Variation In Expression Levels Research Articles

The potential of ESCO2 as a prognostic and immunotherapeutic marker of pan-cancer and its role in anti-PD1 treatment of bladder cancer

Background Establishment of sister chromatid cohesion N-acetyltransferase 2 (ESCO2), a member of the EFO2 family, is implicated in the pathogenesis and progression of various cancers. However, there has been limited comprehensive pan-cancer analysis conducted on ESCO2 thus far. Methods Publicly available databases, such as the UCSC Xena database, were utilized to examine differential expression patterns across various cancer types. In addition, variations in expression levels were investigated across distinct clinical stages. Univariate Cox regression and Kaplan-Meier survival analyses were conducted to evaluate the impact on overall survival (OS), disease-specific survival (DSS), disease-free interval (DFI), and progression-free interval (PFI) at a pan-cancer level. The correlation between ESCO2 expression and immune cell infiltration was examined to gain insight into the tumor microenvironment in different cancers. The results of the bioinformatic analysis were validated using immunotherapy clinical trials and pathological specimens. CCK-8 and transwell assay experiments were performed to investigate the biological function of ESCO2. Results ESCO2 expression was found to be up-regulated in most cancers, with a correlation to TNM stages. Prognostic analysis indicated that overexpression of ESCO2 was associated with poor prognosis in various cancers. Furthermore, the correlation between ESCO2 expression and immune cell infiltration suggested its potential as a predictor for immunotherapy efficacy. Notably, ESCO2 expression showed positive associations with immunoinhibitor, immnostimulator, MHC molecule, chemokine receptor, tumor mutation burden (TMB) and microsatellite instability (MSI) levels in bladder cancer (BLCA). The validation cohort for immunotherapy corroborated these findings and substantiated that ESCO2 could function as an autonomous prognostic biomarker and a promising target for cancer treatment via immunotherapy. In addition, in vitro experiments confirmed the role of ESCO2 in influcing the proliferation, invasion and migration of BLCA cells. Conclusion ESCO2 participates in regulating the immune infiltration and affecting the prognosis of patients in many cancers, especially in BLCA. ESCO2 may serve as a prognostic and immunotherapy biomarker in future treatment of human cancer.

SPP1 is a plasma biomarker associated with the dia gnosis and prediction of prognosis in sepsis

In this study, peripheral whole blood samples from 22 hospitalized patients and 10 healthy individuals were analyzed using a combination of Data Independent Acquisition (DIA) and Enzyme-Linked Immunosorbent Assay (ELISA) techniques to identify differentially expressed proteins (DEPs) in sepsis patients’ plasma. The aim was to provide accurate and detailed biomarkers, such as SPP1, for determining the pathological stages of sepsis. SPP1, known as osteopontin1 is a pleiotropic protein with a wide distribution and multifunctional effects. Its protein expression is associated with inflammatory changes, including variations in expression levels in infectious diseases, allergic diseases, and situations involving tissue damage. The registration number was ChiCTR1900021261.The full date of first registration year is 2018. In the affiliated hospital of southwest medical university, 22 sepsis patients were hospitalized from January 2019 to September 2020 and 10 normal healthy individuals were selected for DIA-based quantitative proteomics analysis. In addition to gene ontology analysis and Kyoto Encyclopedia of genes and genomes analysis, enrichment analysis of data was performed and target protein network was screened through joint protein–protein interaction and visualization techniques. The selected protein targets were then validated by Elisa kit. The software was used to analyze the differences comparing the control group to the sepsis group and the sepsis group, as well as between sepsis survivals and non-survivals, and a ROC curve was drawn to evaluate the diagnostic value and prognostic effect of the method of the corresponding target proteins. A total of 174 DEPs were screened by bioinformatics analysis. An analysis of go pathway enrichment revealed the following: These proteins were mainly involved in biological processes among them are the inflammation response, the metabolism of extracellular matrix, the secretion of cell secretions, the activation of cells, and the immune response. According to the Kegg pathway analysis, they were mainly involved in complement cascade polymerization, extracellular protease and glycosylase activation, protein synthesis process, biotin metabolism, leukocyte transmembrane migration, bacterial infection and phagosome formation. SPP1 was identified as a possible plasma biomarker and was therefore further validated using Elisa. As a result of experiments, it has been demonstrated that level in sepsis patients is significantly compared to the normal control group and the level is also higher in non-survivals of sepsis. The ROC curve can be used to see that it can diagnose sepsis more accurately and improve prognostic ability prediction. Cell experiments confirm that SPP1 is highly expressed in sepsis. There is a significant difference in the levels of SPP1 protein between the normal group and the sepsis group; it not only has good diagnostic significance for sepsis, but also provides corresponding reference value for patient prognosis; Therefore, it is more likely to become a biological marker of sepsis over time.

