BACH1 Stabilization by Antioxidants Stimulates Lung Cancer Metastasis.

For tumors to progress efficiently, cancer cells must overcome barriers of oxidative stress. Although dietary antioxidant supplementation or activation of endogenous antioxidants by NRF2 reduces oxidative stress and promotes early lung tumor progression, little is known about its effect on lung cancer metastasis. Here, we show that long-term supplementation with the antioxidants N-acetylcysteine and vitamin E promotes KRAS-driven lung cancer metastasis. The antioxidants stimulate metastasis by reducing levels of free heme and stabilizing the transcription factor BACH1. BACH1 activates transcription of Hexokinase 2 and Gapdh and increases glucose uptake, glycolysis rates, and lactate secretion, thereby stimulating glycolysis-dependent metastasis of mouse and human lung cancer cells. Targeting BACH1 normalized glycolysis and prevented antioxidant-induced metastasis, while increasing endogenous BACH1 expression stimulated glycolysis and promoted metastasis, also in the absence of antioxidants. We conclude that BACH1 stimulates glycolysis-dependent lung cancer metastasis and that BACH1 is activated under conditions of reduced oxidative stress.

Global loss of cellular m6A RNA methylation following infection with different SARS-CoV-2 variants.

Insights into host-virus interactions during SARS-CoV-2 infection are needed to understand COVID-19 pathogenesis and may help to guide the design of novel antiviral therapeutics. N 6-Methyladenosine modification (m6A), one of the most abundant cellular RNA modifications, regulates key processes in RNA metabolism during stress response. Gene expression profiles observed postinfection with different SARS-CoV-2 variants show changes in the expression of genes related to RNA catabolism, including m6A readers and erasers. We found that infection with SARS-CoV-2 variants causes a loss of m6A in cellular RNAs, whereas m6A is detected abundantly in viral RNA. METTL3, the m6A methyltransferase, shows an unusual cytoplasmic localization postinfection. The B.1.351 variant has a less-pronounced effect on METTL3 localization and loss of m6A than did the B.1 and B.1.1.7 variants. We also observed a loss of m6A upon SARS-CoV-2 infection in air/liquid interface cultures of human airway epithelia, confirming that m6A loss is characteristic of SARS-CoV-2-infected cells. Further, transcripts with m6A modification are preferentially down-regulated postinfection. Inhibition of the export protein XPO1 results in the restoration of METTL3 localization, recovery of m6A on cellular RNA, and increased mRNA expression. Stress granule formation, which is compromised by SARS-CoV-2 infection, is restored by XPO1 inhibition and accompanied by a reduced viral infection in vitro. Together, our study elucidates how SARS-CoV-2 inhibits the stress response and perturbs cellular gene expression in an m6A-dependent manner.

METTL3 drives telomere targeting of TERRA lncRNA through m6A-dependent R-loop formation: a therapeutic target for ALT-positive neuroblastoma.

Telomerase-negative tumors maintain telomere length by alternative lengthening of telomeres (ALT), but the underlying mechanism behind ALT remains poorly understood. A proportion of aggressive neuroblastoma (NB), particularly relapsed tumors, are positive for ALT (ALT+), suggesting that a better dissection of the ALT mechanism could lead to novel therapeutic opportunities. TERRA, a long non-coding RNA (lncRNA) derived from telomere ends, localizes to telomeres in a R-loop-dependent manner and plays a crucial role in telomere maintenance. Here we present evidence that RNA modification at the N6 position of internal adenosine (m6A) in TERRA by the methyltransferase METTL3 is essential for telomere maintenance in ALT+ cells, and the loss of TERRA m6A/METTL3 results in telomere damage. We observed that m6A modification is abundant in R-loop enriched TERRA, and the m6A-mediated recruitment of hnRNPA2B1 to TERRA is critical for R-loop formation. Our findings suggest that m6A drives telomere targeting of TERRA via R-loops, and this m6A-mediated R-loop formation could be a widespread mechanism employed by other chromatin-interacting lncRNAs. Furthermore, treatment of ALT+ NB cells with a METTL3 inhibitor resulted in compromised telomere targeting of TERRA and accumulation of DNA damage at telomeres, indicating that METTL3 inhibition may represent a therapeutic approach for ALT+ NB.

