Downregulation Of Specific Genes Research Articles

Integrative single-cell analysis: dissecting CD8 + memory cell roles in LUAD and COVID-19 via eQTLs and Mendelian Randomization

Lung adenocarcinoma exhibits high incidence and mortality rates, presenting a significant health concern. Concurrently, the COVID-19 pandemic has emerged as a grave global public health challenge. Existing literature suggests that T cells, pivotal components of cellular immunity, are integral to both antiviral and antitumor responses. Yet, the nuanced alterations and consequent functions of T cells across diverse disease states have not been comprehensively elucidated. We gathered transcriptomic data of peripheral blood mononuclear cells from lung adenocarcinoma patients, COVID-19 patients, and healthy controls. We followed a standardized analytical approach for quality assurance, batch effect adjustments, and preliminary data processing. We discerned distinct T cell subsets and conducted differential gene expression analysis. Potential key genes and pathways were inferred from GO and Pathway enrichment analyses. Additionally, we implemented Mendelian randomization to probe the potential links between pivotal genes and lung adenocarcinoma susceptibility. Our findings underscored a notable reduction in mature CD8 + central memory T cells in both lung adenocarcinoma and COVID-19 cohorts relative to the control group. Notably, the downregulation of specific genes, such as TRGV9, could impede the immunological efficacy of CD8 + T cells. Comprehensive multi-omics assessment highlighted genetic aberrations in genes, including TRGV9, correlating with heightened lung adenocarcinoma risk. Through rigorous single-cell transcriptomic analyses, this investigation meticulously delineated variations in T cell subsets across different pathological states and extrapolated key regulatory genes via an integrated multi-omics approach, establishing a robust groundwork for future functional inquiries. This study furnishes valuable perspectives into the etiology of multifaceted diseases and augments the progression of precision medicine.

An updated review on cell signaling pathways regulated by candidate miRNAs in coronary artery disease

MicroRNAs (miRNAs) are small endogenous non-coding RNA, size range from 17 to 25 nucleotides that regulate gene expression at the post-transcriptional level. More than 2000 different types of miRNAs have been identified in humans which regulate about 60% of gene expression, since the discovery of the first miRNA in 1993. MicroRNA performs many functions such as being involved in the regulation of various biological pathways for example cell migration, cell proliferation, cell differentiation, disease progression, and initiation. miRNAs also play an important role in the development of atherosclerosis lesions, cardiac fibroblast, cardiac hypertrophy, cancer, and neurological disorders. Abnormal activation of many cell signaling pathways has been observed in the development of coronary artery disease. Abnormal expression of these candidate miRNA genes leads to up or downregulation of specific genes, these specific genes play an important role in the regulation of cell signaling pathways involved in coronary artery disease. Many studies have found that miRNAs play a key role in the regulation of crucial signaling pathways that are involved in the pathophysiology of coronary artery disease. This review is designed to investigate the role of cell signaling pathways regulated by candidate miRNAs in Coronary artery disease.

Deciphering the antibiofilm potential of 2-Phenylethyl methyl ether (PEME), a bioactive compound of Kewda essential oil against Staphylococcus aureus

