bluuebunny/crossref_metadata_2025 huggingface dataset:
The Effectiveness of Interventions for Improving the Research Literacy of Nurses: A Systematic Review
Authors: Hines Sonia , Ramsbotham Joanne , Coyer Fiona
Published: September 2015

Background

Internationally, a considerable body of research exists examining why nurses do not use evidence in practice. Consistently, the research finds that lack of knowledge about research or discomfort with understanding research terminology are among the chief reasons given. Research education is commonly included in undergraduate nursing degree programs, but this does not seem to translate into a strong understanding of research following graduation, or an ability to use it in practice.

Aim

The objective of this review was to identify the effectiveness of workplace, tertiary‐level educational, or other interventions designed to improve or increase postregistration nurses’ understanding of research literature and ability to critically interact with research literature with the aim of promoting the use of research evidence in practice in comparison to no intervention, other intervention, or usual practice.

Methods

A wide range of databases were searched for quantitative studies of registered nurses receiving educational interventions designed to increase or improve their understanding of research literature in tertiary or workplace settings. Two reviewers working independently critically appraised the relevant papers and extracted the data using Joanna Briggs Institute instruments. Data are presented as a narrative summary as no meta‐analysis was possible.

Results

Searching identified 4,545 potentially relevant papers, and after the sifting of titles and abstracts, 96 papers were selected for retrieval. On examination of full‐text versions, 10 of the 96 retrieved papers were found to meet the inclusion criteria. Included studies were low to moderate quality. Interactive or activity‐based learning seems to be effective in terms of improving research knowledge, critical appraisal ability, and research self‐efficacy. Utilizing a program with a strong base in an appropriate theory also seems to be associated with greater effectiveness, particularly for workplace interventions.

Linking Evidence to Action

The included studies strongly favored interactive interventions, and those utilizing theory in their construction. Therefore, these types of interventions should be implemented to improve the effectiveness of research education for nurses as well as their research literacy.
Is immune system‐related hypertension associated with ovarian hormone deficiency?
Authors: Sandberg Kathryn , Ji Hong , Einstein Gillian , Au April , Hay Meredith
Published: September 2015

New Findings

What is the topic of this review?

This review summarizes recent data on the role of ovarian hormones and sex in inflammation‐related hypertension.

What advances does it highlight?

The adaptive immune system has recently been implicated in the development of hypertension in males but not in females. The role of the immune system in the development of hypertension in women and its relationship to ovarian hormone production are highlighted.

The immune system is known to contribute to the development of high blood pressure in males. However, the role of the immune system in the development of high blood pressure in females and the role of ovarian hormones has only recently begun to be studied. In animal studies, both the sex of the host and the T cell are critical biological determinants of susceptibility and resistance to hypertension induced by angiotensin II. In women, natural menopause is known to result in significant changes in the expression of genes regulating the immune system. Likewise, in animal models, ovariectomy results in hypertension and an upregulation in T‐cell tumour necrosis factor‐α‐related genes. Oestrogen replacement results in decreases in inflammatory genes in the brain regions involved in blood pressure regulation. Together, these studies suggest that the response of the adaptive immune system to ovarian hormone deficiency is a significant contributor to hypertension in women.
State Space Model for Thermal Analysis of Integrated Structure of Flat Plate Solar Collector and Building Envelope
Authors: Yu Guoqing , Zhou Jirui , Tang Yongqiang
Published: September 2015