FOXK1 and FOXK2 regulate aerobic glycolysis.

Adaptation to the environment and extraction of energy are essential for survival. Some species have found niches and specialized in using a particular source of energy, whereas others-including humans and several other mammals-have developed a high degree of flexibility1. A lot is known about the general metabolic fates of different substrates but we still lack a detailed mechanistic understanding of how cells adapt in their use of basic nutrients2. Here we show that the closely related fasting/starvation-induced forkhead transcription factors FOXK1 and FOXK2 induce aerobic glycolysis by upregulating the enzymatic machinery required for this (for example, hexokinase-2, phosphofructokinase, pyruvate kinase, and lactate dehydrogenase), while at the same time suppressing further oxidation of pyruvate in the mitochondria by increasing the activity of pyruvate dehydrogenase kinases 1 and 4. Together with suppression of the catalytic subunit of pyruvate dehydrogenase phosphatase 1 this leads to increased phosphorylation of the E1α regulatory subunit of the pyruvate dehydrogenase complex, which in turn inhibits further oxidation of pyruvate in the mitochondria-instead, pyruvate is reduced to lactate. Suppression of FOXK1 and FOXK2 induce the opposite phenotype. Both in vitro and in vivo experiments, including studies of primary human cells, show how FOXK1 and/or FOXK2 are likely to act as important regulators that reprogram cellular metabolism to induce aerobic glycolysis.

Identifying novel inhibitors targeting Exportin-1 for the potential treatment of COVID-19.

The nuclear export protein 1 (XPO1) mediates the nucleocytoplasmic transport of proteins and ribonucleic acids (RNAs) and plays a prominent role in maintaining cellular homeostasis. XPO1 has emerged as a promising therapeutic approach to interfere with the lifecycle of many viruses. In our earlier study, we proved the inhibition of XPO1 as a therapeutic strategy for managing SARS-COV-2 and its variants. In this study, we have utilized pharmacophore-assisted computational methods to identify prominent XPO1 inhibitors. After several layers of screening, a few molecules were shortlisted for further experimental validation on the in vitro SARS-CoV-2 cell infection model. It was observed that these compounds reduced spike positivity, suggesting inhibition of SARS-COV-2 infection. The outcome of this study could be considered further for developing novel antiviral therapeutic strategies against SARS-CoV-2.

Gas chromatography mass spectrometer determination of dimethylamine impurity in N,N-dimethylformamide solvent by derivatization of N,N-dimethylbenzamide with benzoyl chloride agent.

This study describes the development of a reliable and linear analytical method for precisely determining dimethylamine impurity in N,N-dimethylformamide solvent utilizing a benzoyl chloride derivatization reagent and a gas chromatography mass spectrometer. Benzoyl chloride was used to derivatize dimethylamine. At normal temperature, benzoyl chloride combined with dimethylamine, producing N,N-dimethylbenzamide. This method separated N,N-dimethylbenzamide using Rtx-5 amine (30 m × 0.32 mm × 1.50 µm) as the stationary phase, helium as the carrier gas, argon as the collision gas, and methanol as the diluent. The column flow rate was 2 mL/min. The retention time of N,N-dimethylbenzamide was determined to be 8.5 min. Precision, linearity, and accuracy were tested using ICH Q2 (R2) and USP<1225> guidelines. The percentage coefficient of variation (CV) for N,N-dimethylbenzamide in the system suitability parameter was 1.1%. The correlation coefficient of N,N-dimethylbenzamide was found to be >0.99. In the method precision parameter, the % CV for N,N-dimethylbenzamide was found to be 1.9%, whereas the % CV for N,N-dimethylbenzamide was 1.2% in intermediate precision. The percentage recovery of N,N-dimethylbenzamide was determined to be between 80% and 98%.

