Pre- to post-implantation transition is an essential step for early mammalian development. The epigenome dynamics such as DNA methylome and transcriptome of embryos have been analyzed, while the coordination between these two molecular layers controlling lineage fate remained elusive. Here, with a multidimensional profiling of peri-implantation embryos, we present the emerging lineage-specific DNA methylation (DNAme) patterns across species. The maternal and paternal DNA methylation levels exhibit differential dynamics in transposon elements. Genes with lineage-specific unmethylated promoters early in development retain this state and are expressed later, suggesting a role in lineage fate determination. By measuring both molecular dimensions simultaneously in individual developing cells, we identified a group of gene promoters with positive correlation between DNAme and RNAex along epiblast development. Functional validation suggested DNAme at these promoters contributes to non-canonical DNAme-RNAex dynamics, potentially via complex TF-mediated or epigenetic regulation. This study provides clues for understanding molecular regulation of peri-implantation embryogenesis.

Breast cancer is one of the most common malignant tumors, threatening women's health and life globally. Despite significant treatment advances, its prognosis still faces great challenges. With the rapid development of molecular biology and genomics, the role of detoxification enzymes in breast cancer occurrence, development, and prognosis has gained increasing attention. This paper aims to establish a prognostic model based on detoxification enzymes-related genes to predict breast cancer patient survival.

Unsupervised clustering was used to analyze breast cancer samples based on detoxification enzymes-related genes expression. Lasso cox regression analysis and univariate and multivariate Cox analysis were used to process the data, and machine learning algorithm was used to construct breast cancer prognosis model. The effect of detoxification enzymes-related genes on breast cancer was analyzed by single cell analysis.

The samples were classified into two subtypes, and a breast cancer prognosis model based on detoxification enzymes-related genes was constructed and validated using TCGA and GEO cohorts. Significant differences in pathways, immune infiltration, immunotherapy response, and drug sensitivity were observed between high- and low-risk groups. Single-cell analysis revealed that SQLE, a detoxification enzymes-related gene, was highly expressed in breast cancer epithelial cells (cancer cells), where SQLE + epithelial cells primarily influenced exhausted CD8 + T cells via the MIF signaling pathway.

In summary, the detoxification enzymes-related genes-based prognostic model developed in this study provides an effective tool for predicting breast cancer prognosis and offers new insights for diagnosis and treatment.

The nucleoprotein structures known as telomeres provide genomic integrity by protecting the ends of chromosomes. Tumorigenesis is associated with alterations in telomere function and stability. This narrative review provides evidence of the potential prognostic value of telomere length and telomerase in leukemias. On the one hand, oxidative stress and mitochondrial dysfunction can accelerate telomere shortening, leading to higher susceptibility and the progression of leukemia. On the other hand, cytogenetic alterations (such as gene fusions and chromosomal abnormalities) and genomic complexity can result from checkpoint dysregulation, the induction of the DNA damage response (DDR), and defective repair signaling at telomeres. This review thoroughly outlines the ways by which telomere dysfunction can play a key role in the development and progression of four primary leukemias, including chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), and acute leukemias of myeloid or lymphoid origin, highlighting the potential prognostic value of telomere length in this field. However, telomerase, which is highly active in leukemias, can prevent the rate of telomere attrition. In line with this, leukemia cells can proliferate, suggesting telomerase as a promising therapeutic target in leukemias. For this reason, telomerase-based immunotherapy is analyzed in the fight against leukemias, leveraging the immune system to eliminate leukemia cells with uncontrolled proliferation.

Marfan syndrome (MFS) is a genetic disorder primarily affecting the connective tissue, with cardiovascular complications as the leading cause of mortality. While mutations in the FBN1 gene are the primary cause, the severity and progression of the disease can vary significantly among individuals. DNA methylation, a key epigenetic regulatory mechanism, has garnered attention in MFS research, particularly regarding methylation changes in the FBN1 locus and their effects on fibrillin-1 expression. Differential methylation and expression of genes related to inflammation (e.g., interleukin (IL)-10, IL-17) and oxidative stress (e.g., PON2, TP53INP1) have been linked to MFS aortic pathology. These alterations likely contribute to disease progression by influencing inflammatory responses, smooth muscle cell apoptosis, and biomechanical properties of the aorta. The transforming growth factor-beta (TGF-β) signaling pathway plays a central role in MFS pathology, with aberrant methylation of related genes potentially elevating active TGF-β levels and exacerbating aortic lesions. Notably, tissue-specific methylation patterns, especially in smooth muscle cells of the aorta, remain poorly understood. A deeper understanding of DNA methylation's role in MFS could pave the way for early interventions and epigenetic-targeted therapies.

Telomerase reverse transcriptase (TERT) is one of the most well-established oncogenes in tumor development and progression. It is widely known that TERT promoter hypermethylation is associated with its transcription activation. Despite its canonical role in maintaining telomere length in cancer cells, TERT is also involved in various oncogenic processes independent of its enzymatic activity. However, the role of TERT in the tumor immune microenvironment has been largely unexplored. Hence, we assessed the associations between TERT promoter methylation and its expression, clinicopathological features, overall survival, immune cell infiltration, and response to immune checkpoint inhibitor therapy in clear cell renal cell carcinoma.

A single-sample gene-set enrichment analysis algorithm was used to quantify the relative abundance of each type of immune cell infiltration in the tumor microenvironment (TME) of the TCGA KIRC cohort. We used Spearman's rank correlation to calculate the correlation coefficients between TERT promoter methylation and immune cell infiltration. The relative methylation of cg11625005 in our validation cohort was detected by pyrosequencing and the relative infiltration of CD4 + and CD8 + T cells infiltration in the TME was measured by immunohistochemistry.

The TERT promoter was significantly hypermethylated in clear cell renal cell tumor tissues, which was related to the transcriptional activation of TERT. TERT promoter hypermethylation was significantly correlated with aggressive phenotypes and poor survival in clear cell renal cell carcinoma patients. Furthermore, TERT promoter methylation was significantly positively correlated with CD4 + /CD8 + T cells infiltration and immune checkpoint molecule (CTLA-4, TIGIT, PD-1 and LAG3) expression. And TERT promoter methylation was correlated with the therapeutic response to anti-PD1 immunotherapy.

TERT promoter methylation is a promising predictive biomarker of immune cell infiltration, overall survival, clinicopathological characteristics and response to anti-PD1 immunotherapy treatment in clear cell renal cell carcinoma patients.

