PubMed Central (PMC3 - NLM DTD)
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Archive of life sciences journal literature at the U.S. National Institutes of Health (NIH), developed and managed by NIH's National Center for Biotechnology Information (NCBI) in the National Library of Medicine (NLM).
Mostrando recursos 101 - 120 de 59,293
101.
Fission yeast Arp6 is required for telomere silencing, but functions independently of Swi6 - Ueno, Masaru; Murase, Tadashi; Kibe, Tatsuya; Ohashi, Noriyuki; Tomita, Kazunori; Murakami, Yota; Uritani, Masahiro; Ushimaru, Takashi; Harata, Masahiko
The actin-related proteins (Arps), which are subdivided into at least eight subfamilies, are conserved from yeast to humans. A member of the Arp6 subfamily in Drosophila, Arp4/Arp6, co-localizes with heterochromatin protein 1 (HP1) in pericentric heterochromatin. Fission yeast Schizosaccharomyces pombe possesses both an HP1 homolog and an Arp6 homolog. However, the function of S.pombe Arp6 has not been characterized yet. We found that deletion of arp6+ impaired telomere silencing, but did not affect centromere silencing. Chromatin immunoprecipitation assays revealed that Arp6 bound to the telomere region. However, unlike Drosophila Arp4/Arp6, S.pombe Arp6 was distributed throughout nuclei. The binding of Arp6...
102.
Transcription regulation of TNF-?-early response genes by poly(ADP-ribose) polymerase-1 in murine heart endothelial cells - Carrillo, Ana; Monreal, Yolanda; Ramírez, Pablo; Marin, Luis; Parrilla, Pascual; Oliver, Francisco Javier; Yélamos, José
Poly(ADP-ribose) polymerase-1 (PARP-1) has been involved in endothelial cell dysfunction associated with various pathophysiological conditions. The intrinsic mechanism of PARP-1-mediated endothelial cell dysfunction could be related to PARP-1 overactivation, NAD+ consumption and ATP depletion. An alternative way could involve transcription regulation. By using high-density microarrays, we examined early tumor necrosis factor ? (TNF-?)-stimulated gene expression profiles in PARP-1+/+ and PARP-1/ murine heart endothelial cells. TNF-? modulated a significant number of genes in both cell types. We have identified a set of genes whose expression in response to TNF-? is modulated by PARP-1, whereas the expression of others is PARP-1-independent. Up-regulation...
103.
A new ?-interferon-inducible promoter and splice variants of an anti-angiogenic human tRNA synthetase - Liu, Jianming; Shue, Eveline; Ewalt, Karla L.; Schimmel, Paul
Two forms of human tryptophanyl-tRNA synthetase (TrpRS) are produced in vivo through alternative mRNA splicing. The two forms, full-length TrpRS and mini TrpRS, are catalytically active, but are distinguished by the striking anti-proliferative and anti-angiogenic activity specific to mini TrpRS. Here we describe two new splice variants of human TrpRS mRNA. Their production was strongly regulated by ?-interferon (IFN-?), an anti-proliferative cytokine known to stimulate the expression of other anti-angiogenic factors. A new IFN-?-sensitive promoter was demonstrated to drive production of these splice variants. In human endothelial cells, both the newly discovered and a previously reported promoter were shown to...
104.
PCR amplification of DNA containing non-standard base pairs by variants of reverse transcriptase from Human Immunodeficiency Virus-1 - Sismour, A. Michael; Lutz, Stefan; Park, Jeong-Ho; Lutz, Michael J.; Boyer, Paul L.; Hughes, Stephen H.; Benner, Steven A.
As the next step towards generating a synthetic biology from artificial genetic information systems, we have examined variants of HIV reverse transcriptase (RT) for their ability to synthesize duplex DNA incorporating the non-standard base pair between 2,4-diaminopyrimidine (pyDAD), a pyrimidine presenting a hydrogen bond donoracceptordonor pattern to the complementary base, and xanthine (puADA), a purine presenting a hydrogen bond acceptordonoracceptor pattern. This base pair fits the WatsonCrick geometry, but is joined by a pattern of hydrogen bond donor and acceptor groups different from those joining the GC and AT pairs. A variant of HIV-RT where Tyr 188 is replaced by...