Active types of integration of flat plate solar collectors and building envelopes are studied in this paper. The integrated structure of flat plate solar collector and building envelope includes glass cover, absorber plate, tubes, back insulation, and building envelope (we will call it integrated structure later in this paper). With the solar collector integrated with building, the boundary conditions of heat transfer both for the solar collector and the building envelope are changed significantly, and the thermal performance of solar collection and building heat transfer characteristics influences each other. The state space model for thermal analysis of the integrated structure is proposed in this paper, and method for solving this state space model is provided. Moreover, thermal analysis for a particular integrated structure was conducted both by state space model and fluent simulation, then the results were compared and agree well. The state space model has great advantages in time-spending over fluent simulation and it can be used for long-term (several months or a whole year) simulation of the integrated structure. Comparison were made between the integrated structure, detached solar collector and detached single wall based on results calculated by state space method. It shows that (1) integration has little impact on the thermal efficiency of solar collection and the useful heat gain of the integrated structure are nearly the same as that for the single detached solar collector under the same ambient conditions; (2) integration has significant impact on the heat flux across the wall, and the heat flux of the integrated structure is much less than the detached single wall.
Numerical Analysis of Carrier Particle Motion in a Dry Powder Inhaler
Authors: Sommerfeld Martin , Schmalfuß Silvio
Published: September 2015

The efficiency of dry powder inhalers (DPIs) for drug delivery is still very low and is therefore the objective of intensive research. Thus, numerical calculations (computational fluid dynamics (CFD)) using the Euler/Lagrange approach without coupling are being performed in order to analyze flow structure and carrier particle motion within a typical inhaler device. These computations are being performed for a steady-state situation with a flow rate of 100 l/min. Essential for the detachment of the very fine drug powder (i.e., between 1 and 5 μm) from the carrier particles are the fluid stresses experienced by such particles (i.e., relative velocity, turbulence, and fluid shear) as well as wall collisions, which are both evaluated in the present study. Since the carrier particles are rather large (i.e., normally 50–100 μm), first the importance of different relevant fluid forces, especially transverse lift forces, is investigated. Moreover, the significance of the parameters in the particle–wall collision model is highlighted and a statistical analysis of particle–wall collisions in an inhaler is conducted. The improved understanding of particle motion in the normally very complex flows of inhalers will be the basis for optimizing inhaler design.
Thermal-Hydraulic Performance of SiC-Water and Al2O3-Water Nanofluids in the Minichannel
Authors: Zhang Ji , Diao Yanhua , Zhao Yaohua , Zhang Yanni
Published: September 2015

The single-phase flow and heat transfer behaviors of SiC and Al2O3 nanoparticles dispersed in water were studied experimentally in a multiport minichannel flat tube (MMFT). The volume concentrations of the two nanofluids ranged from 0.001% to 1%. Their effective particle sizes, thermal conductivities, and viscosities were also measured. Results indicated that these nanofluids as a working fluid could enhance heat transfer but increase pressure drop and the Nusselt number by up to 85%. The two nanofluids exhibited a common optimal volume concentration of 0.01% for heat transfer. Effective particle size was also found to have a significant effect on heat transfer.
Effect of Domain Size on Fluid–Particle Statistics in Homogeneous, Gravity-Driven, Cluster-Induced Turbulence
Authors: Capecelatro Jesse , Desjardins Olivier , Fox Rodney O.
Published: September 2015

Scaling of volume fraction and velocity fluctuations with domain size is investigated for high-mass-loading suspensions of finite-size inertial particles subject to gravity. Results from highly resolved Euler–Lagrange simulations are evaluated via an adaptive spatial filter with an averaging volume that varies with the local particle concentration. This filter enables the instantaneous particle velocity to be decomposed into a spatially correlated contribution used in defining the particle-phase turbulent kinetic energy (TKE), and a spatially uncorrelated contribution used in defining the granular temperature. The total granular energy is found to grow nearly linearly with the domain size, while the balance between the separate contributions remains approximately constant.
Lessons from Ebola: Improving infectious disease surveillance to inform outbreak management
Authors: Woolhouse Mark E. J. , Rambaut Andrew , Kellam Paul
Published: September 2015