Zebrafish (Danio rerio) have become a valuable model in biomedical research due to their genetic similarity to humans, rapid development, and suitability for high-throughput studies. Metabolomic analyses in zebrafish provide critical insights into the biochemical pathways underlying health and disease. This review explores the applications of metabolomics in zebrafish research, highlighting its contributions to understanding embryonic development, tuberculosis, neurodegenerative disorders such as Alzheimer's disease, obesity-related metabolic dysfunction, and drug-induced toxicity through a thorough literature review. Zebrafish metabolomics reveals dynamic metabolite shifts during vertebrate development. In tuberculosis research, zebrafish models have helped identify metabolic biomarkers with potential translational relevance. Studies on Alzheimer's disease suggest that metabolomics can elucidate neuroprotective mechanisms, while investigations into obesity have provided insights into metabolic imbalances associated with kidney dysfunction. Furthermore, toxicometabolomic studies have demonstrated the utility of zebrafish in assessing drug-induced renal injury. Despite their advantages, zebrafish metabolomics faces challenges, including differences in metabolic rates compared to mammals, the need for standardized protocols, and limitations in metabolite database annotations. Nonetheless, integrating metabolomics with other omics approaches holds great promise for advancing disease research and paving the way for personalized medicine.

DNA 5-methylcytosine (5mC) and RNA N6-methyladenosine (m6A) methylation are prevalent modifications in eukaryotes, both playing crucial roles in gene regulation. Recent studies have explored their crosstalk and impact on transcription. However, the intricate relationships among 5mC, m6A, and gene expression remain incompletely elucidated.

We collect data on 5mC, m6A, and gene expression from samples from three tissues from each of four pregnant cattle and sheep. We construct a comprehensive genome-wide self-interaction (same gene) and across-interaction (across genes) network of 5mC and m6A within gene-bodies or promoters and gene expression in both species. Qualitative analysis identifies uniquely expressed genes with specific m6A methylation in each tissue from both species. A quantitative comparison of gene expression ratio between methylated and unmethylated genes for m6A within gene body and promoter, and 5mC within gene body and promoter confirms the positive effect of RNA methylation on gene expression. Importantly, the influence of RNA methylation on gene expression is stronger than that of DNA methylation. The predominant self- and across-interactions are between RNA methylation within gene bodies and gene expression, as well as between RNA methylation within promoters and gene expression in both species.

RNA methylation has a stronger effect on gene expression than does DNA methylation within gene bodies and promoters. DNA and RNA methylation in gene-bodies has a greater impact on gene expression than those in promoters. These findings deepen comprehension of the dynamics and complex relationships among the epigenome, epitranscriptome, and transcriptome, offering fresh insights for advancing epigenetics research.

Ras signaling regulates many cellular processes in cancer development. While well-known Ras-related oncogenes, such as KRAS, have been extensively explored, the role of other Ras-related genes in cancer remains poorly studied. Dexamethasone-induced Ras-related protein 1 (RASD1), a member of the Ras superfamily, is widely expressed across various tissues and is involved in inhibiting cell growth and inducing apoptosis, suggesting a potential role as a tumor suppressor. Here, we investigated RASD1 expression across multiple tissues and cancers, utilizing data from The Cancer Genome Atlas (TCGA), Human Protein Atlas, and Genotype-Tissue Expression (GTEx) databases. Our analysis revealed a significant downregulation of RASD1 mRNA expression in several cancer types compared to normal tissues, correlating with low levels of promoter methylation. Interestingly, high RASD1 expression correlated with a favorable prognosis in multiple cancers. Immune cell infiltration analysis indicated that elevated RASD1 expression is associated with an increased infiltration of CD4+ T cells and myeloid-derived dendritic cells in cancer. Furthermore, the expression of genes exhibiting similar expression patterns as RASD1 suggest that RASD1 may play a role in interleukin-4-mediated apoptosis and could regulate the transcription of the phosphatase and tensin homolog (PTEN) gene. Overall, these findings suggest that RASD1 may modulate immune signaling and tumor suppressive pathways.

Sex differences in physiological and disease traits are pervasive and begin during early development, but the genetic architecture of these differences is largely unknown. Here, we leverage the human placenta, a transient organ during pregnancy critical to fetal development, to investigate the impact of sex in the regulatory landscape of placental autosomal methylome and transcriptome, and its relevance to health and disease. We find that placental methylation and its genetic regulation are extensively impacted by fetal sex, whereas sex differences in placental gene expression and its genetic regulation are limited. We identify molecular processes and regulatory targets that are enriched in a sex-specific manner, and find enrichment of imprinted genes in sex-differentiated placental methylation, including female-biased methylation within the well-known KCNQ1OT1/CDKN1C imprinting cluster of genes expressed in a parent-of-origin dependent manner. We establish that several sex-differentiated genetic effects on placental methylation and gene expression colocalize with birthweight and adult disease genetic associations, facilitating mechanistic insights on early life origins of health and disease outcomes shaped by sex.

Methylated stool DNA (sDNA) is a reliable noninvasive biomarker for early colorectal cancer (CRC) diagnosis. However, there are barely any diagnostic panels that can achieve both a sensitivity and specificity exceeding 90% simultaneously.

We aimed to identify a novel methylated sDNA panel and model for the early diagnosis of CRC.

We conducted methyl-CpG binding domain isolated genome sequencing (MiGS) on CpG island methylation phenotype (CIMP)-positive (n = 3) and CIMP-negative CRC tissues (n = 3) and their corresponding normal adjacent tissues. Subsequently, by utilizing both the aforementioned data and public datasets, we identified a set of promising methylated sDNA markers for CRC. Next, we validated 5 of these genes using pyrosequencing in CRC patients (n = 31). Then, we developed a combined diagnostic model (CDM) for CRC based on the methylation status of PRDM12, FOXE1, and SDC2 by a Training cohort (n = 231). Finally, the performance of CDM was evaluated in an independent multicenter Validation cohort (n = 800).

A total of 1062 participants were included in this study. The area under the curve (AUC) of the CDM was 0.979 (95% CI: 0.960-0.997), and the optimal sensitivity and specificity were 97.35% and 99.05%, respectively, in the training cohort (n = 231). In the independent validation cohort (n = 800), the AUC was 0.950 (95% CI: 0.927-0.973), along with the optimal sensitivity of 92.75% and specificity of 97.21%. When CRC and advanced adenoma (AAD) were used as diagnostic targets, the model AUC was 0.945 (95% CI: 0.922-0.969), with an optimal sensitivity of 91.89% and a specificity of 95.21%. The model sensitivity for nonadvanced adenoma patients was 68.66%.