Opportunistic pathogenic bacteria and their pathogenicity linked with biofilm infections become a severe issue as they resist the actions of multiple antimicrobial drugs. Naturally derived drugs having antibiofilm properties are more effective than chemically synthesized drugs. The plant derived essential oils are a rich source of phytoconstituents with widespread pharmacological values. In the present investigation, a major phytoconstituent, 2-Phenyl Ethyl Methyl Ether (PEME) of Kewda essential oil extracted from the flowers of Pandanus odorifer was explored for its prospective antimicrobial and anti-biofilm properties against ESKAPE pathogenic bacterial strains, Staphylococcus aureus and MTCC 740. The minimum inhibitory concentration (MIC) of PEME was found to be 50 mM against the tested bacterial strains. A gradual decrease in biofilm production was observed when PEME was treated with the sub-MIC concentration. The reduction in biofilm formation was noticeable from qualitative assay i.e., Congo Red Agar Assay (CRA) and further quantified by crystal violet staining assay. The decline in exopolysaccharides production was quantified, with the highest inhibition against MTCC 740 with a decrease of 71.76 ± 4.56% compared to untreated control. From the microscopic analysis (light and fluorescence microscopic method), PEME exhibited inhibitory effect on biofilm formation on the polystyrene surface. The in silico studies stated that PEME could invariably bind to biofilm associated target proteins. Further, transcriptomic data analysis suggested the role of PEME in the down-regulation of specific genes, agrA, sarA, norA and mepR, which are critically associated with bacterial virulence, biofilm dynamics and drug resistance patterns in S. aureus. Further, qRT-PCR analysis validated the role of PEME on biofilm inhibition by relative downregulation of agrA, sarA, norA and mepR genes. Further, advanced in silico methodologies could be employed in future investigations to validate its candidature as promising anti-biofilm agent.

Chromosomal Rearrangements and Altered Nuclear Organization: Recent Mechanistic Models in Cancer.

Simple SummaryNew methodologies and technologies developed in the last few decades have highlighted the precise spatial organization of the genome into the cell nucleus, with chromatin architecture playing a central role in controlling several genome functions. Genes are expressed in a well-defined way and at a well-defined time during cell differentiation, and alterations in genome organization can lead to genetic diseases, such as cancers. Here we review how the genome is organized in the cell nucleus and the evidence of genome misorganization leading to cancer diseases.The last decade has seen significant progress in understanding how the genome is organized spatially within interphase nuclei. Recent analyses have confirmed earlier molecular cytogenetic studies on chromosome positioning within interphase nuclei and provided new information about the topologically associated domains (TADs). Examining the nuances of how genomes are organized within interphase nuclei will provide information fundamental to understanding gene regulation and expression in health and disease. Indeed, the radial spatial positioning of individual gene loci within nuclei has been associated with up- and down-regulation of specific genes, and disruption of normal genome organization within nuclei will result in compromised cellular health. In cancer cells, where reorganization of the nuclear architecture may occur in the presence of chromosomal rearrangements such as translocations, inversions, or deletions, gene repositioning can change their expression. To date, very few studies have focused on radial gene positioning and the correlation to gene expression in cancers. Further investigations would improve our understanding of the biological mechanisms at the basis of cancer and, in particular, in leukemia initiation and progression, especially in those cases where the molecular consequences of chromosomal rearrangements are still unclear. In this review, we summarize the main milestones in the field of genome organization in the nucleus and the alterations to this organization that can lead to cancer diseases.

Differential root and shoot magnetoresponses in Arabidopsis thaliana

The geomagnetic field (GMF) is one of the environmental stimuli that plants experience continuously on Earth; however, the actions of the GMF on plants are poorly understood. Here, we carried out a time-course microarray experiment to identify genes that are differentially regulated by the GMF in shoot and roots. We also used qPCR to validate the activity of some genes selected from the microarray analysis in a dose-dependent magnetic field experiment. We found that the GMF regulated genes in both shoot and roots, suggesting that both organs can sense the GMF. However, 49% of the genes were regulated in a reverse direction in these organs, meaning that the resident signaling networks define the up- or downregulation of specific genes. The set of GMF-regulated genes strongly overlapped with various stress-responsive genes, implicating the involvement of one or more common signals, such as reactive oxygen species, in these responses. The biphasic dose response of GMF-responsive genes indicates a hormetic response of plants to the GMF. At present, no evidence exists to indicate any evolutionary advantage of plant adaptation to the GMF; however, plants can sense and respond to the GMF using the signaling networks involved in stress responses.

