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Elena Feinstein

Publications and source records attributed to Elena Feinstein.

13 recordsLinked to original sources
PubMed2005

Studies on the transcriptional regulation of cholesterol 24-hydroxylase (CYP46A1): marked insensitivity toward different regulatory axes.

Mammalian CNS contains a disproportionally large and remarkably stable pool of cholesterol. Despite an efficient recycling there is some requirement for elimination of brain cholesterol. Conversion of cholesterol into 24S-hydroxycholesterol by the cholesterol 24-hydroxylase (CYP46A1) is the quantitatively most important mechanism. Based on the protein expression and plasma levels of 24S-hydroxycholesterol, CYP46A1 activity appears to be highly stable in adults. Here we have made a structural and functional characterization of the promoter of the human CYP46A1 gene. No canonical TATA or CAAT boxes were found in the promoter region. Moreover this region had a high GC content, a feature often found in genes considered to have a largely housekeeping function. A broad spectrum of regulatory axes using a variety of promoter constructs did not result in a significant transcriptional regulation. Oxidative stress caused a significant increase in transcriptional activity. The possibility of a substrate-dependent transcriptional regulation was explored in vivo in a sterol-deficient mouse model (Dhcr24 null) in which almost all cholesterol had been replaced with desmosterol, which is not a substrate for CYP46A1. Compared with heterozygous littermates there was no statistically significant difference in the mRNA levels of Cyp46a1. During the first 2 weeks of life in the wild-type mouse, however, a significant increase of Cyp46a1 mRNA levels was found, in parallel with an increase in 24S-hydroxycholesterol level and a reduction of cholesterol synthesis. The failure to demonstrate a significant transcriptional regulation under most conditions is discussed in relation to the turnover of brain and neuronal cholesterol.

Alzheimer Disease↗
PubMed2005

p53 is a suppressor of inflammatory response in mice.

Chronic inflammation is known to promote cancer, suggesting that negative regulation of inflammation is likely to be tumor suppressive. We found that p53 is a general inhibitor of inflammation that acts as an antagonist of nuclear factor kappaB (NFkappaB). We first observed striking similarities in global gene expression profiles in human prostate cancer cells LNCaP transduced with p53 inhibitory genetic element or treated with TNF, suggesting that p53 inhibits transcription of TNF-inducible genes that are largely regulated by NFkappaB. Consistently, ectopically expressed p53 acts as an inhibitor of transcription of NFkappaB-dependent promoters. Furthermore, suppression of inflammatory response by p53 was observed in vivo in mice by comparing wild-type and p53 null animals at molecular (inhibition of transcription of genes encoding cytokines and chemokines, reducing accumulation of reactive oxygen species and protein oxidation products), cellular (activation of macrophages and neutrophil clearance) and organismal (high levels of metabolic markers of inflammation in tissues of p53-deficient mice and their hypersensitivity to LPS) levels. These observations indicate that p53, acting through suppression of NFkappaB, plays the role of a general "buffer" of innate immune response in vivo that is well consistent with its tumor suppressor function and frequent constitutive activation of NFkappaB in tumors.

Animals↗
PubMed2005

Ero1-L alpha plays a key role in a HIF-1-mediated pathway to improve disulfide bond formation and VEGF secretion under hypoxia: implication for cancer.

Oxygen is the ultimate source of oxidizing power for disulfide bond formation, suggesting that under limiting oxygen proper protein folding might be compromised. We show that secretion of vascular endothelial growth factor (VEGF), a protein with multiple disulfide bonds, was indeed impeded under hypoxia and was partially restored by artificial increase of oxidizing equivalents with diamide. Physiologically, the oxireductase endoplasmic reticulum oxidoreductin-1 (Ero1)-L alpha, but not other proteins in the relay of disulfide formation, was strongly upregulated by hypoxia and independently by hypoglycemia, two known accompaniments of tumors. Further, we provide genetic evidence that induction of Ero1-L alpha by hypoxia and hypoglycemia is mediated by the transcription factor hypoxia-inducible factor 1 (HIF-1) but is independent of p53. In natural human tumors, Ero1-L alpha mRNA was specifically induced in hypoxic microenvironments coinciding with that of upregulated VEGF expression. To establish a physiological relevance to modulations in Ero1-L alpha levels, we showed that even a modest, two- to three-fold reduction in Ero1-L alpha production via siRNA leads to significant inhibition of VEGF secretion, a compromised proliferation capacity and enhanced apoptosis. Together, these findings demonstrate that hypoxic induction of Ero1-L alpha is the key adaptive response in a previously unrecognized HIF-1-mediated pathway that operates to improve protein secretion under hypoxia and might be harnessed for inhibiting tumor growth via inhibiting VEGF-driven angiogenesis.