Analytical determination of ethylenediamine impurity in tripelennamine hydrochloride by gas chromatography-mass spectrometry using phthalaldehyde as the derivatizing agent.

In the pharmaceutical industry, effective risk management and control strategies for potential genotoxic impurities are of paramount importance. The current study utilized GC-MS to evaluate a precise, linear, and accurate analytical method for quantifying ethylenediamine present in tripelennamine hydrochloride using phthalaldehyde as a derivatizing agent. When phthalaldehyde is sonicated for 10 min at room temperature, it reacts with ethylenediamine to form (1z,5z)-3,4-dihydrobenzo[f][1,4]diazocine. This approach minimizes matrix interference issues and resolves sample preparation difficulties encountered during ethylenediamine identification in GC-MS. In this method, helium serves as the carrier gas, while methanol acts as the diluent. The stationary phase consists of a DB-5MS column (30 m × 0.25 mm × 0.25 µm) with a flow rate of 1.5 mL/min. The retention time of (1z,5z)-3,4-dihydrobenzo[f][1,4]diazocine was determined to be 6.215 min. The method validation demonstrated limits of detection and quantification for (1z,5z)-3,4-dihydrobenzo[f][1,4]diazocine at 0.4 and 1.0 ppm, respectively, with a linearity range spanning from 1 to 30 ppm concentration with respect to the specification level. System suitability, precision, linearity, and accuracy of the current method were assessed in accordance with guidelines, yielding results deemed suitable for the intended use.

Electroweak Multi-Higgs Production: A Smoking Gun for the Type-I Two-Higgs-Doublet Model.

Extending the Higgs sector of the standard model (SM) by just one additional Higgs doublet field leads to the two-Higgs-doublet model (2HDM). In the type-I Z_{2}-symmetric limit of the 2HDM, all the five new physical Higgs states can be fairly light, O(100) GeV or less, without being in conflict with current data from the direct Higgs boson searches and the B-physics measurements. In this Letter, we establish that the new neutral as well as the charged Higgs bosons in this model can all be simultaneously observable in the multi-b final state. The statistical significance of the signature for each of these Higgs states, resulting from the electroweak (EW) production of their pairs, can exceed 5σ at the 13 TeV high-luminosity Large Hadron collider (HL-LHC). Since the parameter space configurations where this is achievable are precluded in the other, more extensively pursued, 2HDM types, an experimental validation of our findings would be a clear indication that the true underlying Higgs sector in nature is the type-I 2HDM.

Insights on the pathogenesis of type 2 diabetes as revealed by signature genomic classifiers in an African American population in the Washington, DC area.

AIMS: African Americans (AA) in the United States have a high risk of type 2 diabetes mellitus (T2DM) and suffer from disparities in the prevalence, mortality, and comorbidities of the disease compared to other Americans. The present study aimed to shed light on the molecular mechanisms of disease pathogenesis of T2DM among AA in the Washington, DC region. METHODS: We performed TaqMan Low Density Arrays (TLDA) on 24 genes of interest that belong to three categories: metabolic disease and disorders, cancer-related genes, and neurobehavioural disorders genes. The 18 genes, viz. ARNT, CYP2D6, IL6, INSR, RRAD, SLC2A2 (metabolic disease and disorders), APC, BCL2, CSNK1D, MYC, SOD2, TP53 (Cancer-related), APBA1, APBB2, APOC1, APOE, GSK3B, and NAE1 (neurobehavioural disorders), were differentially expressed in T2DM participants compared to controls. RESULTS: Our results suggest that factors including gender, smoking habits, and the severity or lack of control of T2DM (as indicated by HbA1c levels) were significantly associated with differential gene expression. APBA1 was significantly (p-value <0.05) downregulated in all diabetes participants. Upregulation of APOE and CYP2D6 genes and downregulation of the INSR gene were observed in the majority of diabetes patients. CONCLUSIONS: Tobacco smoking and gender were significantly associated with case-control differences in expression of the APBA1 and APOE genes (connected with Alzheimer's disease) and the INSR and CYP2D6 (associated with metabolic disorders). The results highlight the need for more effective management of T2DM and for tobacco smoking cessation interventions in this community, and further research on the associations of T2DM with other disease processes, including cancer and neurobehavioral pathways.