105.
Involvement of Hus1 in the chain elongation step of DNA replication after exposure to camptothecin or ionizing radiation - Wang, Xiang; Guan, Jun; Hu, Baocheng; Weiss, Robert S.; Iliakis, George; Wang, Ya
DNA damage-induced S phase (S) checkpoint includes inhibition of both replicon initiation and chain elongation. The precise mechanism for controlling the two processes remains unclear. In this study, we showed that Hus1-deficient mouse cells had an impaired S checkpoint after exposure to DNA strand break-inducing agents such as camptothecin (CPT) (?1.0 µM), or ionizing radiation (IR) (?15 Gy). The Hus1-dependent S checkpoint contributes to cell resistance to CPT. This impaired S checkpoint induced by CPT or IR in Hus1-deficient cells reflected mainly the chain elongation step of DNA replication and was correlated with the reduction of dissociation of PCNA from...
106.
Structure of a palindromic amplicon junction implicates microhomology-mediated end joining as a mechanism of sister chromatid fusion during gene amplification - Okuno, Yukiko; Hahn, Peter J.; Gilbert, David M.
Amplification of the copy number of oncogenes is frequently associated with tumor progression. Often, the amplified DNA consists of large (tens to hundreds of kilobases) head-to-head inverted repeat palindromes (amplicons). Several mechanisms have been proposed to explain palindrome formation but their relative contributions in nature have been difficult to assess without precise knowledge of the sequences involved at the junction of natural amplicons. Here, we have sequenced one such junction and compared this sequence to the un-rearranged structure, allowing us to pinpoint the site of sister chromatid fusion. Our results support a novel model, consistent with all described sister chromatid...
108.
VITO-1 is an essential cofactor of TEF1-dependent muscle-specific gene regulation - Günther, Stefan; Mielcarek, Michal; Krüger, Marcus; Braun, Thomas
The expression of several muscle-specific genes is partially or completely regulated by MCAT elements, which bind members of the TEF family of transcription factors. TEF1 itself is unable to activate reporter plasmids bearing TEF1-binding sites, suggesting that additional bridging or co-activating factors are necessary to allow interaction of TEF1 with the transcriptional machinery. In addition, none of the known TEF genes are exclusively expressed in the cardiac or skeletal muscle lineage to account for the muscle-specific expression of MCAT-dependent genes. Here we describe that VITO-1, a new SID (scalloped interaction domain)-containing protein, binds to TEF1 in vitro and strongly stimulates...
109.
Sequence-specific transcriptional repression by an MBD2-interacting zinc finger protein MIZF - Sekimata, Masayuki; Homma, Yoshimi
MBD2 is a member of the methyl-CpG-binding protein family that plays an important role in methylated DNA silencing. We have recently identified a novel zinc finger protein, MIZF, as an MBD2-binding partner. To understand the physiological function of MIZF in MBD2-mediated gene silencing, we investigated the DNA-binding properties of MIZF and its potential target genes. Using a cyclic amplification and selection of targets technique, the consensus sequence CGGACGTT, which contains a conserved CGGAC core, was determined as sufficient for MIZF binding. Deletion of individual zinc fingers revealed that five of the seven zinc fingers are required for DNA binding. Reporter...
110.
Fission yeast global repressors regulate the specificity of chromatin alteration in response to distinct environmental stresses - Hirota, Kouji; Hasemi, Tomoko; Yamada, Takatomi; Mizuno, Ken-ich; Hoffman, Charles S.; Shibata, Takehiko; Ohta, Kunihiro
The specific induction of genes in response to distinct environmental stress is vital for all eukaryotes. To study the mechanisms that result in selective gene responses, we examined the role of the fission yeast Tup1 family repressors in chromatin regulation. We found that chromatin structure around a cAMP-responsive element (CRE)-like sequence in ade6-M26 that is bound by Atf1·Pcr1 transcriptional activation was altered in response to osmotic stress but not to heat and oxidative stresses. Such chromatin structure alteration occurred later than the Atf1 phosphorylation but correlated well with stress-induced transcriptional activation at ade6-M26. This chromatin structure alteration required components for...
