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At least 1,693 records · Page 94Linked to original sources
PubMed1990

The virD genes from the vir region of the Ti plasmid: T-region border dependent processing steps in different rec mutants of Escherichia coli.

We evaluated the substrate requirements for virD-mediated T-circle formation in an in vivo binary test system in Escherichia coli. Two copies of the 25-bp sequence which defines the right border of the T-DNA (transferred DNA) are sufficient, and the right and the left copy of the border are equivalent in function in this system. Experiments with different rec mutants show that the occurrence and frequency of circular double-stranded and single-stranded T-DNA equivalents strongly depend on rec functions of the host. These results are discussed in the context of processing of the tumor-inducing Ti plasmid preceding the T-DNA transfer from agrobacteria to plants.

Blotting, Southern↗
PubMed2002

Characterization of the mtTFA gene and identification of a processed pseudogene in rat.

Mitochondrial DNA replication and transcription are regulated from essential nucleus-encoded components that interact with the mitochondrial (mt) D-loop region. Among these there is the mitochondrial transcription factor A (mtTFA or Tfam). We have determined the sequence of the cDNA mtTFA in rat and have demonstrated that the gene has a mosaic organization with six introns whose sizes we have calculated. A differential splicing transcript lacking exon 5 has been detected in all assayed tissues and represents 9.85% of the full length transcript. Beside the gene which is homologous to the one found in man and mouse, rat nuclear genome contains at least 12 copies of this gene or genome fragments with high similarity to mtTFA. We have determined the sequence of one of these copies. This resulted to have 76.26% similarity to the active gene but to lack introns, suggesting it might be a processed pseudogene. RT-PCR experiments have demonstrated that this pseudogene (psi mtTFA) is transcribed in liver tissue.

Alternative Splicing↗
PubMed2000

Structure of the highly conserved HERC2 gene and of multiple partially duplicated paralogs in human.

Recombination between chromosome-specific low-copy repeats (duplicons) is an underlying mechanism for several genetic disorders. Recently, a chromosome 15 duplicon was discovered in the common breakpoint regions of Prader-Willi and Angelman syndrome deletions. We identified previously the large HERC2 transcript as an ancestral gene in this duplicon, with approximately 11 HERC2-containing duplicons, and demonstrated that recessive mutations in mouse Herc2 lead to a developmental syndrome, juvenile development and fertility 2 (jdf2). We have now constructed and sequenced a genomic contig of HERC2, revealing a total of 93 exons spanning approximately 250 kb and a CpG island promoter. A processed ribosomal protein L41 pseudogene occurs in intron 2 of HERC2, and putative VNTRs occur in intron 70 (28 copies, approximately 76-bp repeat) and 3' exon 40 through intron 40 (6 copies, approximately 62-bp repeat). Sequence comparisons show that HERC2-containing duplicons have undergone several deletion, inversion, and dispersion events to form complex duplicons in 15q11, 15q13, and 16p11. To further understand the developmental role of HERC2, a highly conserved Drosophila ortholog was characterized, with 70% amino acid sequence identity to human HERC2 over the carboxy-terminal 743 residues. Combined, these studies provide significant insights into the structure of complex duplicons and into the evolutionary pathways of formation, dispersal, and genomic instability of duplicons. Our results establish that some genes not only have a protein coding function but can also play a structural role in the genome.

Amino Acid Sequence↗
PubMed1992

Recognition efficiency of the hepatitis B virus polyadenylation signals is tissue specific in transgenic mice.

The hepatitis B virus genome contains a unique polyadenylation (TATAAA) signal which is differentially utilized in the formation of the various hepatitis B virus transcripts. A head-to-tail multiple-copy insertion of a viral fragment comprising the viral enhancer, the X promoter, the X open reading frame, and the viral poly(A) signal in transgenic mice allowed us to monitor tissue-specific differences in the expression of transcripts initiating from the X promoter. These transcripts are efficiently processed at the first polyadenylation site in the liver, while in the kidney, the brain, and the testis, a portion of the transcripts covers two copies of the transgene, since only the second polyadenylation site is properly recognized. As discussed in this article, this observation suggests a tissue-specific distribution of cellular factors involved in polyadenylation.

Animals↗
PubMed2003

Concurrent transcriptional activation of ppa and ppx genes by phosphate deprivation in the cyanobacterium Synechocystis sp. strain PCC 6803.

