cells are restored and experimental FA is attenuated by inhibiting DNMT1.

SIP-metagenomics reveals key drivers of rhizospheric Benzo[a]pyrene bioremediation via bioaugmentation with indigenous soil microbes

Rhizoremediation and bioaugmentation have proven effective in promoting benzo[a]pyrene (BaP) degradation in contaminated soils. However, the mechanism underlying bioaugmented rhizospheric BaP degradation with native microbes is poorly understood. In this study, an indigenous BaP degrader (Stenotrophomonas BaP-1) isolated from petroleum-contaminated soil was introduced into ryegrass rhizosphere to investigate the relationship between indigenous degraders and rhizospheric BaP degradation. Stable isotope probing and 16S rRNA gene amplicon sequencing subsequently revealed 15 BaP degraders, 8 of which were directly associated with BaP degradation including Bradyrhizobium and Streptomyces. Bioaugmentation with strain BaP-1 significantly enhanced rhizospheric BaP degradation and shaped the microbial community structure. A correlation of BaP degraders, BaP degradation efficiency, and functional genes identified active degraders and genes encoding polycyclic aromatic hydrocarbon-ring hydroxylating dioxygenase (PAH-RHD) genes as the primary drivers of rhizospheric BaP degradation. Furthermore, strain BaP-1 was shown to not only engage in BaP metabolism but also to increase the abundance of other BaP degraders and PAH-RHD genes, resulting in enhanced rhizospheric BaP degradation. Metagenomic and correlation analyses indicated a significant positive relationship between glyoxylate and dicarboxylate metabolism and BaP degradation, suggesting a role for these pathways in rhizospheric BaP biodegradation. By identifying BaP degraders and characterizing their metabolic characteristics within intricate microbial communities, our study offers valuable insights into the mechanisms of bioaugmented rhizoremediation with indigenous bacteria for high-molecular-weight PAHs in petroleum-contaminated soils.

Single-Cell Analysis Reveals That CD47 mRNA Expression Correlates with Immune Cell Activation, Antiviral Isgs, and Cytotoxicity.

Immune cells are reported to upregulate CD47 during infection, however, the role of CD47 in innate and adaptive immune cells remains unclear. To bridge this knowledge gap, we analysed our single cell (sc)-RNA dataset along with other publicly available sc-RNA datasets from healthy controls, people with HIV-1 (PWH) and COVID-19 patients. We characterized each immune cell based on low, intermediate, and high expression of CD47 . Our analyses revealed that CD47 high pDCs and monocytes exhibited relatively higher expression of IFN-α regulatory genes, antiviral interferon-stimulated genes (ISGs) and MHC-I associated genes compared to CD47 inter. and CD47 low cells. Furthermore, CD47 high NK and CD8+ T cells showed higher expression of antiviral ISGs, as well as genes encoding for cytotoxic markers like granzyme B, perforin, granulysin, interferon gamma and NKG7. Additionally, CD47 high CD8+ T cells expressed higher levels of PD-1 and LAG-3 genes. Lastly, we found that CD47 high B cells had enriched expression of genes involved in cell activation and humoral responses. Overall, our analyses revealed that innate and adaptive immune cells expressing elevated activation and functional gene signatures also express higher CD47 levels.

CPT2 Deficiency Modeled in Zebrafish: Abnormal Neural Development, Electrical Activity, Behavior, and Schizophrenia-Related Gene Expression.

Carnitine palmitoyltransferase 2 (CPT2) is an inner mitochondrial membrane protein of the carnitine shuttle and is involved in the beta-oxidation of long chain fatty acids. Beta-oxidation provides an alternative pathway of energy production during early development and starvation. CPT2 deficiency is a genetic disorder that we recently showed can be associated with schizophrenia. We hypothesize that CPT2 deficiency during early brain development causes transcriptional, structural, and functional abnormalities that may contribute to a CNS environment that is susceptible to the emergence of schizophrenia. To investigate the effect of CPT2 deficiency on early vertebrate development and brain function, CPT2 was knocked down in a zebrafish model system. CPT2 knockdown resulted in abnormal lipid utilization and deposition, reduction in body size, and abnormal brain development. Axonal projections, neurotransmitter synthesis, electrical hyperactivity, and swimming behavior were disrupted in CPT2 knockdown zebrafish. RT-qPCR analyses showed significant increases in the expression of schizophrenia-associated genes in CPT2 knockdown compared to control zebrafish. Taken together, these data demonstrate that zebrafish are a useful model for studying the importance of beta-oxidation for early vertebrate development and brain function. This study also presents novel findings linking CPT2 deficiency to the regulation of schizophrenia and neurodegenerative disease-associated genes.