111.
A programmed 1 ribosomal frameshift signal can function as a cis-acting mRNA destabilizing element - Plant, Ewan P.; Wang, Pinger; Jacobs, Jonathan L.; Dinman, Jonathan D.
Nonsense-mediated mRNA decay (NMD) directs rapid degradation of premature termination codon (PTC)-containing mRNAs, e.g. those containing frameshift mutations. Many viral mRNAs encode polycistronic messages where programmed 1 ribosomal frameshift (1 PRF) signals direct ribosomes to synthesize polyproteins. A previous study, which identified consensus 1 PRF signals in the yeast genome, found that, in contrast to viruses, the majority of predicted 1 PRF events would direct translating ribosomes to PTCs. Here we tested the hypothesis that a 1 PRF signal can function as a cis-acting mRNA destabilizing element by inserting an L-A viral 1 PRF signal into a PGK1 reporter construct...
112.
High-throughput protein analysis integrating bioinformatics and experimental assays - del Val, Coral; Mehrle, Alexander; Falkenhahn, Mechthild; Seiler, Markus; Glatting, Karl-Heinz; Poustka, Annemarie; Suhai, Sandor; Wiemann, Stefan
The wealth of transcript information that has been made publicly available in recent years requires the development of high-throughput functional genomics and proteomics approaches for its analysis. Such approaches need suitable data integration procedures and a high level of automation in order to gain maximum benefit from the results generated. We have designed an automatic pipeline to analyse annotated open reading frames (ORFs) stemming from full-length cDNAs produced mainly by the German cDNA Consortium. The ORFs are cloned into expression vectors for use in large-scale assays such as the determination of subcellular protein localization or kinase reaction specificity. Additionally, all...
114.
Impedance-based detection of DNA sequences using a silicon transducer with PNA as the probe layer - Macanovic, A.; Marquette, C.; Polychronakos, C.; Lawrence, M. F.
Electrochemical impedance measurements were used for the detection of single-strand DNA sequences using a peptide nucleic acid (PNA) probe layer immobilized onto Si/SiO2 chips. An epoxysilane layer is first immobilized onto the Si/SiO2 surface. The immobilization procedure consists of an epoxide/amine coupling reaction between the amino group of the PNA linker and the epoxide group of the silane. A 20-nucleotide sequence of PNA was used. Impedance measurements allow for the detection of the changes in charge distribution at the oxide/solution interface following modifications to the oxide surface. Due to these modifications, there are significant shifts in the semiconductors flat-band potential...
115.
A novel approach for evaluating the efficiency of siRNAs on protein levels in cultured cells - Wu, Weilin; Hodges, Emily; Redelius, Jenny; Höög, Christer
An important aim in the post-sequencing age of functional genomics is to translate gene sequences into protein functions. This shift of focus is particularly necessary for a very large number of human genes, referred to as novel genes, where we have no or very rudimentary information about their biochemical functions. Recently, a new method for investigating human gene functions using small interfering RNA molecules (siRNAs) has become available. siRNAs are powerful reagents for post-transcriptional silencing, where mRNA targeted by the siRNAs is degraded in vivo and the level of the encoded protein is reduced. However, the lack of antibodies against...
116.
Rapid detection of single nucleotide polymorphisms associated with spinal muscular atrophy by use of a reusable fibre-optic biosensor - Watterson, James H.; Raha, Sandeep; Kotoris, Christopher C.; Wust, Christopher C.; Gharabaghi, Farhad; Jantzi, Sarah C.; Haynes, Nicole K.; Gendron, Nathalie H.; Krull, Ulrich J.; Mackenzie, Alex E.; Piunno, Paul A. E.