The cyanobacterium Synechocystis sp. strain PCC 6803 possesses two genes, named ppa and ppx, which, respectively, encode proteins involved in the hydrolysis of inorganic phosphate polymers, namely, inorganic pyrophosphatase (PPA, EC 3.6.1.1), an essential enzyme that hydrolyzes pyrophosphate, and exopolyphosphatase (PPX, EC 3.6.1.11), a processive enzyme that releases the terminal orthophosphate group from linear polyphosphates. Northern blots showed that both single-copy genes are induced by long-term inorganic phosphate (P(i)) starvation, transcript levels being markedly increased (ca. 10- and 20-fold, respectively) relative to P(i)-sufficient cells. Concurrent increases of both PPA and PPX specific activities and protein levels by P(i) deprivation were also observed. On the other hand, a knockout mutant was obtained by insertional mutagenesis of ppx, but it could not be achieved with ppa, thus indicating that PPA function is essential for cell viability. Moreover, whereas the ppx mutant exhibited under P(i)-sufficient conditions lower growth rates than the wild-type and was certainly devoid of PPX activity, it showed a severe reduction of the PPA levels. These results are the first evidence on the involvement of both PPA and PPX in a possible intracellular P(i)-recycling enzymatic process activated under P(i)-starvation.

Base Sequence↗
PubMed2006

The use of whole genome amplification to study chromosomal changes in prostate cancer: insights into genome-wide signature of preneoplasia associated with cancer progression.

BACKGROUND: Prostate cancer (CaP) is a disease with multifactorial etiology that includes both genetic and environmental components. The knowledge of the genetic basis of CaP has increased over the past years, mainly in the pathways that underlie tumourigenesis, progression and drug resistance. The vast majority of cases of CaP are adenocarcinomas that likely develop through a pre-malignant lesion and high-grade prostatic intraepithelial neoplasia (HPIN). Histologically, CaP is a heterogeneous disease consisting of multiple, discrete foci of invasive carcinoma and HPIN that are commonly interspersed with benign glands and stroma. This admixture with benign tissue can complicate genomic analyses in CaP. Specifically, when DNA is bulk-extracted the genetic information obtained represents an average for all of the cells within the sample. RESULTS: To minimize this problem, we obtained DNA from individual foci of HPIN and CaP by laser capture microdissection (LCM). The small quantities of DNA thus obtained were then amplified by means of multiple-displacement amplification (MDA), for use in genomic DNA array comparative genomic hybridisation (gaCGH). Recurrent chromosome copy number abnormalities (CNAs) were observed in both HPIN and CaP. In HPIN, chromosomal imbalances involving chromosome 8 where common, whilst in CaP additional chromosomal changes involving chromosomes 6, 10, 13 and 16 where also frequently observed. CONCLUSION: An overall increase in chromosomal changes was seen in CaP compared to HPIN, suggesting a universal breakdown in chromosomal stability. The accumulation of CNAs, which occurs during this process is non-random and may indicate chromosomal regions important in tumourigenesis. It is therefore likely that the alterations in copy number are part of a programmed cycle of events that promote tumour development, progression and survival. The combination of LCM, MDA and gaCGH is ideally suited for the identification of CNAs from small cell clusters and may assist in the discovery of potential genomic markers for early diagnosis, or identify the location of tumour suppressor genes (TSG) or oncogenes previously unreported in HPIN and CaP.

Chromosome Aberrations↗
PubMedDate not supplied

The role of a conventional simulator in multileaf-plan simulation: a proposal.

Modern computer networks provide satisfying levels of data recording and verification between the treatment planning system (TPS) and the accelerators, while the main weakness of the preparation chain remains the simulation. When a conventional simulator is employed, it may adversely affect the three-dimensional treatment planning system (3DPS) process because of the difficulty to document the leaf positions on the simulator location films and on the patient's skin. With a conventional simulator, hard copies of the DRRs of each field and CT scans at isocenter level are needed. In an attempt to transfer more information displayed from a BEV perspective from the 3DPS to simulator radiographs, this study aimed to reduce the quality loss by using a 2D conventional simulator in a 3DPS process. We realized an acetate photocopy of TPS data for each field, from a BEV perspective, containing: DRR, wire frames of the PTV, organs at risk and MLC aperture. The photocopies, with an appropriate magnification factor to obtain a correct projective value (ratio 1:1) at isocenter level, are carefully placed on the radiographic images on the same hard copy which allows us to better understand possible setup errors and obliges us to correct these. The method provides reliable documentation, facilitates treatment verification, and fulfils the criteria for MLC simulation. It is accurate, simple, and very inexpensive.