Comparison of Different Keratinocyte Cell Line Models for Analysis of NLRP1 Inflammasome Activation

The NLRP1 (nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-1) inflammasome is the most important inflammasome in human keratinocytes. It plays a crucial role in regulating innate immunity in the skin. This study aimed to evaluate NLRP1 inflammasome activation and the corresponding levels of detection in different keratinocyte cell lines to identify a suitable in vitro model for analyzing inflammasome activation in keratinocytes. We compared NLRP1 inflammasome activation, expression, and cell death among primary keratinocytes and immortalized keratinocyte cell lines HaCaT, HaSKpw, and SVTERT upon stimulation with ultraviolet B (UVB) irradiation or talabostat. The effects of both NLRP1 inducers on cell death and the modification of NLRP1 molecules were examined using fluorescence-activated cell sorting analysis, Western blotting, and an enzyme-linked immunosorbent assay. The key inflammasome components had varied expression levels among the keratinocyte cell models, with the highest expression observed in primary keratinocytes. Moreover, our data showed that both UVB and talabostat triggered cell death, and NLRP1 inflammasome activation was readily detected in primary keratinocytes but not in the analyzed immortalized keratinocyte cell lines. Therefore, we do not recommend the use of the immortalized keratinocyte cell lines HaCaT, HaSKpw, and SVTERT for analyzing inflammasome activation in keratinocytes; we strongly recommend the use of primary keratinocytes for these studies.

Characterization of the function and clinical value of ERCC family genes in lung adenocarcinoma

IntroductionERCC genes, responsible for encoding enzymes involved in base excision repair, have been implicated in various cancers, contributing to chemoresistance. However, a comprehensive analysis of the prognostic and therapeutic significance of this gene family in lung adenocarcinoma (LUAD) is lacking.MethodsThis study conducted a multidimensional assessment of ERCC family genes in LUAD using bioinformatic approaches, including mRNA expression level, gene methylation, and copy number variation (CNV), as well as their correlations with clinical outcome, gene set variations, and tumor-infiltrating lymphocytes (TILs). In addition, We evaluated the anti-tumor effects of ERCC8 in cell lines, demonstrating its clinical potential on an experimental level.ResultsOverall, the expression of ERCC genes exhibited a negative correlation with good prognosis, with ERCC6L and ERCC8 demonstrating the most reliable predictive performance. Gene methylation level and CNV increases of ERCC genes generally displayed negative and positive associations with their expression levels, respectively. Additionally, GSVA analysis suggested that ERCC expression was positively correlated with cell cycle and apoptosis pathways but negatively correlated to the TSC/mTOR pathway. Furthermore, the expression of ERCC genes exhibited a complex relationship with TILs and the response to anti-tumor drugs. The results of in vitro cellular experiments show that inhibiting ERCC8 can alleviate the malignant phenotype of LUAD cells.DiscussionOur study revealed the multifaceted biological and clinical significance of ERCC family members in LUAD. These findings provide new insights into the function of ERCC family genes in LUAD and their potential clinical applications.

Profiling Phenotypic Heterogeneity of Circulating Tumor Cells through Spatially Resolved Immunocapture on Nanoporous Micropillar Arrays.