A maximally effective public health response to infectious disease outbreaks requires the framework of an integrated surveillance system.
Efficient modification of
CCR5
in primary human hematopoietic cells using a megaTAL nuclease and AAV donor template
Authors: Sather Blythe D. , Romano Ibarra Guillermo S. , Sommer Karen , Curinga Gabrielle , Hale Malika , Khan Iram F. , Singh Swati , Song Yumei , Gwiazda Kamila , Sahni Jaya , Jarjour Jordan , Astrakhan Alexander , Wagner Thor A. , Scharenberg Andrew M. , Rawlings David J.
Published: September 2015

Therapeutic coding sequences can be targeted to the
CCR5
locus of primary human T cells with high efficiency by using megaTAL nuclease and an AAV donor template.
Ontogenetic Differences in Dietary Fat Influence Microbiota Assembly in the Zebrafish Gut
Authors: Wong Sandi , Stephens W. Zac , Burns Adam R. , Stagaman Keaton , David Lawrence A. , Bohannan Brendan J. M. , Guillemin Karen , Rawls John F.
Published: September 2015

Gut microbiota influence the development and physiology of their animal hosts, and these effects are determined in part by the composition of these microbial communities. Gut microbiota composition can be affected by introduction of microbes from the environment, changes in the gut habitat during development, and acute dietary alterations. However, little is known about the relationship between gut and environmental microbiotas or about how host development and dietary differences during development impact the assembly of gut microbiota. We sought to explore these relationships using zebrafish, an ideal model because they are constantly immersed in a defined environment and can be fed the same diet for their entire lives. We conducted a cross-sectional study in zebrafish raised on a high-fat, control, or low-fat diet and used bacterial 16S rRNA gene sequencing to survey microbial communities in the gut and external environment at different developmental ages. Gut and environmental microbiota compositions rapidly diverged following the initiation of feeding and became increasingly different as zebrafish grew under conditions of a constant diet. Different dietary fat levels were associated with distinct gut microbiota compositions at different ages. In addition to alterations in individual bacterial taxa, we identified putative assemblages of bacterial lineages that covaried in abundance as a function of age, diet, and location. These results reveal dynamic relationships between dietary fat levels and the microbial communities residing in the intestine and the surrounding environment during ontogenesis.

IMPORTANCE
The ability of gut microbiota to influence host health is determined in part by their composition. However, little is known about the relationship between gut and environmental microbiotas or about how ontogenetic differences in dietary fat impact gut microbiota composition. We addressed these gaps in knowledge using zebrafish, an ideal model organism because their environment can be thoroughly sampled and they can be fed the same diet for their entire lives. We found that microbial communities in the gut changed as zebrafish aged under conditions of a constant diet and became increasingly different from microbial communities in their surrounding environment. Further, we observed that the amount of fat in the diet had distinct age-specific effects on gut community assembly. These results reveal the complex relationships between microbial communities residing in the intestine and those in the surrounding environment and show that these relationships are shaped by dietary fat throughout the life of animal hosts.
Insufficient Acidification of Autophagosomes Facilitates Group A Streptococcus Survival and Growth in Endothelial Cells
Authors: Lu Shiou-Ling , Kuo Chih-Feng , Chen Hao-Wen , Yang Yi-Shuan , Liu Ching-Chuan , Anderson Robert , Wu Jiunn-Jong , Lin Yee-Shin
Published: September 2015

Group A streptococcus (GAS) is an important human pathogen, and its invasion via blood vessels is critically important in serious events such as bacteremia or multiorgan failure. Although GAS was identified as an extracellular bacterium, the internalization of GAS into nonphagocytic cells may provide a strategy to escape from immune surveillance and antibiotic killing. However, GAS has also been reported to induce autophagy and is efficiently killed within lysosome-fused autophagosomes in epithelial cells. In this study, we show that GAS can replicate in endothelial cells and that streptolysin O is required for GAS growth. Bacterial replication can be suppressed by altering GAS gene expression in an acidic medium before internalization into endothelial cells. The inhibitory effect on GAS replication can be reversed by treatment with bafilomycin A1, a specific inhibitor of vacuolar-type H
+
-ATPase. Compared with epithelial cells in which acidification causes autophagy-mediated clearance of GAS, there was a defect in acidification of GAS-containing vesicles in endothelial cells. Consequently, endothelial cells fail to maintain low pH in GAS-containing autophagosomes, thereby permitting GAS replication inside LAMP-1- and LC3-positive vesicles. Furthermore, treatment of epithelial cells with bafilomycin A1 resulted in defective GAS clearance by autophagy, with subsequent bacterial growth intracellularly. Therefore, low pH is a key factor for autophagy-mediated suppression of GAS growth inside epithelial cells, while defective acidification of GAS-containing vesicles results in bacterial growth in endothelial cells.