Humans↗
PubMed2006

Klusters, NeuroScope, NDManager: a free software suite for neurophysiological data processing and visualization.

Recent technological advances now allow for simultaneous recording of large populations of anatomically distributed neurons in behaving animals. The free software package described here was designed to help neurophysiologists process and view recorded data in an efficient and user-friendly manner. This package consists of several well-integrated applications, including NeuroScope (http://neuroscope.sourceforce.net), an advanced viewer for electrophysiological and behavioral data with limited editing capabilities, Klusters (http://klusters.sourceforge.net), a graphical cluster cutting application for manual and semi-automatic spike sorting, NDManager (GPL,see http://www.gnu.org/licenses/gpl.html), an experimental parameter and data processing manager. All of these programs are distributed under the GNU General Public License (GPL, see ), which gives its users legal permission to copy, distribute and/or modify the software. Also included are extensive user manuals and sample data, as well as source code and documentation.

Animals↗
PubMed1995

Informed consent in emergency research. Consensus statement from the Coalition Conference of Acute Resuscitation and Critical Care Researchers.

OBJECTIVE: A coalition conference of acute resuscitation researchers was held to discuss the feasibility of applying current federal research regulations regarding informed consent to the emergency setting. This article presents consensus recommendations for regulatory changes for consent in emergency research. PARTICIPANTS: Representatives from the Society for Academic Emergency Medicine and the American Heart Association identified several professional organizations as stakeholders in this issue, including research, clinical, bioethics, legal, and patient advocacy groups. The Office for Protection From Research Risks (OPRR), the Food And Drug Administration (FDA), and staff from specific legislative offices were also invited to observe. Forty-three participants attended, including representatives from 12 professional organizations, five medical institutions, and the FDA and OPRR. This was a closed meeting. Participants were self-funded or sponsored by their professional organizations. EVIDENCE: Before the meeting, a draft of a position statement was developed by the conference organizers based on the current literature and discussions with experts in the field. This draft, copies of the current federal research regulations, and supporting articles were distributed before the conference. CONSENSUS PROCESS: Participants rotated through moderated discussion sessions to comment on subsections of the draft. Following discussion, a working draft was developed and distributed to each participant and represented organizational board for final review. All comments were considered in the final version of the document. CONCLUSIONS: We believe there are circumstances when it is not feasible to obtain prospective or proxy consent for enrollment into an emergency research protocol. In these circumstances, patients are vulnerable, not only to research risks, but also to being denied potentially beneficial therapy when there is no known effective treatment for their life-threatening condition. We offer recommendations that should be met when the critical nature of the illness or injury or the need to apply an investigational therapy rapidly precludes prospective consent for participation in emergency research.

Consensus↗
PubMed1996

Analysis of optical density wave propagation and cell movement during mound formation in Dictyostelium discoideum.

Aggregation fields of Dictyostelium amoebae are organized by propagating concentric or spiral waves of cAMP. These waves coordinate cell movement directed toward the aggregation center. We now systematically investigated dark-field wave propagation and chemotactic cell movement during late aggregation and mound formation. The period and the signal propagation velocity decreased continuously during aggregation leading to a 15-fold decrease of the chemical wavelength. By analyzing the behavior of single GFP-labeled cells in aggregates and mounds we measured cell movement velocity, changes in cell shape, periodicity of cell movement, and cell trajectories. In early mounds of strain AX-3 dark-field waves propagated frequently as multiarmed (high-frequency) spirals. During the high-frequency waves observed in the early mound stage, cell movement speed is low and cell movement rather undirected. During tip formation the wave period decreased again and the cells started to rotate in the mound at unusually high average speeds of 40 microns/min. The rotation was almost monotonic with no clear periodicity. Since at this time the majority of the cells had already differentiated into prespore cells, this implies that prespore cells moved faster than aggregation stage cells. At 12 hr of development cell movement velocity dropped again and became highly periodic. These measurements show that the relay system is characterized by a specific temporal evolution, which is closely correlated with cellular differentiation. The remarkable changes in cell movement speed and period indicate a qualitative change in signal and movement parameters which might well be caused by the observed switch from high- to low-affinity cAMP receptors during mound formation. This switch might be required to copy with the increase in cell density and most likely plays a crucial role in the process of cell sorting.

Animals↗
PubMed1988

Difference between two major size classes of carrot rDNA repeating units is due to reiteration of sequences of about 460 bp in the large spacer.