The phenotype of circulating tumor cells (CTCs) offers valuable insights into monitoring cancer metastasis and recurrence. While microfluidics presents a promising approach for capturing these rare cells in blood, the phenotypic profiling of CTCs remains technically challenging. Herein, we developed a nanoporous micropillar array chip enabling highly efficient capture and in situ phenotypic analysis of CTCs through enhanced and tunable on-chip immunoaffinity binding. The nanoporous micropillar array addresses the fundamental limits in fluidic mass transfer, surface stagnant flow boundary effect, and interface topographic and multivalent reactions simultaneously within a single device, resulting in a synergistic enhancement of CTC immunocapture efficiency. The CTC capture efficiency increased by approximately 40% for cancer cells with low surface marker expressing. By manipulating fluidic velocity (hydrodynamic drag force) on the chip, a cell adhesion gradient was generated in the capture chamber, enabling individual CTCs with varying expression levels of epithelial cellular adhesion molecules to be immunocaptured at the corresponding spatial locations where equilibrium drag force is provided. The clinical utility of the nanoporous micropillar array was demonstrated by accurately distinguishing early and advanced stages of breast cancer and further longitudinally monitoring treatment response. We envision that the nanoporous micropillar array chip will provide an in situ capture and molecular profiling approach for CTCs and enhance the clinical application of CTC liquid biopsy.

Five-gene prognostic model based on autophagy-dependent cell death for predicting prognosis in lung adenocarcinoma

Non-small cell lung adenocarcinoma (LUAD) is the predominant form of lung cancer originating from lung epithelial cells, making it the most prevalent pathological type. Currently, reliable indicators for predicting treatment efficacy and disease prognosis are lacking. Despite extensive validation of autophagy-dependent cell death (ADCD) in solid tumor studies and its correlation with immunotherapy effectiveness and cancer prognosis, systematic research on ADCD-related genes in LUAD is limited. We utilized AddModuleScore, ssGSEA, and WGCNA to identify genes associated with ADCD across single-cell and bulk transcriptome datasets. The TCGA dataset, comprising 598 cases, was randomly divided into training and validation sets to develop an ADCD-related LUAD prediction model. Internal validation was performed using the TCGA validation set. For external validation, datasets GSE13213 (119 LUAD samples), GSE26939 (115 LUAD samples), GSE29016 (39 LUAD samples), and GSE30219 (86 LUAD samples) were employed. We evaluated the model’s accuracy and effectiveness in predicting prognostic risk. Additionally, CIBERSORT, ESTIMATE, and ssGSEA techniques were used to explore immunological characteristics, drug response, and gene expression in LUAD. Real-time RT-PCR was conducted to assess variations in mRNA expression levels of the gene XCR1 between cancerous and normal tissues in 10 lung cancer patients. We identified 249 genes associated with autophagy-dependent cell death (ADCD) at both single-cell and bulk transcriptome levels. Univariate COX regression analysis revealed that 18 genes were significantly associated with overall survival (OS). Using LASSO-Cox analysis, we developed an ADCD signature based on five genes (BIRC3, TAP1, SLAMF1, XCR1, and HLA-DMB) and created the ADCD-related risk scoring system (ADCDRS). Validation of this model demonstrated its ability to predict disease prognosis and its correlation with clinical characteristics, immune cell infiltration, and the tumor microenvironment. To enhance clinical applicability, we integrated an ADCDRS nomogram. Furthermore, we identified potential drugs targeting specific risk subgroups. We successfully identified a model based on five ADCD genes to predict disease prognosis and treatment efficacy in LUAD, as well as to assess the tumor immune microenvironment. An efficient and practical ADCDRS nomogram was designed.

Early apoptosis detection in canine granulosa cells through the analysis of BCL-2 and BAX proteins during the follicular development associated with oocyte maturation

The objective of this study was to investigate the early follicular apoptosis in canine ovarian follicles by examining the expression of anti-apoptotic BCL-2 and pro-apoptotic BAX proteins throughout the estrous cycle associated with oocyte maturation. Follicular cells from preantral and antral follicles of varying sizes were isolated and grouped based on follicle type and estrous phase. Antral follicles underwent flow cytometry analysis, whereas preantral follicles were subjected to Western blotting. The meiotic capacity of oocytes from these different follicle types was evaluated through in vitro maturation. Statistical analysis included ANOVA and Duncan’s test. Results showed fluctuations in BCL-2 and BAX levels across different follicular stages and estrous phases. BCL-2 levels increased (P<0.05) with follicular development in antral follicles, particularly during estrus, while BAX exhibited variations peaking (P<0.05) during estrus. The BCL-2/BAX ratio in antral follicles was higher (P<0.05) in estrus and diestrus compared to anestrus and proestrus. Additionally, BCL-2 and BAX proteins were detected in preantral follicles, with varying expression levels (P<0.05) across estrous phases. The BCL-2/BAX ratio in preantral follicles was highest (P<0.05) during anestrus and estrus and decreased (P<0.05) in proestrus and diestrus. Oocytes from preantral follicles did not reach the MII stage, regardless of the levels of BCL-2/BAX. Conversely, oocytes obtained from large follicles during estrus showed the highest (P<0.05) maturation percentages, which were associated with the highest BCL-2/BAX ratio. These findings provide insights into the dynamic patterns of BCL-2 /BAX in canine follicles across the estrous cycle, shedding light on their potential roles in oocyte development.