MiR-10167-3p targets TCF7L1 to inhibit bovine adipocyte differentiation and promote bovine adipocyte proliferation

MicroRNAs (miRNAs) are widely involved in various lipogenic processes, including adipocyte proliferation and differentiation, lipid droplet formation, and adipocyte-specific gene activation. The present study aimed to investigate the gene expression profiles of bovine preadipocytes under high miR-10167-3p expression using the RNA-seq technique and to verify the functions of its downstream target genes on the proliferation and differentiation of bovine preadipocytes. First, RNA-seq identified 573 differentially expressed genes (DEGs), of which 243 were downregulated and 330 were upregulated. Then, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that 15.19% of the DEGs were enriched in pathways related to lipid metabolism. Meanwhile, dual-luciferase reporter gene assay verified the target-binding relationship between miR-10167-3p and TCF7L1. The function of TCF7L1 was assessed using several experiments in adipocytes with high TCF7L1 expression and RNA interference. The mRNA and protein expression of proliferation, differentiation, and apoptosis marker genes were detected using qPCR and western blot, respectively; lipid droplet synthesis was detected using oil red O, Nile red, and bodipy staining; adipocyte proliferation was detected by EdU; and apoptosis was detected using flow cytometry. The results revealed that TCF7L1 overexpression inhibited bovine preadipocyte differentiation and apoptosis and promoted their proliferation, with opposite results obtained with its RNA interference. These results may provide a reference for the subsequent investigation of the molecular mechanism of bovine fat deposition.

Clear Cell Renal Cell Carcinoma: A Comprehensive Review of its Histopathology, Genetics, and Differential Diagnosis.

Clear cell renal cell carcinoma (ccRCC) is the predominant subtype of renal epithelial tumor, accounting for roughly 2% of all malignancies. Clinically, it often presents in the sixth to seventh decade of life, predominantly in men. Pathologically, these tumors exhibit a distinctive golden yellow cut surface, usually arising from the renal cortex. Their microscopic features are characterized by solid and nested architectures of cells with clear or eosinophilic granular cytoplasm and a prominent vascular network. A hallmark genetic feature is the inactivation of the VHL gene situated on chromosome 3p25. The majority of ccRCCs are sporadic (over 95%), typically presenting as a single mass; and a small percentage have a hereditary basis, often associated with VHL disease, characterized by multiple bilateral tumors with an earlier onset. Immunohistochemically, ccRCC tumors express PAX8, CA9 box like pattern, and CD10 but are generally negative for AMACR (35% positive) and KRT7 (15% positive). The prognosis of ccRCC is largely determined by its TNM stage, ISUP/WHO nucleolar grade, and the presence of specific aggressive features. This review article delves into the detailed gross, microscopic, molecular, and clinical features of ccRCC, offering comprehensive insights into its diagnosis, management, and prognosis.

An insight into the role of branched-chain α-keto acid dehydrogenase (BKD) complex in branched-chain fatty acid biosynthesis and virulence of Listeria monocytogenes.

Listeria monocytogenes is a foodborne bacterial pathogen that causes listeriosis. Positive regulatory factor A (PrfA) is a pleiotropic master activator of virulence genes of L. monocytogenes that becomes active upon the entry of the bacterium into the cytosol of infected cells. L. monocytogenes can survive and multiply at low temperatures; this is accomplished through the maintenance of appropriate membrane fluidity via branched-chain fatty acid (BCFA) synthesis. Branched-chain α-keto acid dehydrogenase (BKD), which is composed of four polypeptides encoded by lpd, bkdA1, bkdA2, and bkdB, is known to play a vital role in BCFA biosynthesis. Here, we constructed BKD-deficient Listeria strains by in-frame deletion of lpd, bkdA1, bkdA2, and bkdB genes. To determine the role in in vivo and in vitro, mouse model challenges, plaque assay in murine L2 fibroblast, and intracellular replication in J744A.1 macrophage were conducted. BKD-deficient strains exhibited defects in BCFA composition, virulence, and PrfA-regulon function within the host cells. Transcriptomics analysis revealed that the transcript level of the PrfA-regulon was lower in ΔbkdA1 strain than those in the wild-type. This study demonstrates that L. monocytogenes strains lacking BKD complex components were defective in PrfA-regulon function, and full activation of wild-type prfA may not occur within host cells in the absence of BKD. Further study will investigate the consequences of BKD deletion on PrfA function through altering BCFA catabolism.IMPORTANCEListeria monocytogenes is the causative agent of listeriosis, a disease with a high mortality rate. In this study, we have shown that the deletion of BKD can impact the function of PrfA and the PrfA-regulon. The production of virulence proteins within host cells is necessary for L. monocytogenes to promote its intracellular survival and is likely dependent on membrane integrity. We thus report a link between L. monocytogenes membrane integrity and the function of PrfA. This knowledge will increase our understanding of L. monocytogenes pathogenesis, which may provide insight into the development of antimicrobial agents.