Based on the previous observation that the different regions between two major size classes of carrot rRNA genes are located in their large spacer, detailed physical maps of the spacer region were constructed by cloning, followed by restriction analysis. As a result, the different regions were restricted to BamHI segments of 1.3 kb and 1.8 kb. Sequence analysis of these segments revealed that the shorter one carried one truncated and two complete copies of about 460 bp of repetitious sequences, while the longer one contained one truncated and three full copies of the repetitious sequences. S1 nuclease mapping data suggest that either transcription initiation sites or processing sites of precursor rRNA are located in the repetitious sequence closest to the 18 S rRNA coding region.

Base Sequence↗
PubMed2002

Molecular characterization of a mouse cDNA encoding Dicer, a ribonuclease III ortholog involved in RNA interference.

Members of the ribonuclease III superfamily of double-stranded(ds)-RNA-specific endoribonucleases participate in diverse cellular RNA maturation and degradation pathways. A recently identified eukaryotic RNase III family member, named "Dicer", functions in the RNA interference (RNAi) pathway by producing 21--23 bp dsRNAs which target the selective destruction of homologous RNAs. RNAi is operative in animals, plants, and fungi, where it is proposed to inhibit viral reproduction and retroposon movement, as well as to participate in developmental pathways. RNAi functions in mammalian cells, including mouse oocytes and embryos. This article reports the cDNA sequence characterization and expression analysis of the mouse Dicer ortholog. On the basis of the cDNA sequence, the Dicer polypeptide is 1906 amino acids and has a predicted molecular mass of 215 kDa. Mouse Dicer contains a DExH/DEAH helicase motif; a PAZ domain; a tandem repeat of RNase III catalytic domain sequences; and a dsRNA-binding motif. The Dicer gene maps to a single locus on the distal portion of mouse Chromosome (Chr) 12. The Dicer transcript is expressed from the embryonic through adult stages of development. The Dicer transcript is also present in a wide variety of adult mouse organs. The highly conserved set of functional domains and the occurrence of a single-copy gene strongly indicate that the encoded protein is the RNase III ortholog responsible for dsRNA processing in the RNAi pathway.

Amino Acid Sequence↗
PubMed2005

Functional constraint and divergence in the G protein family in Caenorhabditis elegans and Caenorhabditis briggsae.

Part of the challenge of the post-genomic world is to identify functional elements within the wide array of information generated by genome sequencing. Although cross-species comparisons and investigation of rates of sequence divergence are an efficient approach, the relationship between sequence divergence and functional conservation is not clear. Here, we use a comparative approach to examine questions of evolutionary rates and conserved function within the guanine nucleotide-binding protein (G protein) gene family in nematodes of the genus Caenorhabditis. In particular, we show that, in cases where the Caenorhabditis elegans ortholog shows a loss-of-function phenotype, G protein genes of C. elegans and Caenorhabditis briggsae diverge on average three times more slowly than G protein genes that do not exhibit any phenotype when mutated in C. elegans, suggesting that genes with loss of function phenotypes are subject to stronger selective constraints in relation to their function in both species. Our results also indicate that selection is as strong on G proteins involved in environmental perception as it is on those controlling other important processes. Finally, using phylogenetic footprinting, we identify a conserved non-coding motif present in multiple copies in the genomes of four species of Caenorhabditis. The presence of this motif in the same intron in the gpa-1 genes of C. elegans, C. briggsae and Caenorhabditis remanei suggests that it plays a role in the regulation of gpa-1, as well as other loci.

Animals↗
PubMed2005

Identification and functional characterization of Candida albicans CDC4.

The CDC4 gene of Saccharomyces cerevisiae encodes an essential function that is required for G1-S and G2-M transitions during mitosis and at various stages during meiosis. We have isolated a functional homologue of CDC4 (CaCDC4) from the pathogenic yeast Candida albicans by complementing the S. cerevisiae cdc4-3 mutation with CaCDC4 expressed from its own promoter on a single-copy vector. The predicted product of CaCDC4 has 37% overall identity to the S. cerevisiae Cdc4 protein, although this identity is biased towards the C-terminal region of the two proteins which contains eight copies of the degenerate WD-40 motif, an element found in proteins that regulate diverse biological processes and an F-box domain proximal to the first iteration of the WD-40 motif. Both the F-box domain and WD-40 motifs appear necessary for the mitotic functions of Cdc4 in both yeasts. In contrast to its conserved role in mitosis, C. albicans CDC4 is unable to rescue the meiotic deficiency in a S. cerevisiae cdc4 homozygous diploid under restrictive conditions, even when expressed from an efficient S. cerevisiae promoter. In opposition to S. cerevisiae CDC4 being essential, C. albicans CDC4 appears to be nonessential and in its absence is critical for filamentous growth in C. albicans.