The combination of FLCWK with 5-FU inhibits colon cancer and multidrug resistance by activating PXR to suppress the IL-6/STAT3 pathway.

5-fluorouracil (5-FU) is a preferred chemotherapeutic agent for the treatment of colon cancer. Nonetheless, its clinical effectiveness is frequently hampered by suboptimal therapeutic outcomes and the emergence of drug resistance. Therefore, there exists a pressing demand for novel therapeutic agents to circumvent chemoresistance. The pregnane X receptor (PXR) exerts a pivotal regulatory influence on the proliferation, invasion, and chemoresistance mechanisms in colon cancer. Activation of PXR drives up the transcription of the multidrug resistance gene (MDR1), thus prompting the expression of P-glycoprotein (P-gp) responsible for conferring tumour resistance. This study scrutinized the potential of Fengliao Changweikang (FLCWK) in augmenting the efficacy of 5-FU in the management of colon cancer. To this end, we engineered colon cancer cells with varied levels of PXR expression via lentiviral transfection, subsequently validating the findings in nude mice. By means of MTT assays, flow cytometry apoptosis analysis, Western blotting and immunofluorescence, we probed into the prospective impacts of FLCWK and 5-FU on cellular viability and resistance. Our results revealed that while upregulation of PXR amplified the therapeutic benefits in colon cancer treatment, it concurrently heightened resistance levels. FLCWK demonstrated a capacity to reduce P-gp expression, with the combined administration of FLCWK and 5-FU effectively reversing resistance mechanisms. Furthermore, activation of PXR was found to impede the IL-6/STAT3 signalling pathway. In an effort to mimic the development of colon cancer, we established an azomethane oxide (AOM)/ dextran sodium sulfate (DSS) mouse model, showing that FLCWK bolstered the inhibitory effects of 5-FU, impeding the progression of colon cancer. In summation, our findings point towards the potential of FLCWK in the treatment of colon cancer, particularly in strengthening the therapeutic efficacy of 5-FU in the prevention and control of the disease.

MIKC*-type MADS transcription factors control JINGUBANG expression and the degree of pollen dormancy in Arabidopsis.

While pollen dormancy has been proposed to play a necessary role in sexual reproduction, it remains poorly understood. Here, we used traditional pollen germination assays to characterize dormancy. Our results underscore variation in the degree of dormancy between individual pollen grains. In addition, we provide evidence that JINGUBANG (JGB), previously defined as a negative regulator of pollen germination in Arabidopsis (Arabidopsis thaliana), is responsible for the uneven degrees of pollen dormancy, as asynchronous pollen germination in vitro reflected varied expression levels of JGB. We identified five cis-acting elements, including four CArG-boxes and the previously uncharacterized element ERE7, as essential for the initiation and enhancement of JGB expression. A 10-bp sequence between CArG-box 3 and ERE7, likely the result of an inverse DNA loop formed between CArG-box 3 and CArG-box 4, was required for robust gene expression. In addition, the pollen-specific AtMIKC*-type MADS transcription factors AGAMOUS-LIKE 30 (AGL30), AGL65, AGL66, AGL94, and AGL104 activated JGB transcription. Notably, the transactivation levels differed among the obligate AtMIKC* heterodimers tested. Our results indicate that distinct AtMIKC* complexes formed in individual pollen grains direct pollen dormancy to uneven degrees, which is likely an adaptive trait that ensures broader pollen dispersal under adverse environmental conditions.