The sDNA diagnostic model CDM, developed from both CIMP-P and CIMP-N, exhibited exceptional performance in CRC and could serve as a potential alternative strategy for CRC screening.

Uterine corpus endometrial carcinoma (UCEC) is one of the most common gynecologic tumors. Due to the high recurrence and metastasis of UCEC, it is crucial for patients to find new biomarkers for diagnosis and therapy. In this study, R software and the TCGA database were used to screen candidate UCEC predictive markers. Western blot and RT-qPCR were performed to detect protein and mRNA expression of EXT1 in UCEC cell lines. In addition, MTT assay, flow cytometry, transwell assay, and wound healing assay were conducted to assess the cell viability, apoptosis, invasion, and migration in UCEC cells. Overlap-extension PCR technique was employed to construct the vector targeting the deletion of the methylated segment of EXT1. The results showed that a total of 11 candidate genes were obtained and EXT1 was identified as a potential target. The expression and methylation levels of EXT1 were both increased in UCEC tissues and cell lines, as well as elevated EXT1 was closely related to the poor prognosis of patients. Besides, the knockdown of EXT1 significantly inhibited the malignant biological behaviors in UCEC cells. Additionally, the current study also found that the deletion of 1559-2146 bp CpG island segment upregulated EXT1 expression and promoted malignant biological behaviors in UCEC cells. Furthermore, the presence of m7G RNA methylation in UCEC cells also was found. In conclusion, the methylation of EXT1 influenced the gene expression, thereby affecting the malignant biological behaviors in UCEC cells and regulating the pathological progression of UCEC.

DNA methylation at cytosine bases (5-methylcytosine, 5mC) is a heritable epigenetic mark regulating gene expression. While enzymes that metabolize 5mC are well-characterized, endogenous signaling molecules that regulate DNA methylation machinery have not been described. We report that physiological nitric oxide (NO) concentrations reversibly inhibit the DNA demethylases TET and ALKBH2 by binding to the mononuclear non-heme iron atom forming a dinitrosyliron complex (DNIC) and preventing cosubstrates from binding. In cancer cells treated with exogenous NO, or endogenously synthesizing NO, 5mC and 5-hydroxymethylcytosine (5hmC) increase, with no changes in DNA methyltransferase activity. 5mC is also significantly increased in NO-producing patient-derived xenograft tumors from mice. Genome-wide methylome analysis of cells chronically treated with NO (10 days) shows enrichment of 5mC and 5hmC at gene-regulatory loci, correlating with altered expression of NO-regulated tumor-associated genes. Regulation of DNA methylation is distinctly different from canonical NO signaling and represents a unique epigenetic role for NO.

Promoter DNA methylation is a key epigenetic mark, commonly associated with gene silencing. However, we noticed that a positive association between promoter DNA methylation and expression is surprisingly common in cancer. Here, we use hit-and-run CRISPR/dCas9 epigenome editing to evaluate how deposition of DNA methylation can regulate gene expression dependent on pre-existing chromatin environment. While the predominant effect of DNA methylation in non-bivalent promoters is gene repression, we show that in bivalent promoters this often leads to gene activation. We demonstrate that gain of DNA methylation leads to reduced MTF2 binding and eviction of H3K27me3, a repressive mark that guards bivalent genes against activation. Our cancer patient data analyses reveal that in cancer, this mechanism likely leads to activation of a large group of transcription factors regulating pluripotency, apoptosis, and senescence signalling. In conclusion, our study uncovers an activating role of DNA methylation in bivalent promoters, with broad implications for cancer and development.

ARPC1B has been identified as a key regulator of malignant biological behavior in various tumors. However, its specific role in clear cell renal cell carcinoma (ccRCC) remains poorly understood. This study aims to evaluate the influence of ARPC1B on the prognosis and disease progression in ccRCC patients.

Multi-omics data and clinical information from public databases were analyzed to determine the associations between ARPC1B and prognosis, clinical features, immune microenvironment, and drug sensitivity in ccRCC. Co-expression and gene set enrichment analyses were conducted to elucidate the potential role of ARPC1B in ccRCC pathogenesis. Functional assays, including RT-qPCR, CCK8 assays, colony formation assays, immunofluorescence, immunohistochemistry, and xenograft tumor formation in nude mice, were performed to assess ARPC1B's impact on cell proliferation and apoptosis. Flow cytometry and Western blotting were further employed to investigate the underlying molecular mechanisms of ARPC1B in ccRCC.

ARPC1B expression was significantly elevated in ccRCC and associated with an unfavorable prognosis. Both independent and meta-analyses confirmed that ARPC1B is an independent prognostic risk factor in ccRCC. Furthermore, ARPC1B expression significantly correlated with the immune microenvironment and drug sensitivity. In vitro, experiments demonstrated that ARPC1B knockdown suppressed ccRCC cell proliferation and induced apoptosis through the BAX-Bcl-2/c-caspase3/c-PARP axis, which was further validated by in vivo studies.

ARPC1B overexpression is associated with poor prognosis, altered immune status, and drug sensitivity in ccRCC. Furthermore, ARPC1B promotes the malignant behavior of ccRCC cells and holds potential as a prognostic biomarker and therapeutic target for ccRCC.

Super-enhancers (SEs) are typically located in the regulatory regions of genes, driving high-level gene expression. Identifying SEs is crucial for a deeper understanding of gene regulatory networks, disease mechanisms, and the development and physiological processes of organisms, thus exerting a profound impact on research and applications in the life sciences field. Traditional experimental methods for identifying SEs are costly and time-consuming. Existing methods for predicting SEs based solely on sequence data use deep learning for feature representation and have achieved good results. However, they overlook biological features related to physicochemical properties, leading to low interpretability. Additionally, the complex model structure often requires extensive labeled data for training, which limits their further application in biological data. In this paper, we integrate the strengths of different models and proposes an ensemble model based on an integration strategy to enhance the model's generalization ability. It designs a multi-angle feature representation method that combines local structure and global information to extract high-dimensional abstract relationships and key low-dimensional biological features from sequences. This enhances the effectiveness and interpretability of the model's input features, providing technical support for discovering cell-specific and species-specific patterns of SEs. We evaluated the performance on both mouse and human datasets using five metrics, including area under the receiver operating characteristic curve accuracy, and others. Compared to the latest models, EnsembleSE achieved an average improvement of 4.5% in F1 score and an average improvement of 8.05% in recall, demonstrating the robustness and adaptability of the model on a unified test set. Source codes are available at https://github.com/2103374200/EnsembleSE-main.