Transcriptomics of MASLD Pathobiology in African American Patients in the Washington DC Area †.

Metabolic-dysfunction-associated steatotic liver disease (MASLD) is becoming the most common chronic liver disease worldwide and is of concern among African Americans (AA) in the United States. This pilot study evaluated the differential gene expressions and identified the signature genes in the disease pathways of AA individuals with MASLD. Blood samples were obtained from MASLD patients (n = 23) and non-MASLD controls (n = 24) along with their sociodemographic and medical details. Whole-blood transcriptomic analysis was carried out using Affymetrix Clarion-S Assay. A validation study was performed utilizing TaqMan Arrays coupled with Ingenuity Pathway Analysis (IPA) to identify the major disease pathways. Out of 21,448 genes in total, 535 genes (2.5%) were significantly (p < 0.05) and differentially expressed when we compared the cases and controls. A significant overlap in the predominant differentially expressed genes and pathways identified in previous studies using hepatic tissue was observed. Of note, TGFB1 and E2F1 genes were upregulated, and HMBS was downregulated significantly. Hepatic fibrosis signaling is the top canonical pathway, and its corresponding biofunction contributes to the development of hepatocellular carcinoma. The findings address the knowledge gaps regarding how signature genes and functional pathways can be detected in blood samples ('liquid biopsy') in AA MASLD patients, demonstrating the potential of the blood samples as an alternative non-invasive source of material for future studies.

Peroxisome Proliferator-Activated Receptor-α (PPARα) Expression in a Clinical Population of Pakistani Patients with Type 2 Diabetes and Dyslipidemia.

Poor glycemic control and dyslipidemia are hallmarks of type 2 diabetes mellitus (T2DM), which predispose to cardiovascular diseases. Peroxisome proliferator-activated receptor-α (PPARα) has been associated with atherosclerosis, but its role in T2DM is less clear. Previously, we studied PPARα expression levels in diabetics with and without dyslipidemia (DD). In this study we described the association with fasting blood glucose, HbA1c levels and lipid levels of the study population. Patient demography and biochemical data were collected from hospitals in Islamabad, Pakistan, and RT-PCR data of PPARα expression were retrieved from our previous study from the same cohort. We performed t-tests and regression analysis to evaluate the relationships between PPARα expression and demographic and clinical variables. As expected, body mass index and HbA1c were elevated in T2DM and DD patients compared to controls. Blood lipids (total cholesterol, triglycerides, LDL and HDL) were significantly higher in the DD group compared to the other two groups. In the T2DM and DD groups, the PPARα expression was not associated with any of the physical and biochemical parameters measured in this study. Expression of the PPARα gene was independent of blood lipids and glycemic control in this study. Further research is necessary to better understand the biological parameters of PPARα expression.

The long noncoding RNA TUNAR modulates Wnt signaling and regulates human ß-cell proliferation.

Many long noncoding RNAs (lncRNAs) are enriched in pancreatic islets and several lncRNAs are linked to type 2 diabetes (T2D). Although they have emerged as potential players in ß-cell biology and T2D, little is known about their functions and mechanisms in human ß-cells. We identified an islet-enriched lncRNA, TUNAR (TCL1 upstream neural differentiation-associated RNA), which was upregulated in ß-cells of patients with T2D and promoted human ß-cell proliferation via fine-tuning of the Wnt pathway. TUNAR was upregulated following Wnt agonism by a glycogen synthase kinase-3 (GSK3) inhibitor in human ß-cells. Reciprocally, TUNAR repressed a Wnt antagonist Dickkopf-related protein 3 (DKK3) and stimulated Wnt pathway signaling. DKK3 was aberrantly expressed in ß-cells of patients with T2D and displayed a synchronized regulatory pattern with TUNAR at the single cell level. Mechanistically, DKK3 expression was suppressed by the repressive histone modifier enhancer of zeste homolog 2 (EZH2). TUNAR interacted with EZH2 in ß-cells and facilitated EZH2-mediated suppression of DKK3. These findings reveal a novel cell-specific epigenetic mechanism via islet-enriched lncRNA that fine-tunes the Wnt pathway and subsequently human ß-cell proliferation.NEW & NOTEWORTHY The discovery that long noncoding RNA TUNAR regulates ß-cell proliferation may be important in designing new treatments for diabetes.