Genome-Wide Identification and Analysis of Phospholipase C Gene Family Reveals Orthologs, Co-Expression Networks, and Expression Profiling Under Abiotic Stress in Sorghum bicolor

Phospholipase C (PLC) is an essential enzyme involved in lipid signaling pathways crucial for regulating plant growth and responding to environmental stress. In sorghum, 11 PLC genes have been identified, comprising 6 PI-PLCs and 5 NPCs. Through phylogenetic and interspecies collinearity analyses, structural similarities between SbPLCs and ZmPLCs proteins have been observed, with a particularly strong collinearity between SbPLCs and OsPLCs. Promoter function analysis has shown that SbPLCs are significantly enriched under abiotic stress and hormonal stimuli, like ABA, jasmonic acid, drought, high temperature, and salt. Gene co-expression networks, constructed using a weighted gene co-expression network analysis (WGCNA), highlight distinct expression patterns of SbPLC1, SbPLC3a, and SbPLC4 in response to abiotic stress, providing further insights into the expression patterns and interactions of SbPLCs under various environmental stimuli. qRT-PCR results reveal variations in expression levels among most SbPLCs members under different stress conditions (drought, NaCl, NaHCO3), hormone treatments (ABA), and developmental stages, indicating both specific and overlapping expression patterns. This comprehensive analysis offers valuable insights into the roles of SbPLCs in sorghum, shedding light on their specific expression patterns, regulatory elements, and protein interactions across different environmental stimuli and developmental stages.

Evolutionary and Expression Analyses of the bZIP Family in Tea Plants (Camellia sinensis) and Functional Characterization of CsbZIP3/42/6 in Response to Environmental Stresses.

Basic leucine zipper (bZIP) transcription factors play crucial roles in various biological processes and responses to environmental stresses. However, the functions of the bZIP family in tea plants remain largely unexplored. Here, we identified 74 bZIP genes in tea plants (Camellia sinensis) and classified them into 12 phylogenetic groups, supported by analyses of conserved motifs and gene structures. Cis-element analysis provided insights into the potential roles of CsbZIP genes in phytohormone signaling and stress responses. Tissue-specific expression analysis demonstrated differential expression profiles of CsbZIP genes, suggesting their tissue- and stage-specific functions. Additionally, varying expression levels under different abiotic stresses indicated functional divergence of the CsbZIP family during the long-term evolution. Notably, CsbZIP3/42/6 were identified as positive regulators of drought and salt stress responses but negative regulators in response to pathogen infection, and CsbZIP42 could interact with CsbZIP3 and CsbZIP6 in regulating these environmental stresses. This study provides valuable information on potential applications for improving stress tolerance and overall plant health of tea plants.

Expression, regulation and physiological roles of the five Rsm proteins in Pseudomonas syringae pv. tomato DC3000

Proteins belonging to the RsmA (regulator of secondary metabolism)/CsrA (carbon storage regulator) family are small RNA-binding proteins that play crucial roles post-transcriptionally regulating gene expression in many Gram-negative and some Gram-positive bacteria. Although most of the bacteria studied have a single RsmA/CsrA gene, Pseudomonas syringae pv. tomato (Pto) DC3000 encodes five Rsm proteins: RsmA/CsrA2, RsmC/CsrA1, RsmD/CsrA4, RsmE/CsrA3, and RsmH/CsrA5. This work aims to provide a comprehensive analysis of the expression of these five rsm protein-encoding genes, elucidate the regulatory mechanisms governing their expression, as well as the physiological relevance of each variant. To achieve this, we examined the expression of rsmA, rsmE, rsmC, rsmD, and rsmH within their genetic contexts, identified their promoter regions, and assessed the impact of both their deletion and overexpression on various Pto DC3000 phenotypes. A novel finding is that rsmA and rsmC are part of an operon with the upstream genes, whereas rsmH seems to be co-transcribed with two downstream genes. We also observed significant variability in expression levels and RpoS dependence among the five rsm paralogs. Thus, despite the extensive repertoire of rsm genes in Pto DC3000, only rsmA, rsmE and rsmH were significantly expressed under all tested conditions (swarming, minimal and T3SS-inducing liquid media). Among these, RsmE and RsmA were corroborated as the most important paralogs at the functional level, whereas RsmH played a minor role in regulating free life and plant-associated phenotypes. Conversely, RsmC and RsmD did not seem to be functional under the conditions tested.