Rapid (<2 min) and quantitative genotyping for single nucleotide polymorphisms (SNPs) associated with spinal muscular atrophy was done using a reusable (approximately 80 cycles of application) fibre-optic biosensor over a clinically relevant range (04 gene copies). Sensors were functionalized with oligonucleotide probes immobilized at high density (?7 pmol/cm2) to impart enhanced selectivity for SNP discrimination and used in a total internal reflection fluorescence detection motif to detect 202 bp PCR amplicons from patient samples. Real-time detection may be done over a range of ionic strength conditions (0.11.0 M) without stringency rinsing to remove non-selectively bound materials and without loss of...
117.
Construction of long DNA molecules using long PCR-based fusion of several fragments simultaneously - Shevchuk, Nikolai A.; Bryksin, Anton V.; Nusinovich, Yevgeniya A.; Cabello, Felipe C.; Sutherland, Margaret; Ladisch, Stephan
A procedure for precise assembly of linear DNA constructs as long as 20 kb is proposed. The method, which we call long multiple fusion, has been used to assemble up to four fragments simultaneously (for a 10.8 kb final product), offering an additional improvement on the combination of long PCR and overlap extension PCR. The method is based on Pfu polymerase mix, which has a proofreading activity. We successfully assembled (and confirmed by sequencing) seven different linear constructs ranging from 3 to 20 kb, including two 20 kb products (from fragments of 11, 1.7 and 7.5 kb), two 10.8 kb...
118.
Highly sensitive amperometric detection of genomic DNA in animal tissues - Xie, Hong; Yu, Yuan Hong; Mao, Pei-Lin; Gao, Zhiqiang
A simple and highly sensitive method for the detection of genomic DNA in tissue samples is described. It is based on amperometric detection of target DNA by forming an analyte/polymeric activator bilayer on a gold electrode. The biotinylated target DNA is hybridized to oligonucleotide capture probes immobilized on the gold electrode, forming the first layer. A subsequent binding of glucose oxidase avidin conjugate to the target DNA and the introduction of a second layer of a redox polymer to the electrode, via layer-by-layer electrostatic self-assembly, allow for electrochemical detection of the catalytic oxidation current of glucose in a PBS solution....
119.
Rapid and accurate characterisation of short tandem repeats by MALDI-TOF analysis of endonuclease cleaved RNA transcripts - Seichter, Doris; Krebs, Stefan; Förster, Martin
We describe the application of matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) for the characterisation of short tandem repeat (STR) sequences by the analysis of endonuclease cleaved RNA transcripts. Several simple bovine STR loci as well as interrupted and compound microsatellites were chosen as model loci to evaluate the capabilities of MALDI-TOF MS for STR analysis. In short, the described approach consists of a PCR amplification of the investigated STR sequence, which then is transcribed into RNA and cleaved by G-specific RNase T1. Base-specific cleavage of the transcript results in high informative fragment patterns from both the repetitive core...
120.
Two-dimensional conformation-dependent electrophoresis (2D-CDE) to separate DNA fragments containing unmatched bulge from complex DNA samples - Gunnarsson, Gudmundur H.; Thormar, Hans G.; Gudmundsson, Bjarki; Akesson, Lina; Jonsson, Jon J.
DNA fragments containing mispaired and modified bases, bulges, lesions and specific sequences have altered conformation. Methods for separating complex samples of DNA fragments based on conformation but independent of length have many applications, including (i) separation of mismatched or unmatched DNA fragments from those perfectly matched; (ii) simultaneous, diagnostic, mismatch scanning of multiple fragments; (iii) isolation of damaged DNA fragments from undamaged fragments; and (iv) estimation of reannealing efficiency of complex DNA samples. We developed a two-dimensional conformation-dependent electrophoresis (2D-CDE) method for separating DNA fragments based on length and conformation in the first dimension and only on length in the...
101.