Energy Metabolism in the Inner Retina in Health and Glaucoma

Glaucoma, the leading cause of irreversible blindness, is a heterogeneous group of diseases characterized by progressive loss of retinal ganglion cells (RGCs) and their axons and leads to visual loss and blindness. Risk factors for the onset and progression of glaucoma include systemic and ocular factors such as older age, lower ocular perfusion pressure, and intraocular pressure (IOP). Early signs of RGC damage comprise impairment of axonal transport, downregulation of specific genes and metabolic changes. The brain is often cited to be the highest energy-demanding tissue of the human body. The retina is estimated to have equally high demands. RGCs are particularly active in metabolism and vulnerable to energy insufficiency. Understanding the energy metabolism of the inner retina, especially of the RGCs, is pivotal for understanding glaucoma’s pathophysiology. Here we review the key contributors to the high energy demands in the retina and the distinguishing features of energy metabolism of the inner retina. The major features of glaucoma include progressive cell death of retinal ganglions and optic nerve damage. Therefore, this review focuses on the energetic budget of the retinal ganglion cells, optic nerve and the relevant cells that surround them.

Conditional knockdown of integrin beta-3 reveals its involvement in osteolytic and soft tissue lesions of breast cancer skeletal metastasis

Integrin β3 (ITGB3) is probably related to skeletal metastasis, which is the most frequent complication in breast cancer progression. We aimed to define its role and suitability as target for anti-metastatic therapy. We generated two MDA-MB-231 cell clones with conditional miRNA-mediated ITGB3 knockdown for analyzing the resulting effects in vitro regarding mRNA expression, proliferation and migration, as well the impact on skeletal metastasis in a nude rat model. Furthermore, ITGB3 levels were analyzed in exosomes from plasma of rats with skeletal metastases, and from MDA-MB-231 cells incubated with these vesicles, as well as from exosomes secreted by cells with conditional ITGB3 knockdown. This inhibition of ITGB3 expression decreased cellular proliferation and more distinctly inhibited cellular migration. Reduction and even complete remissions of respective soft tissue and osteolytic lesions were detected after ITGB3 knockdown in vivo. Furthermore, ITGB3 levels were increased in exosomes isolated from plasma of rats harboring MDA-MB-231 lesions as well as in respective cells incubated with these vesicles in vitro. ITGB3 was distinctly decreased in exosomes from cells with ITGB3 knockdown. The observed in vitro and in vivo anti-ITGB3 effects can be explained by downregulation of specific genes, which have roles in angiogenesis (NPTN, RRM2), tumor growth (NPTN), energy metabolism (ISCA1), cytokinesis (SEPT11), migration (RRM2, STX6), cell proliferation, invasiveness, senescence, tumorigenesis (RRM2) and vesicle trafficking (SEPT11, STX6). ITGB3 has a role in breast cancer skeletal metastasis via gene expression modulation, as mirrored for ITGB3 in exosomes, thus it could serve as target for anti-metastatic therapy.

Abstract 2962: Molecular mechanisms underlying the sensitizing effect of mutant p53 protein expression for Auranofin treatment of NSCLC and PDAC cells