Animals↗
PubMed2004

Regeneration of peroxiredoxins by p53-regulated sestrins, homologs of bacterial AhpD.

Acting as a signal, hydrogen peroxide circumvents antioxidant defense by overoxidizing peroxiredoxins (Prxs), the enzymes that metabolize peroxides. We show that sestrins, a family of proteins whose expression is modulated by p53, are required for regeneration of Prxs containing Cys-SO(2)H, thus reestablishing the antioxidant firewall. Sestrins contain a predicted redox-active domain homologous to AhpD, the enzyme catalyzing the reduction of a bacterial Prx, AhpC. Purified Hi95 (sestrin 2) protein supports adenosine triphosphate-dependent reduction of overoxidized PrxI in vitro, indicating that unlike AhpD, which is a disulfide reductase, sestrins are cysteine sulfinyl reductases. As modulators of peroxide signaling and antioxidant defense, sestrins constitute potential therapeutic targets.

Amino Acid Sequence↗
PubMed2004

Identification and handling of artifactual gene expression profiles emerging in microarray hybridization experiments.

Mathematical methods of analysis of microarray hybridizations deal with gene expression profiles as elementary units. However, some of these profiles do not reflect a biologically relevant transcriptional response, but rather stem from technical artifacts. Here, we describe two technically independent but rationally interconnected methods for identification of such artifactual profiles. Our diagnostics are based on detection of deviations from uniformity, which is assumed as the main underlying principle of microarray design. Method 1 is based on detection of non-uniformity of microarray distribution of printed genes that are clustered based on the similarity of their expression profiles. Method 2 is based on evaluation of the presence of gene-specific microarray spots within the slides' areas characterized by an abnormal concentration of low/high differential expression values, which we define as 'patterns of differentials'. Applying two novel algorithms, for nested clustering (method 1) and for pattern detection (method 2), we can make a dual estimation of the profile's quality for almost every printed gene. Genes with artifactual profiles detected by method 1 may then be removed from further analysis. Suspicious differential expression values detected by method 2 may be either removed or weighted according to the probabilities of patterns that cover them, thus diminishing their input in any further data analysis.

Algorithms↗
PubMed2004

CMF608-a novel mechanical strain-induced bone-specific protein expressed in early osteochondroprogenitor cells.

Microarray gene expression analysis was utilized to identify genes upregulated in primary rat calvaria cultures in response to mechanical force. One of the identified genes designated CMF608 appeared to be novel. The corresponding full-length cDNA was cloned and characterized in more details. It encodes a putative 2597 amino acid protein containing N-terminal signal peptide, six leucine-rich repeats (LRRs), and 12 immunoglobulin-like repeats, 10 of which are clustered within the C-terminus. Expression of CMF608 is bone-specific and the main type of CMF608-positive cells is mesenchymal osteochondroprogenitors with fibroblast-like morphology. These cells reside in the perichondral fibrous ring of La Croix, periosteum, endosteum of normal bone as well as in the activated periosteum and early fibrous callus generated postfracture. Expression of CMF608 is notably absent from the regions of endochondral ossification. Mature bone cell types do not produce CMF608 with the exception of chondrocytes of the tangential layer of the articular cartilage, which are thought to be under constant mechanical loading. Ectopic expression of CMF608 in HEK293T cells shows that the protein is subjected to post-translational processing and its N-terminal approximately 90 kDa polypeptide can be found in the conditioned medium. Ectopic expression of either the full-length cDNA of CMF608 or of its N-terminal region in CMF608-negative ROS17/2.8 rat osteosarcoma cells results in transfected clones displaying increased proliferation rate and the characteristics of less-differentiated osteoblasts compared to the control cells. Our data indicate that CMF608 is a unique marker of early osteochondroprogenitor cells. We propose that it could be functionally involved in maintenance of the osteochondroprogenitor cells pool and its down-regulation precedes terminal differentiation.