Revealing disease subtypes and heterogeneity in common variable immunodeficiency through transcriptomic analysis

Common Variable Immunodeficiency (CVID) is a primary immunodeficiency characterized by reduced levels of specific immunoglobulins, resulting in frequent infections, autoimmune disorders, increased cancer risk, and diminished antibody production despite an adequate B cell count. With its clinical manifestations being highly variable, the classification of CVID, including the widely recognized Freiburg classification, is primarily based on clinical symptoms and genetic variations. Our study aims to refine the classification of CVID by analyzing transcriptomics data to identify distinct disease subtypes. We utilized the GSE51405 dataset, examining transcriptomic profiles from 30 CVID patients without complications. Employing a combination of clustering techniques—KMeans, hierarchical agglomerative clustering, spectral clustering, and Gaussian Mixture models—and differential gene expression analysis with R’s limma package, we integrated molecular findings with demographic data (age and gender) through correlation analysis and identified common genes among clusters. Three distinct clusters of CVID patients were identified using KMeans, Agglomerative Clustering, and Gaussian Mixture Models, highlighting the disease’s heterogeneity. Differential expression analysis unveiled 31 genes with variable expression levels across these clusters. Notably, nine genes (EIF5A, RPL21, ANP32A, DTX3L, NCF2, CDC42EP3, CHP1, FOLR3, and DEFA4) exhibited consistent differential expression across all clusters, independent of demographic factors. The study recommends categorizing patients based on the four genes, NCF2, CHP1, FOLR3, and DEFA4—as they may assist in prognostic prediction. Transcriptomic analysis of common variable immunodeficiency (CVID) patients identified three distinct clusters based on gene expression, independent of age and gender. Nine differentially expressed genes were identified across these clusters, suggesting potential biomarkers for CVID subtype classification. These findings highlight the genetic heterogeneity of CVID and provide novel insights into disease classification and potential personalized treatment approaches.

Deciphering the ghost proteome in ovarian cancer cells by deep proteogenomic characterization

Proteogenomics is becoming a powerful tool in personalized medicine by linking genomics, transcriptomics and mass spectrometry (MS)-based proteomics. Due to increasing evidence of alternative open reading frame-encoded proteins (AltProts), proteogenomics has a high potential to unravel the characteristics, variants, expression levels of the alternative proteome, in addition to already annotated proteins (RefProts). To obtain a broader view of the proteome of ovarian cancer cells compared to ovarian epithelial cells, cell-specific total RNA-sequencing profiles and customized protein databases were generated. In total, 128 RefProts and 30 AltProts were identified exclusively in SKOV-3 and PEO-4 cells. Among them, an AltProt variant of IP_715944, translated from DHX8, was found mutated (p.Leu44Pro). We show high variation in protein expression levels of RefProts and AltProts in different subcellular compartments. The presence of 117 RefProt and two AltProt variants was described, along with their possible implications in the different physiological/pathological characteristics. To identify the possible involvement of AltProts in cellular processes, cross-linking-MS (XL-MS) was performed in each cell line to identify AltProt-RefProt interactions. This approach revealed an interaction between POLD3 and the AltProt IP_183088, which after molecular docking, was placed between POLD3-POLD2 binding sites, highlighting its possibility of the involvement in DNA replication and repair.

Multiplexed In Situ Imaging of Site-Specific m6A Methylation with Proximity Hybridization Followed by Primer Exchange Amplification (m6A-PHPEA).

Post-transcriptional modification of N6-methyladenosine (m6A) is crucial for ribonucleic acid (RNA) metabolism and cellular function. The ability to visualize site-specific m6A methylation at the single-cell level would markedly enhance our understanding of its pivotal regulatory functions in the field of epitranscriptomics. Despite this, current in situ imaging techniques for site-specific m6A are constrained, posing a significant barrier to epitranscriptomic studies and pathological diagnostics. Capitalizing on the precise targeting capability of deoxyribonucleic acid (DNA) hybridization and the high specificity of the m6A antibody, we present a method, termed proximity hybridization followed by primer exchange amplification (m6A-PHPEA), for the site-specific imaging of m6A methylation within cells. This approach enables high-resolution, single-cell imaging of m6A methylation across various RNA molecules coupled with efficient signal amplification. We successfully imaged three distinct m6A methylation sites concurrently in multiple cell types, revealing cell-to-cell variability in expression levels. This method promises to illuminate the dynamics of m6A-modified RNAs, potentially revolutionizing epitranscriptomic research and the development of advanced pathological diagnosis for chemical modifications.