H3K4me2 and WDR5 enriched chromatin interacting long non-coding RNAs maintain transcriptionally competent chromatin at divergent transcriptional units.

Recently lncRNAs have been implicated in the sub-compartmentalization of eukaryotic genome via genomic targeting of chromatin remodelers. To explore the function of lncRNAs in the maintenance of active chromatin, we characterized lncRNAs from the chromatin enriched with H3K4me2 and WDR5 using chromatin RNA immunoprecipitation (ChRIP). Significant portion of these enriched lncRNAs were arranged in antisense orientation with respect to their protein coding partners. Among these, 209 lncRNAs, commonly enriched in H3K4me2 and WDR5 chromatin fractions, were named as active chromatin associated lncRNAs (active lncCARs). Interestingly, 43% of these active lncCARs map to divergent transcription units. Divergent transcription (XH) units were overrepresented in the active lncCARs as compared to the inactive lncCARs. ChIP-seq analysis revealed that active XH transcription units are enriched with H3K4me2, H3K4me3 and WDR5. WDR5 depletion resulted in the loss of H3K4me3 but not H3K4me2 at the XH promoters. Active XH CARs interact with and recruit WDR5 to XH promoters, and their depletion leads to decrease in the expression of the corresponding protein coding genes and loss of H3K4me2, H3K4me3 and WDR5 at the active XH promoters. This study unravels a new facet of chromatin-based regulation at the divergent XH transcription units by this newly identified class of H3K4me2/WDR5 chromatin enriched lncRNAs.

Novel Triazole linked 2-phenyl benzoxazole derivatives induce apoptosis by inhibiting miR-2, miR-13 and miR-14 function in Drosophila melanogaster.

Apoptosis is an important phenomenon in multi cellular organisms for maintaining tissue homeostasis and embryonic development. Defect in apoptosis leads to a number of disorders like- autoimmune disorder, immunodeficiency and cancer. 21-22 nucleotides containing micro RNAs (miRNAs/miRs) function as a crucial regulator of apoptosis alike other cellular pathways. Recently, small molecules have been identified as a potent inducer of apoptosis. In this study, we have identified novel Triazole linked 2-phenyl benzoxazole derivatives (13j and 13h) as a negative regulator of apoptosis inhibiting micro RNAs (miR-2, miR-13 and miR-14) in a well established in vivo model Drosophila melanogaster where the process of apoptosis is very similar to human apoptosis. These compounds inhibit miR-2, miR-13 and miR-14 activity at their target sites, which induce an increased caspase activity, and in turn influence the caspase dependent apoptotic pathway. These two compounds also increase the mitochondrial reactive oxygen species (ROS) level to trigger apoptotic cell death.

Kcnq1ot1 antisense noncoding RNA mediates lineage-specific transcriptional silencing through chromatin-level regulation.