Abstract Introduction: Auranofin (AF) is an orally available, organogold compound that is FDA-approved for its use in rheumatoid arthritis. AF gained interest as an anti-cancer agent since it acts as a thioredoxin reductase (TrxR) inhibitor. Data extracted from The Cancer Genome Atlas show that TXNRD1 is abundantly expressed in lung adenocarcinoma patients and negatively affects their outcome, making TrxR1 an attractable target in non-small cell lung cancer (NSCLC) patients. The goal of this study was to determine if mutant p53 expression can be used as a predictive biomarker for the treatment of AF in NSCLC. In addition, we aimed to obtain a profound understanding of the molecular mechanisms that govern the sensitizing effect of mutant p53 towards AF-treated NSCLC cells by investigating the type of cell death. Material and methods: We determined IC50-values using the sulforhodamine-B assay for AF in a panel of NSCLC and pancreatic ductal adenocarcinoma (PDAC) cell lines with different p53 mutations and expression levels. In addition, we correlated these IC50-values to baseline expression levels of different antioxidant related proteins. Results were confirmed in the NSCLC cell line NCI-H1299 (p53 deletion, null) and its two isogenic derivates (mutant p53 R175H or R273H knock-in). mRNA-Seq of the isogenic cell line panel was performed after exposure to a medium dose of AF. Finally, the IncuCyte ZOOM system was used to determine the percentage of AF-induced cell death by the means of the Cytotox Green reagent after pretreatment of the tumor cells with a panel of apoptosis, necroptosis and ferroptosis inhibitors. Results: A strong inverse correlation was observed between mutant p53 protein expression and AF IC50-values and baseline Trx levels in the NSCLC and PDAC cells, indicating that mutant p53 can sensitize cancer cells for AF. This sensitizing effect was confirmed in the p53Mutant isogenic NSCLC cells. Transcriptome analysis revealed the up- and downregulation of specific genes involved in ferroptosis. More specifically, AF induced cell death through various mechanisms, including apoptosis and ferroptosis, which were dependent on the expression levels of p53 rather than the type of p53 mutation. In cells with high expression levels of mutant p53, AF induced ferroptosis and lipid peroxidation. At lower expression levels of p53, the apoptosis pathway was triggered by AF with an increase in caspase-3/7 positive cells and inhibition of cell death by the caspase inhibitor Z-VAD-FMK. Conclusion: Mutant p53 protein expression renders NSCLC and PDAC cells more susceptible to AF. In an isogenic setting, medium levels of mutant p53 sensitize NSCLC cells for apoptosis while cells with higher levels are effectively killed through ferroptosis. Altogether, AF presents a potential novel therapeutic strategy to efficiently kill p53 mutated NSCLC tumor cells by various types of cell death. Citation Format: Laurie Freire Boullosa, Filip Lardon, Evelien Smits, Christophe Deben. Molecular mechanisms underlying the sensitizing effect of mutant p53 protein expression for Auranofin treatment of NSCLC and PDAC cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2962.

Changes in gene expression and metabolic profile of drupes of Olea europaea L. cv Carolea in relation to maturation stage and cultivation area

BackgroundOlive (Olea europaea L.) is an emblematic oil tree crop in the Mediterranean basin. Currently, despite olive features as a moderately thermophilic species, its cultivation is worldwide spreading due to the health-related impact of olive products on human nutrition. A point of concern for the expanding olive cultivation is related to the influence that, in addition to genotype, environmental factors exerts on drupe development and metabolism with consequent impact on fruit key traits. In this context, the aim of the present work was to gain further information on the genetic networks controlling drupe maturation phase and, mainly, on their modulation in response to environmental cues.ResultsTo achieve this goal, a comparative transcriptome-wide investigation was carried out on drupes of Olea europaea cultivar Carolea, collected from plants growing in areas at different altitude level and therefore experiencing different climatic conditions. Two maturation stages of drupe were analysed: green mature and turning-purple. Metabolic characterization of drupe was also performed. At both transcriptomic and metabolic level differences were detected in the pathway of fatty acids (FAs) and phenol compounds, in relation to both drupe maturation stage and cultivation area. Among the most relevant differences detected during the transition from GM to TP stages there were: the upregulation of FADs genes in the drupes of population growing at 700 masl, the upregulation of phenol biosynthesis-related genes in drupes growing at 10 and 200 masl and very interestingly the downregulation of specific genes involved in secoiridoids production in drupes growing at 700 masl. Globally, these results suggested that stability of FAs and phenols, mainly of secoiridoids group, is promoted at high altitude, while at lower altitude phenol biosynthesis is prolonged.ConclusionThe obtained results showed a differential modulation of genetic pathways related to olive compound quality in relation to the cultivation area, likely imposed by the different temperature impending at each altitude. The derived molecular information appears of interest for both breeding and biotechnological programs of olive species, especially with respect to the modulation of antioxidant secoiridoid compounds which play a key role in conferring both sensorial and healthy characteristic to olive products.