The interaction between TF binding and DNA methylation is increasingly recognized as a key player in the regulation of gene expression. However, the role of this interaction in regulating ICAM1 expression in breast cancer has not been elucidated. CpG methylation in the ICAM1 promoter is negatively correlated with ICAM1 expression, and ICAM1 expression is significantly positively correlated with DNMT and TET3 expression in breast cancer. TF binding attenuates ICAM1 promoter CpG methylation and promotes ICAM1 transcription. DNA methylation regulation enhances ICAM1 expression in breast cancer by promoting the transcription of transcription factors. In terms of mechanisms, RELA and STATs recruit TET3 to prevent DNMT-mediated DNA methylation, thereby maintaining CpG island hypomethylation in the ICAM1 promoter. Therefore, TF occupancy limits DNA methylation and affects ICAM1 expression in breast cancer.

DNA methylation, also known as 5-methylcytosine, is an epigenetic modification that has crucial functions in plant growth, development and adaptation. The cellular DNA methylation level is tightly regulated by the combined action of DNA methyltransferases and demethylases. Protein complexes involved in the targeting and interpretation of DNA methylation have been identified, revealing intriguing roles of methyl-DNA binding proteins and molecular chaperones. Structural studies and in vitro reconstituted enzymatic systems have provided mechanistic insights into RNA-directed DNA methylation, the main pathway catalysing de novo methylation in plants. A better understanding of the regulatory mechanisms will enable locus-specific manipulation of the DNA methylation status. CRISPR-dCas9-based epigenome editing tools are being developed for this goal. Given that DNA methylation patterns can be stably transmitted through meiosis, and that large phenotypic variations can be contributed by epimutations, epigenome editing holds great promise in crop breeding by creating additional phenotypic variability on the same genetic material.

Exercise is a potent stimulus for combatting skeletal muscle ageing. To study the effects of exercise on muscle in a preclinical setting, we developed a combined endurance-resistance training stimulus for mice called progressive weighted wheel running (PoWeR). PoWeR improves molecular, biochemical, cellular and functional characteristics of skeletal muscle and promotes aspects of partial epigenetic reprogramming when performed late in life (22-24 months of age). In this investigation, we leveraged pan-mammalian DNA methylome arrays and tandem mass-spectrometry proteomics in skeletal muscle to provide detailed information on late-life PoWeR adaptations in female mice relative to age-matched sedentary controls (n = 7-10 per group). Differential CpG methylation at conserved promoter sites was related to transcriptional regulation genes as well as Nr4a3, Hes1 and Hox genes after PoWeR. Using a holistic method of -omics integration called binding and expression target analysis (BETA), methylome changes were associated with upregulated proteins related to global and mitochondrial translation after PoWeR (P = 0.03). Specifically, BETA implicated methylation control of ribosomal, mitoribosomal, and mitochondrial complex I protein abundance after training. DNA methylation may also influence LACTB, MIB1 and UBR4 protein induction with exercise - all are mechanistically linked to muscle health. Computational cistrome analysis predicted several transcription factors including MYC as regulators of the exercise trained methylome-proteome landscape, corroborating prior late-life PoWeR transcriptome data. Correlating the proteome to muscle mass and fatigue resistance revealed positive relationships with VPS13A and NPL levels, respectively. Our findings expose differential epigenetic and proteomic adaptations associated with translational regulation after PoWeR that could influence skeletal muscle mass and function in aged mice. KEY POINTS: Late-life combined endurance-resistance exercise training from 22-24 months of age in mice is shown to improve molecular, biochemical, cellular and in vivo functional characteristics of skeletal muscle and promote aspects of partial epigenetic reprogramming and epigenetic age mitigation. Integration of DNA CpG 36k methylation arrays using conserved sites (which also contain methylation ageing clock sites) with exploratory proteomics in skeletal muscle extends our prior work and reveals coordinated and widespread regulation of ribosomal, translation initiation, mitochondrial ribosomal (mitoribosomal) and complex I proteins after combined voluntary exercise training in a sizeable cohort of female mice (n = 7-10 per group and analysis). Multi-omics integration predicted epigenetic regulation of serine β-lactamase-like protein (LACTB - linked to tumour resistance in muscle), mind bomb 1 (MIB1 - linked to satellite cell and type 2 fibre maintenance) and ubiquitin protein ligase E3 component N-recognin 4 (UBR4 - linked to muscle protein quality control) after training. Computational cistrome analysis identified MYC as a regulator of the late-life training proteome, in agreement with prior transcriptional analyses. Vacuolar protein sorting 13 homolog A (VPS13A) was positively correlated to muscle mass, and the glycoprotein/glycolipid associated sialylation enzyme N-acetylneuraminate pyruvate lyase (NPL) was associated to in vivo muscle fatigue resistance.

TMEM189 is a recently discovered transmembrane protein involved in ether glycerophospholipid synthesis and ferroptosis regulation. However, its role in tumors is not well understood.

This study aimed to elucidate the oncogenic effects and prognostic values of TMEM189 in tumors.

We performed a pan-cancer analysis of TMEM189 using various databases, bioinformatics and statistical tools, and tissue microarray analysis.

TMEM189 was upregulated in tumors compared to normal tissues. High TMEM189 expression was found to be linked to reduced promoter methylation. Moreover, TMEM189 exhibited a negative correlation with immunogenic markers, immune cell infiltration, and expression of Immune Checkpoint Genes (ICGs) in most cancers, implicating its immunosuppressive role in tumor microenvironments (TME). The genes that interact and are similar to TMEM189 were involved in hotspot signaling pathways in pan-cancer. TMEM189 overexpression was found to be usually associated with poor prognosis, especially an independent prognostic risk factor for BLCA, BRCA, LUAD, MESO, LIHC, and SKCM.

TMEM189 is overexpressed and exerts immunosuppressive effects in many tumors with a significant association with poor prognosis, suggesting its potential as a therapeutic target in cancer treatment.