Recent investigations have implicated long antisense noncoding RNAs in the epigenetic regulation of chromosomal domains. Here we show that Kcnq1ot1 is an RNA polymerase II-encoded, 91 kb-long, moderately stable nuclear transcript and that its stability is important for bidirectional silencing of genes in the Kcnq1 domain. Kcnq1ot1 interacts with chromatin and with the H3K9- and H3K27-specific histone methyltransferases G9a and the PRC2 complex in a lineage-specific manner. This interaction correlates with the presence of extended regions of chromatin enriched with H3K9me3 and H3K27me3 in the Kcnq1 domain in placenta, whereas fetal liver lacks both chromatin interactions and heterochromatin structures. In addition, the Kcnq1 domain is more often found in contact with the nucleolar compartment in placenta than in liver. Taken together, our data describe a mechanism whereby Kcnq1ot1 establishes lineage-specific transcriptional silencing patterns through recruitment of chromatin remodeling complexes and maintenance of these patterns through subsequent cell divisions occurs via targeting the associated regions to the perinucleolar compartment.

Long noncoding RNA-mediated maintenance of DNA methylation and transcriptional gene silencing.

Establishment of silencing by noncoding RNAs (ncRNAs) via targeting of chromatin remodelers is relatively well investigated; however, their role in the maintenance of silencing is poorly understood. Here, we explored the functional role of the long ncRNA Kcnq1ot1 in the maintenance of transcriptional gene silencing in the one mega-base Kcnq1 imprinted domain in a transgenic mouse model. By conditionally deleting the Kcnq1ot1 ncRNA at different stages of mouse development, we suggest that Kcnq1ot1 ncRNA is required for the maintenance of the silencing of ubiquitously imprinted genes (UIGs) at all developmental stages. In addition, Kcnq1ot1 ncRNA is also involved in guiding and maintaining the CpG methylation at somatic differentially methylated regions flanking the UIGs, which is a hitherto unknown role for a long ncRNA. On the other hand, silencing of some of the placental-specific imprinted genes (PIGs) is maintained independently of Kcnq1ot1 ncRNA. Interestingly, the non-imprinted genes (NIGs) that escape RNA-mediated silencing are enriched with enhancer-specific modifications. Taken together, this study illustrates the gene-specific maintenance mechanisms operational at the Kcnq1 locus for tissue-specific transcriptional gene silencing and activation.

Maintenance of epigenetic information: a noncoding RNA perspective.

Along the lines of established players like chromatin modifiers and transcription factors, noncoding RNA (ncRNA) are now widely accepted as one of the key regulatory molecules in epigenetic regulation of transcription. With increasing evidence of ncRNAs in the establishment of gene silencing through their ability to interact with major chromatin modifiers, in the current review, we discuss their prospective role in the area of inheritance and maintenance of these established silenced states which can be reversible or irreversible in nature. In addition, we attempt to understand and speculate how these RNA dependent or independent maintenance mechanisms differ between each other in a developmental stage, tissue, and gene-specific manner in different biological contexts by utilizing known/unknown regulatory factors.

Headspace Gas Chromatography Mass Spectrometry Method for Determination of class-I Residual Solvents in Several Drug Substances: Method Evaluation by Quality by Design Statistical Tool.

BACKGROUND: Class-I residual solvents such as 1,1-dichloroethene, 1,1,1-trichloroethane, carbon tetrachloride, benzene, 1,2-dichloroethane are toxic, environmental hazard and carcinogenic to human. Headspace gas chromatography mass spectrometer is the sophisticated instrument for quantification of residual solvents at lower limits. OBJECTIVE: An exact, sensitive, reliable and fast method was developed to determine the 1,1-dichloroethene, 1,1,1-trichloroethane, carbon tetrachloride, benzene and 1,2-dichloroethane present in different drug substances by using a headspace gas chromatography mass spectrometer. METHOD: Helium is used as a carrier gas in this method. N-Methyl-2-pyrrolidone is used as a diluent and the stationary phase is a DB-624 (60 m x 0.25 mm x 1.4 µm film thickness) column with a flow rate of 1.5 mL/min. RESULTS: The concentration of limit of detection for 1,1-dichloroethene, 1,1,1-trichloroethane, carbon tetrachloride, benzene and 1,2-dichloroethane were achieved with 0.24 ppm, 5 ppm, 0.12 ppm, 0.06 ppm, 0.15 ppm. Subsequently, the concentrations of limit of quantification for aforementioned impurities were achieved with 0.8 ppm, 15 ppm, 0.4 ppm, 0.2 ppm, 0.5 ppm. The linearity was covered in the range of LOQ to 120% with respect to the specification level. CONCLUSION: The current method's system suitability, precision, linearity, accuracy parameters were performed in accordance to the USP < 1225> and ICH Q2(R2) and results were lies within the acceptance criteria. HIGHLIGHTS: No research studies are reported for the determination of class-I residual solvents in lincomycin hydrochloride, dapagliflozin, vonoprazan fumarate, telmisartan drug substances. The proposed study is to develop the common method for quantification of class-I residual solvents for drug substances. QbD concept is utilized in the method performance verification.