Gene expression profile in long-term non progressor HIV infected patients: In search of potential resistance factors

Long-term non-progressors (LTNP) represent a minority (1–5%) of HIV-infected individuals characterized by documented infection for more than 7–10 years, a stable CD4+ T cell count over 500/mm3 and low viremia in the absence of antiretroviral treatment. Protective factors described so far such as the CCR5delta32 deletion, protective HLA alleles, or defective viruses fail to fully explain the partial protection phenotype. The existence of additional host resistance mechanisms in LTNP patients was investigated here using a whole human genome microarray study comparing gene expression profiles of unstimulated peripheral blood mononuclear cells from LTNP patients, HIV-1 infected patients under antiretroviral therapy with CD4+ T cell levels above 500/mm3 (ST), as well as healthy individuals. Genes that were up- or downregulated exclusively in LTNP, ST or in both groups in comparison to controls were identified and classified in functional categories using Ingenuity Pathway Analysis. ST and LTNP patient groups revealed distinct genetic profiles, regarding gene number in each category and up- or downregulation of specific genes, which could have a bearing on the outcome of each group. We selected some relevant genes to validate the differential expression using quantitative real-time qRT-PCR. Among others, we found several genes related to the canonical Wnt/beta-catenin signaling pathway. Our results identify new possible host genes and molecules that could be involved in the mechanisms leading to the slower progression to AIDS and sustained CD4+ T cell counts that is peculiar to LTNP patients.

Target Gene Knockdown by 2′,4′-BNA/LNA Antisense Oligonucleotides in Zebrafish

Gene knockdowns using oligonucleotide-based approaches are useful for studying gene function in both in vitro cell culture systems and in vivo animal models. We evaluated the efficacy of 2',4'-bridged nucleic acids (BNA)-modified antisense oligonucleotides (AONs) for gene knockdown in zebrafish. We used the tcf7l1a gene as a model for testing the knockdown efficacy of 2',4'-BNA AONs and examined how the target sites/affinity and RNase H induction activity of 2',4'-BNA AONs affect knockdown efficacy. We found that tcf7l1a gene function was knocked down by 2',4'-BNA AONs that target the start codon and induce RNase H activity. Although nonspecific p53-mediated developmental defects were observed at higher doses, the effective dose of the 2',4'-BNA AONs for tcf7l1a is much lower than that of morpholino oligonucleotides. Our data thus show a potential application for 2',4'-BNA AONs in the downregulation of specific genes in zebrafish.

Bioconcentration and metabolism of BDE-209 in the presence of titanium dioxide nanoparticles and impact on the thyroid endocrine system and neuronal development in zebrafish larvae

Interactions between organic toxicants and nanoparticles (NPs) in the aquatic environment may modify toxicant bioavailability and consequently the toxicant’s environmental fate and toxicity. Therefore, we investigated the influence of titanium dioxide NPs (nano-TiO2) on deca-BDE (BDE-209; a polybrominated diphenyl ether congener) bioconcentration, metabolism and its effects on the thyroid endocrine system in zebrafish (Danio rerio) larvae. Zebrafish embryos were exposed to various concentrations of BDE-209 alone or in combination with nano-TiO2 (0.1 mg/L) until 7-day post-fertilization. Nano-TiO2 can adsorb BDE-209 and nano-TiO2 is taken up into developing zebrafish larvae. Chemical measurements showed that BDE-209 was bioconcentrated and metabolized in zebrafish larvae, and BDE-209 uptake was enhanced by nano-TiO2. Furthermore, increased BDE-209 metabolites were detected in larvae co-exposed with nano-TiO2. BDE-209 exposure significantly increased whole-body thyroid hormone contents (T3 and T4); T4 content significantly increased in the larvae co-exposed with nano-TiO2. Nano-TiO2 exposure alone did not induce generation of reactive oxygen species, lipid peroxidative oxidation, gene transcription or thyroid hormone levels. Upregulation of several gene transcriptions (tshβ, tg, dio2) in the hypothalamic–pituitary–thyroid axis was also observed. Furthermore, co-exposure of nano-TiO2 and BDE-209 caused a decrease in locomotion activity and downregulation of specific genes and proteins involved in the central nervous system of developing zebrafish larvae (e.g. myelin basic protein and α1-tubulin). These results indicate nano-TiO2 enhances BDE-209 bioavailability and metabolism, leading to thyroid endocrine disruption and developmental neurotoxicity in zebrafish.