Phosphodiesterase 2A (PDE2A) plays a pivotal role in modulating cyclic nucleotide metabolism. Recent studies have shown that PDE2A is associated with some tumors, but its expression profiles, prognostic significance, and immunological roles in diverse cancer types remain unclear. Utilizing advanced bioinformatics tools, we performed a comprehensive analysis of PDE2A gene expression in multiple human cancers. Our study revealed that PDE2A expression was significantly reduced in the majority of cancer types and clinicopathological stages (I to IV) compared to normal tissues. Additionally, PDE2A expression was closely related to the prognosis of cancers such as stomach adenocarcinoma (STAD), ovarian serous cystadenocarcinoma (OV), and liver hepatocellular carcinoma (LIHC). Cox regression analyses indicated that PDE2A can act as an independent prognostic factor for these cancers. The level of PDE2A DNA methylation was significantly decreased in most cancers. Genetic alterations in PDE2A predominantly manifest in the form of amplifications. Moreover, infiltrating cells and immune checkpoint genes, including PDCD1, exhibited notable correlations with PDE2A expression. Significant associations were observed between PDE2A expression and tumor mutation burden as well as microsatellite instability. Single cell sequencing revealed PDE2A's crucial role in regulating differentiation and angiogenesis of cancer cells. Functional enrichment analysis emphasized the important role of PDE2A in synaptic transmission and tumor development. Aberrant expression of PDE2A influenced the sensitivity of various antitumor and chemotherapy drugs. This research provided a comprehensive analysis of PDE2A in human cancers, highlighting its potential as both a prognostic marker and an immunotherapy target for future research.

The canonical view of DNA methylation, a pivotal epigenetic regulation mechanism in eukaryotes, dictates its role as a suppressor of gene activity, particularly within promoter regions. However, this view is being challenged as it is becoming increasingly evident that the connection between DNA methylation and gene expression varies depending on the genomic location and is therefore more complex than initially thought. We examined DNA methylation levels in the gut epithelium of Atlantic salmon (Salmo salar) using whole-genome bisulfite sequencing, which we correlated with gene expression data from RNA sequencing of the same gut tissue sample (RNA-seq). Assuming epigenetic signals might be pronounced between distinctive phenotypes, we compared large and small fish, finding 22 significant associations between 22 differentially methylated regions and 21 genes. We did not detect significant methylation differences between large and small fish. However, we observed a consistent signal of methylation levels around the transcription start sites (TSS), being negatively correlated with the expression levels of those genes. We found both negative and positive associations of methylation levels with gene expression further upstream or downstream of the TSS, revealing a more unpredictable pattern. The 21 genes showing significant methylation-expression correlations were involved in biological processes related to salmon health, such as growth and immune responses. Deciphering how DNA methylation affects the expression of such genes holds great potential for future applications. For instance, our results suggest the importance of genomic context in targeting epigenetic modifications to improve the welfare of aquaculture species like Atlantic salmon.

Aberrant methylation of genomic DNA has been found in preeclamptic placentas, which is characterized by elevated DNA methylation and hypermethylation of gene body regions, but the underlying mechanism is not yet fully understood.

Global DNA methylation was assessed through ELISA and HPLC. The methylation sites were detected using the Illumina Human Methylation 450 K Microarray. The methylation level of FAM46C promoter and gene body was detected through the bisulfite sequencing. RNA-seq was utilized to investigate the mechanism by which DNMT3A and FAM46C mediate the migration and invasion of trophoblast cells.

We discovered that DNMT3A knockdown led to elevated levels of gene body methylation and FAM46C transcription. FAM46C downregulation completely rescued the suppressive effects caused by DNMT3A knockdown on the migration and invasion of trophoblast cells. Mechanistically, DNMT3A reduction led to an increase in the enrichment of DNMT3B and DNMT1 in the gene body region of FAM46C. The results of transcriptome sequencing showed that DNMT3A and FAM46C regulate the adhesion of trophoblast cells. Elevated expression of FAM46C and increased methylation levels within its gene body region were observed in extravillous trophoblast cells of early-onset preeclamptic placentas.

DNMT3A-mediated aberrant FAM46C gene body methylation is relevant to the development of early-onset preeclampsia.

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most lethal malignancies, characterized by its aggressive progression and dismal prognosis. Advances in epigenetic profiling, specifically DNA methylation analysis, have significantly deepened our understanding of PDAC pathogenesis. This review synthesizes findings from recent genome-wide DNA methylation studies, which have delineated a complex DNA methylation landscape differentiating between normal and cancerous pancreatic tissues, as well as across various stages and molecular subtypes of PDAC. These studies identified specific differentially methylated regions (DMRs) that not only enhance our grasp of the epigenetic drivers of PDAC but also offer potential biomarkers for early diagnosis and prognosis, enabling the customization of therapeutic approaches. The review further explores how DNA methylation profiling could facilitate the development of subtype-tailored therapies, potentially improving treatment outcomes based on precise molecular characterizations. Overall, leveraging DNA methylation alterations as functional biomarkers holds promise for advancing our understanding of disease progression and refining PDAC management strategies, which could lead to improved patient outcomes and a deeper comprehension of the disease's underlying biological mechanisms.

Vibrio parahaemolyticus is the main pathogen causing acute hepatopancreatic necrotic disease in crustaceans. To elucidate the epigenetic regulatory mechanism of crustacean resistance to V. parahaemolyticus infection, we conducted artificial infection studies on Portunus trituberculatus. The results showed that the mortality rate reached the highest at 12 h of artificial infection, which was 23.69 %. At 72 h after V parahaemolyticus infection, the expression level of DNA demethylase (ten-eleven-translocation protein) Tet was significantly decreased, the expression of DNA methyltransferase Dnmt3B fluctuated significantly. Based on the differential expression levels of Tet and Dnmt3B. We depict for DNA methylation profiles of the whole genome of P. trituberculatus at single-base resolution by using whole-genome bisulfite sequencing (WGBS) on hemolymph tissues. The overall DNA methylation level was low at 2.16 % in P. trituberculatus hemolymph. A total of 2590 differentially methylated regions (DMRs) were identified, of which 1329 were hypermethylated and 1261 were hypomethylated, and 1389 genes were annotated in these DMRs. Differently methylated genes (DMGs) were significantly enriched in ribosomes (KO03010), protein kinases (KO01001), cell cycle (HSA04110), endocrine resistance (HSA01522) and FoxO signaling pathway (KO04068). Finally, we selected six differentially methylated genes for quantitative analysis. The results showed that DNA methylation not only has a negative regulatory effect on gene expression, but also has a positive regulatory effect. These results indicated that DNA methylation in the regulation of genes involved in immune responses contributes to the resistance of P. trituberculatus to V. parahaemolyticus, which is valuable for understanding how crustaceans regulate the innate immune system to defend against bacterial infections.