Amino Acid Sequence↗
PubMed2004

Inhibition of oxygen-induced retinopathy in RTP801-deficient mice.

PURPOSE: Ischemic proliferative retinopathy, which occurs as a complication of diabetes mellitus, prematurity, or retinal vein occlusion, is a major cause of blindness worldwide. In addition to retinal neovascularization, it involves retinal degeneration, of which apoptosis is the main cause. A prior report has described the cloning of a novel HIF-1-responsive gene, RTP801, which displays strong hypoxia-dependent upregulation in ischemic cells of neuronal origin, both in vitro and in vivo. Moreover, inducible overexpression of RTP801 promotes the apoptotic death of differentiated neuron-like PC12 cells and increases their sensitivity to ischemic injury and oxidative stress. The purpose of the study was to examine the potential role of RTP801 in the pathogenesis of retinopathy, using RTP801-deficient mice. METHODS: Wild-type and RTP801-knockout mice were used in a model of retinopathy of prematurity (ROP). Their retinas were collected at postnatal day (P)14 and P17. They were examined by fluorescein angiography and by analysis of VEGF expression, neovascularization, and apoptosis. RESULTS: The expression of RTP801 was induced in the wild-type retina after hypoxia treatment. The retinal expression of VEGF after transfer to normoxic conditions was similarly upregulated in both wild-type and knockout mice. Nevertheless, the retinas of the RTP801-knockout mice in an ROP model showed a significant reduction in retinal neovascularization (P < 0.0001) and in the number of apoptotic cells in the inner nuclear layer (P < 0.0001). CONCLUSIONS: In the absence of RTP801 expression, development of retinopathy in the mouse model of ROP was significantly attenuated, thus implying an important role of RTP801 in the pathogenesis of ROP.

Adaptor Proteins, Signal Transducing↗
PubMed2003

Molecular analysis of transitional cell carcinoma using cDNA microarray.

The incidence of transitional cell carcinoma (TCC), the fourth most common neoplasm diagnosed in men, is rising. Despite the development of several noninvasive diagnostic tests, none have gained full recognition by the clinicians. Gene expression profiling of tumors can identify new molecular markers for early diagnosis and disease follow-up. It also allows the classification of tumors into subclasses assisting in disease diagnosis and prognosis, as well as in treatment selection. In this paper, we employed expression profiling for molecular analysis of TCC. A TCC-derived cDNA microarray was constructed and hybridized with 19 probes from normal urothelium and TCC tissues. Hierarchical clustering analysis identified all normal urothelium samples to be tightly clustered and separated from the TCC samples, with 29 of the genes significantly induced (t-test, P<10(-5)) in noninvasive TCC compared to normal urothelium. The identified genes are involved in epithelial cells' functions, tumorigenesis or apoptosis, and could become molecular tools for noninvasive TCC diagnosis. Principal components analysis of the noninvasive and invasive TCC expression profiles further revealed sets of genes that are specifically induced in different tumor subsets, thus providing molecular fingerprints that expand the information gained from classical staging and grading.

Adult↗
PubMed2002

Identification of a novel stress-responsive gene Hi95 involved in regulation of cell viability.

cDNA microarray hybridization was used in an attempt to identify novel genes participating in cellular responses to prolonged hypoxia. One of the identified novel genes, designated Hi95 shared significant homology to a p53-regulated GADD family member PA26. In addition to its induction in response to prolonged hypoxia, the increased Hi95 transcription was observed following DNA damage or oxidative stress, but not following hyperthermia or serum starvation. Whereas induction of Hi95 by prolonged hypoxia or by oxidative stress is most likely p53-independent, its induction in response to DNA damaging treatments (gamma- or UV-irradiation, or doxorubicin) occurs in a p53-dependent manner. Overexpression of Hi95 full-length cDNA was found toxic for many types of cultured cells directly leading either to their apoptotic death or to sensitization to serum starvation and DNA damaging treatments. Unexpectedly, conditional overexpression of the Hi95 cDNA in MCF7-tet-off cells resulted in their protection against cell death induced by hypoxia/glucose deprivation or H(2)O(2). Thus, Hi95 gene seems to be involved in complex regulation of cell viability in response to different stress conditions.