Phospholipase Cβ Binds to C3PO and its Components that Orchestrates RNA Interference

Phospholipase Cβ (PLCβ1-4) is a family of signaling enzymes coupled to G protein Gαq. When activated by Gαq, PLCβ hydrolyses phosphatidylinositol 4,5 bisphosphate to produce two second messengers that lead to activation of protein kinase C and an increase in intracellular calcium. While the majority of PLCβ resides on the plasma membrane, a small but significant population is present in the cytosol and nucleus. Recently, our lab identified translin-associated factor X (TRAX) to be a cellular binding partner of PLCβ with a binding constant only 10 fold lower than Gαq. TRAX is known to bind to translin forming the complex C3PO which assists in RNA interference, along with other potential nuclease functions. Recently, our lab showed that PLCβ can reverse the down regulation of specific genes (GAPDH) presumably through its interaction with TRAX thus linking PLCβ with gene regulation. Here, we have characterized the complexes formed between PLCβ and TRAX, translin and C3PO. Using FRET, we show that PLCβ binds to C3PO with Kd similar to PLCβ and Trax (8nM) whereas the Kd for binding to translin is much weaker (40 nM). We also found that translin, Trax and C3PO occur in multiple oligomeric states as shown by FCS and in native gels. Anisotropy studies using labeled oligonucleotides and translin and C3PO show that PLCβ inhibits a nucleotide binding mode of the proteins. Studies to characterize the change in the composition and stoichiometry of C3PO and translin by PLCβ in the presence and absence of various oligonucleotides using FCS and Photon Counting Histogram (PCH) analysis are currently underway. This work was supported by NIH GM053132.

HrpA, a DEAH-Box RNA Helicase, Is Involved in Global Gene Regulation in the Lyme Disease Spirochete

Spirochetes causing Lyme borreliosis are obligate parasites that can only be found in a tick vector or a vertebrate host. The ability to survive in these two disparate environments requires up and downregulation of specific genes by regulatory circuits that remain largely obscure. In this work on the Lyme spirochete, B. burgdorferi, we show that a disruption of the hrpA gene, which encodes a putative RNA helicase, results in a complete loss in the ability of the spirochetes to infect mice by needle inoculation. Studies of protein expression in culture by 2D gels revealed a change in the expression of 33 proteins in hrpA clones relative to the wild-type parent. Quantitative characterization of protein expression by iTRAQ analysis revealed a total of 187 differentially regulated proteins in an hrpA background: 90 downregulated and 97 upregulated. Forty-two of the 90 downregulated and 65 of the 97 upregulated proteins are not regulated under any conditions by the previously reported regulators in B. burgdorferi (bosR, rrp2, rpoN, rpoS or rrp1). Downregulated and upregulated proteins also fell into distinct functional categories. We conclude that HrpA is part of a new and distinct global regulatory pathway in B. burgdorferi gene expression. Because an HrpA orthologue is present in many bacteria, its participation in global regulation in B. burgdorferi may have relevance in other bacterial species where its function remains obscure. We believe this to be the first report of a role for an RNA helicase in a global regulatory pathway in bacteria. This finding is particularly timely with the recent growth of the field of RNA regulation of gene expression and the ability of RNA helicases to modulate RNA structure and function.