Hydroquinone (HQ), a metabolite of benzene, is frequently utilized as a surrogate for benzene in in vitro studies and is associated with the development of acute myeloid leukemia (AML). In the hemotoxicity caused by benzene and HQ, cell apoptosis plays a key role. However, the molecular mechanisms underlying HQ are unknown. Studies have indicated that Suv39h1 is involved in regulating cell division and proliferation by regulating histone H3K9me3. Meanwhile, the Wnt/β-catenin signaling pathway also plays a significant role in cell proliferation and apoptosis. Therefore, this study was aimed at exploring the regulatory role of Suv39h1 and the Wnt/β-catenin signaling pathway in the effects of HQ on bone marrow mesenchymal stem cells (BMSCs), as well as its influence on cell proliferation and apoptosis. The results demonstrated that HQ elevated the levels of Suv39h1 and H3K9me3 and activated the Wnt/β-catenin signaling pathway by upregulating β-catenin, Wnt2b, C-myc, and Cyclin D1 and downregulating Wnt5a, resulting in an increase in cell growth and a decrease in apoptosis. Suv39h1 knockdown inhibited the Wnt/β-catenin signaling pathway. Meanwhile, inhibition of the Wnt/β-catenin signaling pathway resulted in the down-regulation of Suv39h1 and H3K9me3 in BMSCs. They both promoted cell proliferation and inhibited apoptosis in the effects of HQ on BMSCs by downregulating the expression of Cyt-C, Bax, Caspase 3, and Caspase 9 and upregulating the expression of Bcl-xl. Therefore, we concluded that Suv39h1 and the Wnt/β-catenin signaling pathway may mutually regulate each other in the effects of HQ on BMSCs in order to ameliorate the altered function of BMSCs.

This project focused on evaluating the effects of maternal dietary choline intake on global DNA methylation profiles and related transcriptional changes in rainbow trout offspring. Three experimental diets were formulated to test different levels of choline intake: (a) 2065 ppm choline (Low Choline, 0 % supplementation), (b) 5657 ppm choline (Medium Choline, 0.6 % supplementation), and (c) 9248 ppm choline (High Choline, 1.2 % choline supplementation). Six rainbow trout families were fed experimental diets beginning 18 months post-hatch until spawning; their offspring were fed a commercial diet. Reduced representation bisulfite sequencing (RRBS) was utilized to measure genome-wide methylation in offspring immediately after hatching. When comparing to the Medium Choline offspring, differential DNA methylation occurred more in the Low Choline offspring than High Choline, especially in genic features like promoters. The differentially methylated CpGs (q ≤ 0.01) were identified evenly between CpG islands and shores in the genome, mostly found in the introns of genes. Genes such as fabp2 and leap2B associated with protein binding, fatty acid binding, DNA binding, and response to bacteria were differentially methylated and detected as differentially regulated genes by previous RNA-seq analysis. Although these findings indicate that levels of dietary choline available in broodstock diets alter offspring DNA methylation;, most differentially expressed genes were not associated with differential DNA methylation, suggesting additional mechanisms playing a role in regulating gene expression in response to maternal choline intake.

Japanese encephalitis virus (JEV) is a neurotropic and neuroinvasive flavivirus causing viral encephalitis, which seriously threatens the development of animal husbandry and human health. DNA methylation is a major epigenetic modification involved in viral pathogenesis, yet how DNA methylation affects JEV infection remains unknown. Here, we show genome-wide DNA methylation profiles in the brains of JEV-infected mice compared to mock-infected mice. JEV can significantly increase the overall DNA methylation levels in JEV-infected mouse brains. A total of 14,781 differentially methylated regions associated genes (DMGs) have been identified. Subsequently, KEGG pathway analysis suggested that DNA methylation modulates the tight junction signaling pathway, which can potentially impact the permeability of the blood-brain barrier (BBB). We demonstrate that hypermethylation of the tight junction gene Afdn promoter inhibited AFDN expression and increased monolayer permeability of mouse brain microvascular endothelial (bEnd.3) cells in an in vitro transwell assay. Collectively, this study reveals that DNA methylation is increased in a murine Japanese encephalitis model and that modulation of Afdn expression promotes BBB permeability.

Thyroid cancer (TC) represents one of the most prevalent endocrine malignancies, with a rising incidence worldwide. Epigenetic alterations, which modify gene expression without altering the underlying DNA sequence, have garnered significant attention in recent years. Increasing evidence underscores the pivotal role of epigenetic modifications, including DNA methylation, RNA methylation, and histone methylation, in the pathogenesis of TC. This review provides a comprehensive overview of these reversible and environmentally influenced epigenetic modifications, highlighting their molecular mechanisms and functional roles in TC. Additionally, the clinical implications, challenges associated with studying these epigenetic modifications, and potential future research directions are explored.

Breast cancer is the most common cancer worldwide, the leading cause of cancer death in women, and the fifth leading cause of cancer death. Triple negative breast cancer (TNBC), with high metastasis and mortality rates, is the most challenging subtype in breast cancer treatment. There is an urgent need to develop anti-TNBC drugs with significant efficacy, low side effects and good availability. In early drug screening, blestriarene C was found to have inhibitory effects on TNBC cells. In this article, we further explore the mechanisms associated with blestriarene C for breast cancer.

In this article, we take the approach of network pharmacology combined with in vivo and in vitro experiments. Network pharmacology analysis was used to predict the active components in Baiji, and to investigate the hub targets and related mechanisms of BC in TNBC treatment. The mechanism of anti-TNBC in vitro was evaluated by CCK-8 assay, cell apoptosis and cell cycle assays, wound healing assay, WB assay, and molecular docking analysis. The inhibition effect in vivo was test in subcutaneous tumor models established in mice.