IMPORTANCE
Previous reports showed that GAS can induce autophagy and is efficiently killed within lysosome-fused autophagosomes in epithelial cells. In endothelial cells, in contrast, induction of autophagy is not sufficient for GAS killing. In this study, we provide the first evidence that low pH is required to prevent intracellular growth of GAS in epithelial cells and that this mechanism is defective in endothelial cells. Treatment of GAS with low pH altered GAS growth rate and gene expression of virulence factors and resulted in enhanced susceptibility of GAS to intracellular lysosomal killing. Our findings reveal the existence of different mechanisms of host defense against GAS invasion between epithelial and endothelial cells.
Transcriptional and post-transcriptional control of PHO8 expression by PHO regulatory genes in Saccharomyces cerevisiae.
Authors: Kaneko Y , Tamai Y , Toh-e A , Oshima Y
Published: September 2015

A DNA fragment bearing the PHO8 gene, which encodes repressible alkaline phosphatase of Saccharomyces cerevisiae, was cloned. Northern hybridizations with the PHO8 DNA as probe indicated that the PHO8 transcript is 1.8 kilobases in length and is more abundant in cells grown in low-phosphate medium than in high-phosphate medium. The pho9 mutant, whose phenotype is defective in the activity of repressible alkaline phosphatase, produced as much of the PHO8 transcript as did the PHO9+ cells. Hence, the PHO9 product should act at the post-transcriptional level. The pho4 mutant could not derepress the PHO8 transcript, whereas the pho80 mutant could, irrespective of the amount of Pi in the medium, as has been suggested by genetic study.
NIH/3T3 cells transfected with human tumor DNA containing activated ras oncogenes express the metastatic phenotype in nude mice.
Authors: Thorgeirsson U P , Turpeenniemi-Hujanen T , Williams J E , Westin E H , Heilman C A , Talmadge J E , Liotta L A
Published: September 2015

NIH/3T3 cells transfected with DNA from malignant human tumors produced experimental and spontaneous metastases in nude mice. In contrast, parent or spontaneously transformed NIH/3T3 cells failed to metastasize. The transfected clones contained either activated c-Harvey-ras or N-ras oncogenes. A representative clone (T71-17SA2) which was used to assess selected cellular and host factors relevant to the metastatic process produced lung metastases in 100% of the NIH nude mice recipients, secreted augmented levels of type IV collagenase, and invaded human amnion basement membrane in vitro. Expression of the metastatic phenotype was not related to decreased sensitivity to natural killer cells or macrophage-mediated cytotoxicity. Analysis of the cellular DNA from the T71-17SA2 transfectant and its corresponding metastases, both of which contained activated N-ras oncogenes, revealed a twofold increase in the N-ras-specific DNA sequences in the metastatic cells. Thus, transfection with human tumor DNA containing activated ras oncogenes can induce the complete metastatic phenotype in NIH/3T3 cells by a mechanism apparently unrelated to immune cell killing.
pp60c-src in the developing cerebellum.
Authors: Fults D W , Towle A C , Lauder J M , Maness P F
Published: September 2015