Genome‐wide DNA methylation analysis of neuroblastic tumors reveals clinically relevant epigenetic events and large‐scale epigenomic alterations localized to telomeric regions

The downregulation of specific genes through DNA hypermethylation is a major hallmark of cancer, although the extent and genomic distribution of hypermethylation occurring within cancer genomes is poorly understood. We report on the first genome-wide analysis of DNA methylation alterations in different neuroblastic tumor subtypes and cell lines, revealing higher order organization and clinically relevant alterations of the epigenome. The methylation status of 33,485 discrete loci representing all annotated CpG islands and RefSeq gene promoters was assessed in primary neuroblastic tumors and cell lines. A comparison of genes that were hypermethylated exclusively in the clinically favorable ganglioneuroma/ganglioneuroblastoma tumors revealed that nine genes were associated with poor clinical outcome when overexpressed in the unfavorable neuroblastoma (NB) tumors. Moreover, an integrated DNA methylation and copy number analysis identified 80 genes that were recurrently concomitantly deleted and hypermethylated in NB, with 37 reactivated by 5-aza-deoxycytidine. Lower expression of four of these genes was correlated with poor clinical outcome, further implicating their inactivation in aggressive disease pathogenesis. Analysis of genome-wide hypermethylation patterns revealed 70 recurrent large-scale blocks of contiguously hypermethylated promoters/CpG islands, up to 590 kb in length, with a distribution bias toward telomeric regions. Genome-wide hypermethylation events in neuroblastic tumors are extensive and frequently occur in large-scale blocks with a significant bias toward telomeric regions, indicating that some methylation alterations have occurred in a coordinated manner. Our results indicate that methylation contributes toward the clinicopathological features of neuroblastic tumors, revealing numerous genes associated with poor patient survival in NB.

Gene expression in the placenta: maternal stress and epigenetic responses.

Successful placental development is crucial for optimal growth, development, maturation and survival of the embryo/fetus into adulthood. Numerous epidemiologic and experimental studies have demonstrated the profound influence of intrauterine environment on life, and the diseases to which one is subject as an adult. For the most part, these invidious influences, whether maternal hypoxia, protein or caloric deficiency or excess, and others, represent types of maternal stress. In the present review, we examine certain aspects of gene expression in the placenta as a consequence of maternal stressors. To examine these issues in a controlled manner, and in a species in which the genome has been sequenced, most of these reported studies have been performed in the mouse. Although each individual maternal stress is characterized by up- or down-regulation of specific genes in the placenta, functional analysis reveals some patterns of gene expression common to the several forms of stress. Of critical importance, these genes include those involved in DNA methylation and histone modification, cell cycle regulation, and related global pathways of great relevance to epigenesis and the developmental origins of adult health and disease.

Quantifiable downregulation of endogenous genes in Agaricus bisporus mediated by expression of RNA hairpins.

Functional gene studies in the cultivated white button mushroom Agaricus bisporus have been constrained by the absence of effective gene-silencing tools. Using two endogenous genes from A. bisporus, we have tested the utility of dsRNA hairpin constructs to mediate downregulation of specific genes. Hairpin constructs for genes encoding orotidine 5'- monophosphate decarboxylase (URA3) and carboxin resistance (CBX) were introduced into A. bisporus using Agrobacteriummediated transfection. Although predicted changes in phenotype were not observed in vitro, quantitative-PCR analyses indicated unambiguously that transcripts in several transformants were substantially reduced compared with the non-transformed controls. Interestingly, some hairpin transformants exhibited increased transcription of target genes. Our observations show that hairpin transgenic sequences can mediate downregulation of A. bisporus endogenous genes and that the technology has the potential to expedite functional genomics of the mushroom.