Amino Acid Sequence↗
PubMed2002

Nr-CAM is a target gene of the beta-catenin/LEF-1 pathway in melanoma and colon cancer and its expression enhances motility and confers tumorigenesis.

beta-catenin and plakoglobin (gamma-catenin) are homologous molecules involved in cell adhesion, linking cadherin receptors to the cytoskeleton. beta-catenin is also a key component of the Wnt pathway by being a coactivator of LEF/TCF transcription factors. To identify novel target genes induced by beta-catenin and/or plakoglobin, DNA microarray analysis was carried out with RNA from cells overexpressing either protein. This analysis revealed that Nr-CAM is the gene most extensively induced by both catenins. Overexpression of either beta-catenin or plakoglobin induced Nr-CAM in a variety of cell types and the LEF/TCF binding sites in the Nr-CAM promoter were required for its activation by catenins. Retroviral transduction of Nr-CAM into NIH3T3 cells stimulated cell growth, enhanced motility, induced transformation, and produced rapidly growing tumors in nude mice. Nr-CAM and LEF-1 expression was elevated in human colon cancer tissue and cell lines and in human malignant melanoma cell lines but not in melanocytes or normal colon tissue. Dominant negative LEF-1 decreased Nr-CAM expression and antibodies to Nr-CAM inhibited the motility of B16 melanoma cells. The results indicate that induction of Nr-CAM transcription by beta-catenin or plakoglobin plays a role in melanoma and colon cancer tumorigenesis, probably by promoting cell growth and motility.

3T3 Cells↗
PubMed2002

Expression of prostate specific antigen (PSA) is negatively regulated by p53.

Although prostate-specific antigen (PSA) is considered a uniquely important tumor marker and is broadly used for early detection of prostate cancer, the molecular mechanisms underlying its elevated expression in tumors have been unknown. By using cDNA microarray gene expression profiling, we found a fourfold increase in the PSA mRNA level in prostatic carcinoma cell line LNCaP, in which the p53 pathway was suppressed by a dominant negative p53 mutant. Consistently, p53 suppression caused a 4-8-fold increase in secretion of PSA protein in culture medium, suggesting that PSA gene expression is under negative control of p53. While wild type p53 strongly repressed, dominant negative p53 mutants stimulated PSA promoter-driven transcription and secretion of PSA in transient transfection experiments. The inhibitory effect of wild type p53 was undetectable in the presence of trichostatin A, suggesting the involvement of histone deacetylation in negative regulation of PSA promoter activity. Thus, PSA is likely to be a tissue specific indicator of transformation-associated p53 suppression in prostate cells. This finding provides a plausible explanation for a frequent increase of PSA levels in advanced prostate cancer.

Acetylation↗
PubMed2002

Identification of a novel hypoxia-inducible factor 1-responsive gene, RTP801, involved in apoptosis.

Hypoxia is an important factor that elicits numerous physiological and pathological responses. One of the major gene expression programs triggered by hypoxia is mediated through hypoxia-responsive transcription factor hypoxia-inducible factor 1 (HIF-1). Here, we report the identification and cloning of a novel HIF-1-responsive gene, designated RTP801. Its strong up-regulation by hypoxia was detected both in vitro and in vivo in an animal model of ischemic stroke. When induced from a tetracycline-repressible promoter, RTP801 protected MCF7 and PC12 cells from hypoxia in glucose-free medium and from H(2)O(2)-triggered apoptosis via a dramatic reduction in the generation of reactive oxygen species. However, expression of RTP801 appeared toxic for nondividing neuron-like PC12 cells and increased their sensitivity to ischemic injury and oxidative stress. Liposomal delivery of RTP801 cDNA to mouse lungs also resulted in massive cell death. Thus, the biological effect of RTP801 overexpression depends on the cell context and may be either protecting or detrimental for cells under conditions of oxidative or ischemic stresses. Altogether, the data suggest a complex type of involvement of RTP801 in the pathogenesis of ischemic diseases.

Adaptor Proteins, Signal Transducing↗