pp60c-src was localized in the cerebellum of developing chicken embryos by immunoperoxidase staining with antisera raised against bacterially expressed pp60v-src. Immunoreactivity (IR) appeared in the cerebellum of the chicken embryos at the time of neuronal differentiation. pp60c-src IR was detected in regions of the developing cerebellum where processes of developing neurons and glia are located. In the early embryo (stage 17), pp60c-src IR was localized in the marginal zone of the cerebellar plate. By stage 40, pp60c-src IR was localized in the process-rich molecular layer of the cerebellum and between the cells of the developing internal granular layer. Cell bodies of cerebellar neurons did not show pp60c-src IR at any stage of development. Mitotically active neuroepithelial cells of the metencephalon did not express pp60c-src before the onset of differentiation in the early embryo, nor did proliferating cells of the external granular layer express pp60c-src at later stages. Although it is not possible to ascertain whether pp60c-src is localized in developing neurons or glia at the light microscope level, the time of its appearance and pattern of distribution in the molecular layer is suggestive of a localization within the developing neuronal processes which compose the bulk of this layer. Biochemical analyses of pp60c-src in the developing cerebellum by the immune complex protein kinase activity and sensitivity of the kinase to inhibition by P1,P4-di(adenosine-5')tetraphosphate confirmed that the expression of pp60c-src coincided with the time of neuronal differentiation. We conclude from these results that in the central nervous systems, pp60c-src may be more important in an aspect of cell differentiation or a mature neuronal function than in the proliferation of neuronal or glial precursors.
Efficient correction of mismatched bases in plasmid heteroduplexes injected into cultured mammalian cell nuclei.
Authors: Folger K R , Thomas K , Capecchi M R
Published: September 2015

Heteroduplexes were prepared from two plasmids, pRH4-14/TK and pRH5-8/TK, containing different amber mutations in the neomycin resistance gene (Neor). The Neor gene was engineered to be expressed in both bacterial and mammalian cells. A functional Neor gene conferred kanamycin resistance to bacteria and resistance to the drug G418 to mammalian cells. In addition, the plasmids contained restriction site polymorphisms which did not confer a selectable phenotype but were used to follow the pattern of correction of mismatched bases in the heteroduplexes. In a direct comparison of the efficiency of transforming mouse LMtk- cells to G418r, the injection of heteroduplexes of pRH4-14/TK-pRH5-8/TK was 10-fold more efficient than the coinjection of pRH4-14/TK and pRH5-8/TK linear plasmid DNA. In fact, injection of 5 to 10 molecules of heteroduplex DNA per cell was as efficient in transforming LMtk- cells to G418r as the injection of 5 to 10 molecules of linear plasmid DNA per cell containing a wild-type Neor gene. To determine the pattern of mismatch repair of the injected heteroduplexes, plasmids were "rescued" from the G418r cell lines. From this analysis we conclude that the generation of wild-type Neor genes from heteroduplex DNA proceeds directly by correction of the mismatched bases, rather than by alternative mechanisms such as recombination between the injected heteroduplexes. Our finding that a cell can efficiently correct mismatched bases when confronted with preformed heteroduplexes suggests that this experimental protocol could be used to study a wide range of DNA repair mechanisms in cultured mammalian cells.
Developmentally regulated cytokeratin gene in Xenopus laevis.
Authors: Winkles J A , Sargent T D , Parry D A , Jonas E , Dawid I B
Published: September 2015

We have determined the sequence of cloned cDNAs derived from a 1,665-nucleotide mRNA which transiently accumulates during Xenopus laevis embryogenesis. Computer analysis of the deduced amino acid sequence revealed that this mRNA encodes a 47-kilodalton type I intermediate filament subunit, i.e., a cytokeratin. As is common to all intermediate filament subunits so far examined, the predicted polypeptide, named XK70, contains N- and C-terminal domains flanking a central alpha-helical rod domain. The overall amino acid homology between XK70 and a human 50-kilodalton type I keratin is 47%; homology within the alpha-helical domain is 57%. The N-terminal domain, which is not completely contained in our cDNAs, is basic, contains 42% serine plus alanine, and includes five copies of a six-amino-acid repeating unit. The C-terminal domain has a high alpha-helical content and contains a region with sequence homology to the C-terminal domains of other type I and type III intermediate filament proteins. We suggest that different keratin filament subtypes may have different functional roles during amphibian oogenesis and embryogenesis.
Unequal homologous recombination between tandemly arranged sequences stably incorporated into cultured rat cells.
Authors: Stringer J R , Kuhn R M , Newman J L , Meade J C
Published: September 2015