Histone Deacetylase 11 (HDAC11) as a Novel Molecular Target in Mantle Cell Lymphoma

Mantle Cell Lymphoma (MCL) is a B-cell Non-Hodgkin's Lymphoma characterized by the translocation t(11;14)(q13;32) and a consequent Cyclin D1 over-expression. The clinical evolution of MCL is very aggressive, and currently there is no standard of care for this malignancy.In spite of the known association of Histone Deacetylases (HDACs) and solid malignancy growth, little is known regarding the role of these enzymes in MCL. We recently found newest member of the HDAC family, HDAC11, a molecule with an unknown prior physiological role, is a strong transcriptional repressor of gene expression1. Interestingly, we have found a significant over-expression of HDAC11 in murine as well as in human MCL cell lines. In primary cells from patients with MCL, we observed that higher levels of HDAC11 are associated with a more aggressive disease, while low expressions of HDAC11 are seen in patients with the indolent variant of MCL. Microarray analysis of cells in which HDAC11 expression was silenced revealed significant down regulation of specific genes involved in the control of apoptosis, cell cycle, methylation and B-cell survival. Critical genes regulated by HDAC11 include IL-10 and IL-6, cytokines known to be survival factors for malignant B-cells. In addition, we have unveiled a previously unknown association between HDAC11, cyclin D1, and Cyclin Dependent Kinase Inhibitors (CDKI), molecules that play a central role in the pathogenesis of MCL.

Metastasis‐associated gene expression profile of liver and subcutaneous lesions derived from mouse pheochromocytoma cells

The development of metastatic cancer is associated with overexpression or downregulation of specific genes and cell regulatory pathways. Some of these genes and pathways may be involved in invasion and dissemination of tumor cells, while others may promote seeding, survival or growth of cells at specific distant sites. In this investigation, gene expression profiles of nonmetastasizing tumors generated by injecting mouse pheochromocytoma cells (MPCs) subcutaneously were compared to those of liver tumors generated by injecting the cells intravenously. Both were compared to the cultured parental cell line. Tumors in the liver have a route of spread, anatomical distribution, and growth environment similar to naturally metastasizing pheochromocytomas, while intravenous injection of cells bypasses the initial steps of metastasis occurring spontaneously from a primary tumor. Eight genes were upregulated in liver tumors, 15 in subcutaneous tumors and seven in both compared to the cultured cells. Using quantitative real-time PCR, expression of five genes (Metap2, Reck, S100a4, Timp2, and Timp3) was verified as significantly lower in liver tumors than in subcutaneous tumors. Downregulation of these genes has been previously been associated with malignancy of pheochromocytomas. These findings indicate that different microenvironments can differentially affect the expression of metastasis-related genes in pheochromocytomas, and that overexpression or underexpression of these genes need not be present when the tumor cells are initially disseminated. The hepatic localization of tumors formed by intravenously injected MPC cells and the tumors' gene expression profile resembling that of naturally occurring pheochromocytoma metastases support the use of this model to study pheochromocytoma metastasis.

Constitutional Downregulation of SEMA5A Expression in Autism

There is strong evidence for the importance of genetic factors in idiopathic autism. The results from independent twin and family studies suggest that the disorder is caused by the action of several genes, possibly acting epistatically. We have used cDNA microarray technology for the identification of constitutional changes in the gene expression profile associated with idiopathic autism. Samples were obtained and analyzed from 6 affected subjects belonging to multiplex autism families and from 6 healthy controls. We assessed the expression levels for approximately 7,700 genes by cDNA microarrays using mRNA derived from Epstein-Barr virus-transformed B lymphocytes. The microarray data were analyzed in order to identify up- or downregulation of specific genes. A common pattern with nine downregulated genes was identified among samples derived from individuals with autism when compared to controls. Four of these nine genes encode proteins involved in biological processes associated with brain function or the immune system, and are consequently considered as candidates for genes associated with autism. Quantitative real-time PCR confirms the downregulation of the gene encoding SEMA5A, a protein involved in axonal guidance. Epstein-Barr virus should be considered as a possible source for altered expression, but our consistent results make us suggest SEMA5A as a candidate gene in the etiology of idiopathic autism.