Integrated single-cell and bulk RNA sequencing identifies POSTN as a potential biomarker and therapeutic target for rheumatoid arthritis

BackgroundThis study aimed to integrate single-cell RNA sequencing (scRNA-seq) and bulk RNA-seq data to identify potential biomarkers and therapeutic targets for rheumatoid arthritis (RA). MethodFirstly, we obtained the synovial scRNA-seq data from the Immport database and bulk RNA-seq data from the Gene Expression Omnibus (GEO) database. Then, we used weighted gene correlation network analysis (WGCNA) to screen for module genes most relevant to RA and intersected them with the differentially expressed genes (DEGs) obtained from scRNA-seq and bulk RNA-seq to obtain intersecting genes. Next, we constructed a protein–protein interaction (PPI) network of hub genes using the STRING database and Cytoscape software and validated its expression using external validation cohorts. Finally, we performed immune cell infiltration analysis using CIBERSORT and explored the expression and drug binding activity of key gene using clinical samples and molecular docking, respectively. ResultWe identified six cellular subgroups through dimensionality reduction and clustering, and fibroblasts may be the most important cell cluster in RA based on pseudotime and cell–cell communication analyses. Subsequently, we intersected module genes with DEGs obtained from scRNA-seq and bulk RNA-seq and constructed a PPI network of hub genes (BGN, COL11A1, COL1A1, GUCY1A1, POSTN). In external validation cohorts, POSTN was highly expressed and demonstrated the highest diagnostic performance (AUC = 0.716). In subsequent analyses, we defined POSTN as a key gene and found that its expression level was positively correlated with M2 macrophages in immune cell infiltration analysis. Additionally, POSTN was upregulated in clinical samples and exhibited favorable binding activity with nine anti-rheumatoid arthritis drugs (affinity ≤ −6.0 kcal/mol). ConclusionThrough bioinformatics analysis, clinical sample validation, and molecular docking, we found that POSTN was highly expressed in RA and stably bound to common anti-rheumatoid arthritis drugs, which will provide new insights into potential biomarkers and therapeutic targets for RA.

Engineering of RNase P Ribozymes for Therapy against Human Cytomegalovirus Infection.

Nucleic acid-based gene interference and editing strategies, such as antisense oligonucleotides, ribozymes, RNA interference (RNAi), and CRISPR/Cas9 coupled with guide RNAs, are exciting research tools and show great promise for clinical applications in treating various illnesses. RNase P ribozymes have been engineered for therapeutic applications against human viruses such as human cytomegalovirus (HCMV). M1 ribozyme, the catalytic RNA subunit of RNase P from Escherichia coli, can be converted into a sequence-specific endonuclease, M1GS ribozyme, which is capable of hydrolyzing an mRNA target base-pairing with the guide sequence. M1GS RNAs have been shown to hydrolyze essential HCMV mRNAs and block viral progeny production in virus-infected cell cultures. Furthermore, RNase P ribozyme variants with enhanced hydrolyzing activity can be generated by employing in vitro selection procedures and exhibit better ability in suppressing HCMV gene expression and replication in cultured cells. Additional studies have also examined the antiviral activity of RNase P ribozymes in mice in vivo. Using cytomegalovirus infection as an example, this review summarizes the principles underlying RNase P ribozyme-mediated gene inactivation, presents recent progress in engineering RNase P ribozymes for applications in vitro and in mice, and discusses the prospects of using M1GS technology for therapeutic applications against HCMV as well as other pathogenic viruses.

Apelin modulates inflammation and leukocyte recruitment in experimental autoimmune encephalomyelitis

Demyelination due to autoreactive T cells and inflammation in the central nervous system are principal features of multiple sclerosis (MS), a chronic and highly disabling human disease affecting brain and spinal cord. Here, we show that treatment with apelin, a secreted peptide ligand for the G protein-coupled receptor APJ/Aplnr, is protective in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Apelin reduces immune cell entry into the brain, delays the onset and reduces the severity of EAE. Apelin affects the trafficking of leukocytes through the lung by modulating the expression of cell adhesion molecules that mediate leukocyte recruitment. In addition, apelin induces the internalization and desensitization of its receptor in endothelial cells (ECs). Accordingly, protection against EAE major outcomes of apelin treatment are phenocopied by loss of APJ/Aplnr function, achieved by EC-specific gene inactivation in mice or knockdown experiments in cultured primary endothelial cells. Our findings highlight the importance of the lung-brain axis in neuroinflammation and indicate that apelin targets the transendothelial migration of immune cells into the lung during acute inflammation.