Genome-Wide Analysis of Aquaporins Gene Family in Populus euphratica and Its Expression Patterns in Response to Drought, Salt Stress, and Phytohormones.

Aquaporins (AQPs) play an essential role in membrane water transport during plant responses to water stresses centered on conventional upstream signals. Phytohormones (PHs) regulate plant growth and yield, working with transcription factors to help plants withstand environmental challenges and regulate physiological and chemical processes. The AQP gene family is important, so researchers have studied its function and regulatory system in numerous species. Yet, there is a critical gap the understanding of many of their molecular features, thus our full knowledge of AQPs is far-off. In this study, we undertook a broad examination of the AQP family gene in Populus euphratica via bioinformatics tools and analyzed the expression patterns of certain members in response to drought, salt, and hormone stress. A total of 22 AQP genes were examined in P. euphratica, and were categorized into four main groups, including TIPs, PIPs, SIPs, and NIPs based on phylogenetic analysis. Comparable exon-intron gene structures were found by gene structure examination, and similarities in motif number and pattern within the same subgroup was determined by motif analysis. The PeuAQP gene family has numerous duplications, and there is a distinct disparity in how the members of the PeuAQP family react to post-translational modifications. Abiotic stress and hormone responses may be mediated by AQPs, as indicated by the abundance of stress response elements found in 22 AQP genes, as revealed by the promoter's cis-elements prediction. Expression pattern analysis reveals that selected six AQP genes from the PIP subgroup were all expressed in the leaves, stem, and roots with varying expression levels. Moreover, qRT-PCR analysis discovered that the majority of the selected AQP members were up- or down-regulated in response to hormone treatment and abiotic stress. Remarkably, PeuAQP14 and PeuAQP15 appeared to be highly responsive to drought stress and PeuAQP15 exhibited a high response to salt stress. The foliar application of the phytohormones (SA, IAA, GA3, MeJA, and ABA) were found to either activate or inhibit PeuAQP, suggesting that they may mitigate the effects of water shortage of poplar water stress. The present work enhances our knowledge of the practical roles of AQPs in stress reactions and offers fundamental information for the AQP genes in poplar species. It also highlights a direction for producing new varieties of poplar species with drought, salt, and hormone tolerance and holds substantial scientific and ecological importance, offering a potential contribution to the conservation of poplar species in arid regions.

Impact of Rhg1 copy number variation on a soybean cyst nematode resistance transcriptional network.

Soybean yield loss due to soybean cyst nematode (SCN) infestation has a negative impact on the U.S. economy. Most SCN-resistant soybeans carry a common resistance locus (Rhg1), conferred by copy number variation of a 31.2-kb segment at the Rhg1 locus. To identify the effects of Rhg1 copy number on the plant prior to SCN infection, we investigated genome-wide expression profiles in isogenic Fayette plants carrying different copy numbers at the Rhg1 locus (9-11 copies), that confer different levels of resistance to SCN. We found that even small differences in copy number lead to large changes in expression of downstream defense genes. The co-expression network constructed from differentially expressed genes (DEGs) outside the Rhg1 locus revealed complex effects of Rhg1 copy number on transcriptional regulation involving signal transduction and ethylene-mediated signaling pathways. Moreover, we report a variation in expression levels of phytoalexin biosynthesis-related genes that is correlated with copy number, and the activation of different NBS-LRR gene sets, indicating a broad effect of copy number on defense responses. Using qRT-PCR time series during SCN infection, we validated the SCN responses of DEGs detected in the copy number comparison and showed a stable upregulation of genes related to phytoalexin biosynthesis in resistant Fayette lines during the early stages of the incompatible interaction between soybeans and SCN, before syncytium formation. These results suggest additional genes that could enhance Rhg1-mediated SCN resistance.