Alleles↗
PubMed1985

Nasopharyngeal carcinoma. Brief review.

Nasopharyngeal carcinoma has a well-defined geographic distribution, primarily affecting persons from southern China and Southeast Asia. Environmental factors are numerous and appear to have a secondary role, mainly in the promotion of the neoplastic process. Relationship with the Epstein-Barr virus is indicated by the identification of viral genome copies within the cells and by a persistent host antibody response with restricted specificity for nasopharyngeal malignancies. The World Health Organization has recently adopted a histologic classification categorized into three subtypes according to the degree of epithelial differentiation, keratinization, and stromal lymphocytic infiltration. The tumor expands locally to contiguous structures, spreads through the cervical lymphatics following the jugular chain, and eventually metastasizes to the skeleton and liver. Primary management consists of radiation therapy to cervicofacial fields and usually offers adequate palliation, with a five-year median survival of 67 percent for stage I and 17 percent for stage IV disease.

Antibodies, Viral↗
PubMed1986

Expression and transmission of a rodent retrovirus-like (VL30) gene family.

A transcriptionally active sub-set of the dispersed mouse VL30 family of proviral genetic elements was shown to be highly transmissible as a murine leukaemia virus pseudotype. Newly acquired VL30 proviruses (present at 1 to 2 copies per cell) were shown to be transcriptionally active. These data substantiate the hypothesis that this process of duplicative transposition may have played a major role in the evolution of the gene family and also demonstrate that VL30 elements would be capable of mediating oncogene activation by a promoter-insertion-type mechanism during leukaemia virus-induced tumourgenesis.

Animals↗
PubMed1991

In vitro rescue of an integrated hybrid adeno-associated virus/simian virus 40 genome.

In an in vitro simian virus 40 (SV40) DNA replication assay, we have observed excision of a hybrid adeno-associated virus (AAV)/SV40 insert from a plasmid construct. The excision was dependent on the presence of the palindromic AAV terminal repeat and greatly enhanced by the addition of the SV40 T antigen to the reaction. Analysis of the excision product supports a model in which the palindromic terminal sequences of AAV form a cruciform structure (equivalent to a Holliday recombination intermediate), which is cleaved and resealed so that the excision products are linear duplex pBR322 and linear duplex AAV/SV40 insert. Both the excised linear duplex pBR322 and the excised linear duplex AAV/SV40 insert have each terminus covalently crosslinked by one copy of the palindromic region of the AAV terminal repeat region folded on itself. The excision process may be a model system for cellular homologous recombination. The process as observed was either concomitant with or subsequent to DNA replication.

Adenoviruses, Human↗
PubMed1985

Nucleotide sequences responsible for generation of internally deleted Sendai virus defective interfering genomes.

The deletion points of four internally deleted defective interfering (DI) RNA species (7a, 7b, 7c, and 7d) that reside in a single Sendai virus strain were defined by nucleotide sequencing. DI RNA 7a (Mr 1.24 x 10(6)) retained the entire NP gene with the complete NP protein-coding sequence, except for the last two U residues of the polyadenylation signal, fused to an 1800-nucleotide sequence comprising 5'-terminal genome and adjacent L gene sequences. DI RNA 7b (Mr, 0.70 x 10(6)) consisted of 100 3'-terminal nucleotides fused to 1900 5'-terminal bases; the deletion point in the NP gene precedes the NP protein initiation codon. DI RNA 7c (Mr 0.55 x 10(6)) retained 420 3'-terminal and 1150 5'-terminal nucleotides. The sequence just downstream of the sequenced deletion site is M gene specific, indicating that 7c arose from at least two deletion events and that it comprises NP, M, and L gene fragments. Transcription of RNA 7c could yield an MRNA encoding a fusion protein with a 14,000 Mr (N-terminal NP sequence fused to out of frame M-specific amino acids). DI RNA 7d (Mr 0.92 x 10(6)) retained 1027 3'-terminal nucleotides fused to 1600 bases from the 5'-terminus. It has an open reading frame for a 33,000 Mr N-terminal NP protein fragment. Nucleotide sequences flanking each deletion and just downstream of the NP gene deletion site suggested that these DI genomes were generated by a copy-choice mechanism, involving polymerase jumping during replication of negative polarity virus genome templates. In this process, the termination and reinitiation of RNA synthesis would involve recognition of sequences that regulate virus genome transcription and replication.

Base Sequence↗