Transcriptomic Analysis of Alzheimer's Disease Pathways in a Pakistani Population.

Background: Alzheimer's disease (AD) is a multifactorial neurodegenerative disorder that is most prevalent in elderly individuals, especially in developed countries, and its prevalence is now increasing in developing countries like Pakistan. Objective: Our goal was to characterize key genes and their levels of expression and related molecular transcriptome networks associated with AD pathogenesis in a pilot case-control study in a Pakistani population. Methods: To obtain the spectrum of molecular networks associated with pathogenesis in AD patients in Pakistan (comparing cases and controls), we used high-throughput qRT-PCR (TaqMan Low-Density Array; n = 33 subjects) coupled with Affymetrix Arrays (n = 8) and Ingenuity Pathway Analysis (IPA) to identify signature genes associated with Amyloid processing and disease pathways. Results: We confirmed 16 differentially expressed AD-related genes, including maximum fold changes observed in CAPNS2 and CAPN1. The global gene expression study observed that 61% and 39% of genes were significantly (p-value 0.05) up- and downregulated, respectively, in AD patients compared to healthy controls. The key pathways include, e.g., Amyloid Processing, Neuroinflammation Signaling, and ErbB4 Signaling. The top-scoring networks in Diseases and Disorders Development were Neurological Disease, Organismal Injury and Abnormalities, and Psychological Disorders. Conclusions: Our pilot study offers a non-invasive and efficient way of investigating gene expression patterns by combining TLDA and global gene expression method in AD patients by utilizing whole blood. This provides valuable insights into the expression status of genes related to Amyloid Processing, which could play potential role in future studies to identify sensitive, early biomarkers of AD in general.

Antisense noncoding RNA promoter regulates the timing of de novo methylation of an imprinting control region.

Epigenetic marks at cis acting imprinting control regions (ICRs) regulate parent of origin-specific expression of multiple genes in imprinted gene clusters. Epigenetic marks are acquired during gametogenesis and maintained faithfully thereafter. However, the mechanism by which differential epigenetic marks are established and maintained at ICRs is currently unclear. By using Kcnq1 ICR as a model system, we have investigated the functional role of genetic signatures in the acquisition and maintenance of epigenetic marks. Kcnq1 ICR is methylated on the maternal chromosome but remains unmethylated on the paternal chromosome. Here, we show that a paternal allele of Kcnq1 ICR lacking the Kcnq1ot1 promoter remains unmethylated during spermatogenesis; however, it becomes methylated specifically during pre-implantation development. Analysis of the chromatin structure at the paternal ICR in spermatogenic cells and in E13.5 embryonic tissues revealed that the ICRs of both wild type and mutant mice are enriched with H3K4me2 in spermatiogenic cells of the testicular compartment, but the mutant ICR lost H3K4me2 specifically in epididymal sperm and an increase in repressive marks was observed in embryonic tissues. Interestingly, we also detected a decrease in nucleosomal histone levels at the mutant ICR in comparison to the wild-type ICR in epididymal sperm. Taken together, these observations suggest that the Kcnq1ot1 promoter plays a critical role in establishing an epigenetic memory in the male germline by ensuring that the paternal allele remains in an unmethylated state during pre-implantation development.