A Methyl-Engineered DNAzyme for Endogenous Alkyltransferase Monitoring and Self-Sufficient Gene Regulation.

The on-demand gene regulation is crucial for extensively exploring specific gene functions and developing personalized gene therapeutics, which shows great promise in precision medicines. Although some nucleic acid-based gene regulatory tools (antisense oligonucleotides and small interfering RNAs) are devised for achieving on-demand activation, the introduction of chemical modifications may cause undesired side effects, thereby impairing the gene regulatory efficacy. Herein, a methyl-engineered DNAzyme (MeDz) is developed for the visualization of endogenous alkyltransferase (AGT) and the simultaneous self-sufficiently on-demand gene regulation. The catalytic activity of DNAzyme can be efficiently blocked by O6-methylguanine (O6MeG) modification and specifically restored via the AGT-mediated DNA-repairing pathway. This simply designed MeDz is demonstrated to reveal AGT of varying expression levels in different cells, opening the possibility to explore the AGT-related biological processes. Moreover, the AGT-guided MeDz exhibits cell-selective regulation on the human early growth response-1 (EGR-1) gene, with efficient gene repression in breast cancer cells and low effectiveness in normal cells. The proposed MeDz offers an attractive strategy for on-demand gene regulation, displaying great potential in biomedical applications.

Analysis of miRNAs in milk of four livestock species

BackgroundMilk is essential for mammalian nutrition because it provides vital nutrients for growth and development. Milk composition, which is influenced by genetic and environmental factors, supports lactation, a complex process crucial for milk production and quality. Recent research has focused on noncoding RNAs, particularly microRNAs (miRNAs), which are present in body fluids and regulate gene expression post-transcriptionally. This study comprehensively characterizes miRNAs in milk of four livestock species, namely Bubalus bubalis, Capra hircus, Equus asinus, and Ovis aries and identifies potential target genes.ResultsHigh-throughput sequencing of milk RNA resulted in distinct read counts across species: B. bubalis (8,790,441 reads), C. hircus (12,976,275 reads), E. asinus (9,385,067 reads), and O. aries (7,295,297 reads). E. asinus had the highest RNA mapping rate (94.6%) and O. aries the lowest (84.8%). A substantially greater proportion of miRNAs over other small RNAs was observed for the donkey milk sample (7.74%) compared to buffalo (0.87%), goat (1.57%), and sheep (1.12%). Shared miRNAs, which included miR-200a, miR-200b, miR-200c, and miR-23a among others, showed varying expression levels across species, confirmed by qPCR analysis. Functional annotation of predicted miRNA target genes highlighted diverse roles, with an enrichment in functions linked to metabolism and immunity. Pathway analysis identified immune response pathways as significant, with several miRNAs targeting specific genes across species, suggesting their regulatory function in milk.ConclusionsBoth conserved and species-specific miRNAs were detected in milk of the investigated species. The identified target genes of these miRNAs have important roles in neonatal development, adaptation, growth, and immune response. Furthermore, they influence milk and meat production traits in livestock.

Promoters Constrain Evolution of Expression Levels of Essential Genes in Escherichia coli.

Variability in expression levels in response to random genomic mutations varies among genes, influencing both the facilitation and constraint of phenotypic evolution in organisms. Despite its importance, both the underlying mechanisms and evolutionary origins of this variability remain largely unknown due to the mixed contributions of cis- and trans-acting elements. To address this issue, we focused on the mutational variability of cis-acting elements, that is, promoter regions, in Escherichia coli. Random mutations were introduced into the natural and synthetic promoters to generate mutant promoter libraries. By comparing the variance in promoter activity of these mutant libraries, we found no significant difference in mutational variability in promoter activity between promoter groups, suggesting the absence of a signature of natural selection for mutational robustness. In contrast, the promoters controlling essential genes exhibited a remarkable bias in mutational variability, with mutants displaying higher activities than the wild types being relatively rare compared to those with lower activities. Our evolutionary simulation on a rugged fitness landscape provided a rationale for this vulnerability. These findings suggest that past selection created nonuniform mutational variability in promoters biased toward lower activities of random mutants, which now constrains the future evolution of downstream essential genes toward higher expression levels.