Through network pharmacology analysis and experiments, we screened out BC as the main active ingredient, and found that BC could inhibit the Ras/ERK/c-Fos signaling pathway while downregulating the expression of HSP90AA1 and upregulating the expression of PTGS2, thereby promoting apoptosis, causing S-phase cycle arrest, and inhibiting the proliferation and migration of BT549 cells. The in vivo results illustrated that BC inhibited the growth of TNBC tumors and has a high safety profile. By integrating network pharmacology with in vitro and in vivo experiments, this study demonstrated that BC inhibited the proliferation and migration of TNBC cells by inhibiting the Ras/ERK/c-Fos signaling pathway, promoting apoptosis, and causing S-phase cycle arrest.

This study provides new evidence for the use of BC as a novel drug for TNBC treatment.

The unsustainable use of manmade chemicals poses significant threats to biodiversity and human health. Emerging evidence highlights the potential of certain chemicals to cause transgenerational impacts on metabolic health. Here, we investigate male transmitted epigenetic transgenerational effects of the anti-androgenic herbicide linuron in the pancreas of Xenopus tropicalis frogs, and their association with metabolic phenotypes. Reduced representation bisulfite sequencing (RRBS) was used to assess genome-wide DNA methylation patterns in the pancreas of adult male F2 generation ancestrally exposed to environmentally relevant linuron levels (44 ± 4.7 μg/L). We identified 1117 differentially methylated regions (DMRs) distributed across the X. tropicalis genome, revealing potential regulatory mechanisms underlying metabolic disturbances. DMRs were identified in genes crucial for pancreatic function, including calcium signalling (clstn2, cacna1d and cadps2), genes associated with type 2 diabetes (tcf7l2 and adcy5) and a biomarker for pancreatic ductal adenocarcinoma (plec). Correlation analysis revealed associations between DNA methylation levels in these genes and metabolic phenotypes, indicating epigenetic regulation of glucose metabolism. Moreover, differential methylation in genes related to histone modifications suggests alterations in the epigenetic machinery. These findings underscore the long-term consequences of environmental contamination on pancreatic function and raise concerns about the health risks associated with transgenerational effects of pesticides.

Mammalian neurons lose the ability to regenerate their central nervous system axons as they mature during embryonic or early postnatal development. Neuronal maturation requires a transformation from a situation in which neuronal components grow and assemble to one in which these components are fixed and involved in the machinery for effective information transmission and computation. To regenerate after injury, neurons need to overcome this fixed state to reactivate their growth programme. A variety of intracellular processes involved in initiating or sustaining neuronal maturation, including the regulation of gene expression, cytoskeletal restructuring and shifts in intracellular trafficking, have been shown to prevent axon regeneration. Understanding these processes will contribute to the identification of targets to promote repair after injury or disease.

Cuproptosis, as a unique modality of regulated cell death, requires the involvement of ubiquitin-binding enzyme UBE2D2. However, the prognostic and immunotherapeutic values of UBE2D2 in pan-cancer remain largely unknown.

Using UCSC Xena, TIMER, Clinical Proteomic Tumor Analysis Consortium (CPTAC), and Human Protein Atlas (HPA) databases, we aimed to explore the differential expression pattern of UBE2D2 across multiple cancer types and to evaluate its association with patient prognosis, clinical features, and genetic variations. The association between UBE2D2 and immunotherapy response was assessed by gene set enrichment analysis, tumor microenvironment, immune gene co-expression and drug half maximal inhibitory concentration (IC50) analysis.

The mRNA and protein levels of UBE2D2 were markedly elevated in most cancer types, and UBE2D2 exhibited prognostic significance in liver hepatocellular carcinoma (LIHC), kidney chromophobe (KICH), uveal melanomas (UVM), cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC), and kidney renal papillary cell carcinoma (KIRP). UBE2D2 expression was correlated with clinical features, tumor mutation burden, microsatellite instability, and anti-tumor drug resistance in several tumor types. Gene enrichment analysis showed that UBE2D2 was significantly associated with immune-related pathways. The expression level of UBE2D2 was correlated with immune cell infiltration, including CD4 + T cells、Macrophages M2、CD8 + T cells in pan-cancer. PDCD1, CD274 and CTLA4 expression levels were positively correlated with UBE2D2 level in multiple cancers.

We comprehensively investigated the potential value of UBE2D2 as a prognostic and immunotherapeutic predictor for pan-cancer, providing a novel insight for cancer immunotherapy.

Methylation, a key epigenetic modification, is essential for regulating gene expression and protein function without altering the DNA sequence, contributing to various biological processes, including gene transcription, embryonic development, and cellular functions. Methylation encompasses DNA methylation, RNA methylation and histone modification. Recent research indicates that DNA methylation is vital for establishing and maintaining normal brain functions by modulating the high-order structure of DNA. Alterations in the patterns of DNA methylation can exert significant impacts on both gene expression and cellular function, playing a role in the development of numerous diseases, such as neurological disorders, cardiovascular diseases as well as cancer. Our current understanding of the etiology of neurological diseases emphasizes a multifaceted process that includes neurodegenerative, neuroinflammatory, and neurovascular events. Epigenetic modifications, especially DNA methylation, are fundamental in the control of gene expression and are critical in the onset and progression of neurological disorders. Furthermore, we comprehensively overview the role and mechanism of DNA methylation in in various biological processes and gene regulation in neurological diseases. Understanding the mechanisms and dynamics of DNA methylation in neural development can provide valuable insights into human biology and potentially lead to novel therapies for various neurological diseases.

Prostate adenocarcinoma (PRAD) is a prevalent global malignancy which depends more on lipid metabolism for tumor progression compared to other cancer types. Although Stearoyl-coenzyme A desaturase (SCD) is documented to regulate lipid metabolism in multiple cancers, landscape analysis of its implications in PRAD are still missing at present. Here, we conducted an analysis of diverse cancer datasets revealing elevated SCD expression in the PRAD cohort at both mRNA and protein levels. Interestingly, the elevated expression was associated with SCD promoter hypermethylation and genetic alterations, notably the L134V mutation. Integration of comprehensive tumor immunological and genomic data revealed a robust positive correlation between SCD expression levels and the abundance of CD8+ T cells and macrophages. Further analyses identified significant associations between SCD expression and various immune markers in tumor microenvironment. Single-cell transcriptomic profiling unveiled differential SCD expression patterns across distinct cell types within the prostate tumor microenvironment. The Gene Ontology and Kyoto Encyclopedia of Genes and Genome analyses showed that SCD enriched pathways were primarily related to lipid biosynthesis, cholesterol biosynthesis, endoplasmic reticulum membrane functions, and various metabolic pathways. Gene Set Enrichment Analysis highlighted the involvement of elevated SCD expression in crucial cellular processes, including the cell cycle and biosynthesis of cofactors pathways. In functional studies, SCD overexpression promoted the proliferation, metastasis and invasion of prostate cancer cells, whereas downregulation inhibits these processes. This study provides comprehensive insights into the multifaceted roles of SCD in PRAD pathogenesis, underscoring its potential as both a therapeutic target and prognostic biomarker.

Due to the high heterogeneity of HCC, which leads to poor prognostic outcomes for patients, there is a need to develop a novel predictive model for accurate classification of HCC in order to improve patient survival rates.

The data of the HCV, cirrhosis, and HCC were obtained from TCGA and GEO databases. Multivariable Cox regression analysis and survival analysis was conducted to assess the prognostic relevance of these differentially expressed genes. Single-cell sequencing was used to explore the intercellular interaction patterns and identify relevant signaling pathways. Drug sensitivity analysis was conducted to determine personalized treatment strategies for patients.

In this study, we conducted integrated analysis of hepatitis, cirrhosis, and hepatocellular carcinoma datasets and identified 10 liver disease progression genes associated with prognosis. These genes exhibited significant downregulation in expression as the disease advanced, suggesting their crucial involvement in HCC development. By performing multivariable Cox analysis, we established a prognostic model for liver disease progression to predict the prognosis of HCC patients. The model was validated using ROC analysis, demonstrating good accuracy and stability in prognostic evaluation. Single-cell sequencing analysis revealed that these genes primarily exert their effects through the MIF signaling pathway during HCC progression. Furthermore, we observed that patients in the low-risk group exhibited higher sensitivity to TACE treatment, while patients in the high-risk group showed better response to sorafenib treatment.

In summary, we have elucidated the key genes involved in the progression of liver diseases and established a precise prognostic model for assessing the prognosis of HCC patients. Our study provides novel insights and strategies for the treatment of HCC.

Oligodendroglioma, IDH-mutant and 1p/19q-codeleted has highly variable outcomes that are strongly influenced by patient age. The distribution of oligodendroglioma age is non-Gaussian and reportedly bimodal, which motivated our investigation of age-associated molecular alterations that may drive poorer outcomes. We found that elevated HOXD12 expression was associated with both older patient age and shorter survival in the TCGA (FDR < 0.01, FDR = 1e-5) and the CGGA (p = 0.03, p < 1e-3). HOXD12 gene body hypermethylation was associated with older age, higher WHO grade, and shorter survival in the TCGA (p < 1e-6, p < 0.001, p < 1e-3) and with older age and higher WHO grade in Capper et al. (p < 0.002, p = 0.014). In the TCGA, HOXD12 gene body hypermethylation and elevated expression were independently prognostic of NOTCH1 and PIK3CA mutations, loss of 15q, MYC activation, and standard histopathological features. Single-nucleus RNA and ATAC sequencing data showed that HOXD12 activity was elevated in neoplastic tissue, particularly within cycling and OPC-like cells, and was associated with a stem-like phenotype. A pan-HOX DNA methylation analysis revealed an age and survival-associated HOX-high signature that was tightly associated with HOXD12 gene body methylation. Overall, HOXD12 expression and gene body hypermethylation were associated with an older, atypically aggressive subtype of oligodendroglioma.

Increasing evidence has shown that connexins are involved in the regulation of tumor development, immune escape, and drug resistance. This study investigated the gene expression patterns, prognostic values, and potential mechanisms of connexins in breast cancer.

We conducted a comprehensive analysis of connexins using public gene and protein expression databases and clinical samples from our institution. Connexin mRNA expressions in breast cancer and matched normal tissues were compared, and multiomics studies were performed.

Gap junction beta-2 mRNA was overexpressed in breast cancers of different pathological types and molecular subtypes, and its high expression was associated with poor prognosis. The tumor membrane of the gap junction beta-2 mutated group was positive, and the corresponding protein was expressed. Somatic mutation and copy number variation of gap junction beta-2 are rare in breast cancer. The gap junction beta-2 transcription level in the p110α subunit of the phosphoinositide 3-kinase mutant subgroup was higher than that in the wild-type subgroup. Gap junction beta-2 was associated with the phosphoinositide 3-kinase-Akt signaling pathway, extracellular matrix-receptor interaction, focal adhesion, and proteoglycans in cancer. Furthermore, gap junction beta-2 overexpression may be associated with phosphoinositide 3-kinase and histone deacetylase inhibitor resistance, and its expression level correlated with infiltrating CD8+ T cells, macrophages, neutrophils, and dendritic cells.

Gap junction beta-2 may be a promising therapeutic target for targeted therapy and immunotherapy and may be used to predict breast cancer prognosis.

Mutations in DNA regulatory regions are increasingly being recognized as important drivers of cancer and other complex diseases. These mutations can regulate gene expression by affecting DNA-protein binding and epigenetic profiles, such as DNA methylation in genome regulatory elements. However, identifying mutation hotspots associated with expression regulation and disease progression in non-coding DNA remains a challenge. Unlike most existing approaches that assign a mutation score to individual single nucleotide polymorphisms (SNP), a mutation block (MB)-based approach was introduced in this study to assess the collective impact of a cluster of SNPs on transcription factor-DNA binding affinity, differential gene expression (DEG), and nearby DNA methylation. Moreover, the long-distance target genes of functional MBs were identified using a new permutation-based algorithm that assessed the significance of correlations between DNA methylation at regulatory regions and target gene expression. Two new Python packages were developed. The Differential Methylation Region (DMR-analysis) analysis tool was used to detect DMR and map them to regulatory elements. The second tool, an integrated DMR, DEG, and SNP analysis tool (DDS-analysis), was used to combine the omics data to identify functional MBs and long-distance target genes. Both tools were validated in follicular lymphoma (FL) cohorts, where not only known functional MBs and their target genes (BCL2 and BCL6) were recovered, but also novel genes were found, including CDCA4 and JAG2, which may be associated with FL development. These genes are linked to target gene expression and are significantly correlated with the methylation of nearby DNA sequences in FL. The proposed computational integrative analysis of multiomics data holds promise for identifying regulatory mutations in cancer and other complex diseases.