Cultured rat cells deficient in endogenous thymidine kinase activity (tk) were stably transformed with a recombination-indicator DNA substrate constructed in vitro by rearrangement of the herpes simplex virus tk gene sequences into a partially redundant permutation of the functional gene. The recombination-indicator DNA did not express tk, but was designed to allow formation of a functional tk gene via homologous recombination. A clonal cell line (519) was isolated that harbored several permuted herpes simplex virus tk genes. 519 cells spontaneously produced progeny that survived in medium containing hypoxanthine, aminopterin, and thymidine. Acquisition of resistance to hypoxanthine, aminopterin, and thymidine was accompanied by the rearrangement of the defective tk gene to functional configuration. The rearrangement apparently occurred by unequal exchange between one permuted tk gene and a replicated copy of itself. Recombination was between 500-base-pair tracts of DNA sequence homology that were separated by 3.4 kilobases. Exchanges occurred spontaneously at a frequency of approximately 5 X 10(-6) events per cell per generation. Recombination also mediated reversion to the tk- phenotype; however, the predominant mechanism by which cells escaped death in the presence of drugs rendered toxic by thymidine kinase was not recombination, but rather inactivation of the intact tk gene.
Sequence analysis of the human major histocompatibility gene SX alpha.
Authors: Boss J M , Mengler R , Okada K , Auffray C , Strominger J L
Published: September 2015

The DP subregion of the human major histocompatibility complex contains two closely linked gene pairs, DP alpha, DP beta and SX alpha, SX beta. The exon-intron organization and the complete DNA sequence of the SX alpha gene are reported here. There are several mutations within the SX alpha gene which strongly suggest that it is a pseudogene. These include two frameshift mutations, one in the alpha 1 domain and the other in the cytoplasmic domain. A 5' splice site mutation at the end of the alpha 1 exon also exists. DNA sequence homology between DP alpha and SX alpha suggests that these genes arose through a gene duplication event.
Vector expression of adenovirus type 5 E1a proteins: evidence for E1a autoregulation.
Authors: Smith D H , Kegler D M , Ziff E B
Published: September 2015

We transiently expressed adenovirus type C E1a proteins in wild-type or mutant form from plasmid vectors which have different combinations of E1a and simian virus 40 enhancer elements and which contain the DNA replication origin of SV40 and can replicate in COS 7 cells. We measured the levels of E1a mRNA encoded by the vectors and the transition regulation properties of the protein products. Three vectors encoded equivalent levels of E1a mRNA in COS 7 cells: (i) a plasmid encoding the wt 289-amino acid E1a protein (this complemented the E1a deletion mutant dl312 for early region E2a expression under both replicative and nonreplicative conditions); (ii) a vector for the wt 243-amino acid E1a protein (this complemented dl312 weakly and only under conditions of high multiplicities of dl312); (iii) a mutant, pSVXL105, in which amino acid residues-38 through 44 of the 289-amino acid E1a protein (which includes two highly conserved residues) are replaced by 3 novel amino acids (this also complemented dl312 efficiently). A fourth vector, mutant pSVXL3 with which linker substitution shifts the reading frame to encode a truncated 70-amino acid fragment from the amino terminus of the 289-amino acid protein, was unable to complement dl312. Surprisingly, pSVXL3 overexpressed E1a mRNA approximately 30-fold in COS 7 cells in comparison with the other vectors. The pSVXL3 overexpression could be reversed by cotransfection with a wt E1a vector. We suggest that wt E1a proteins regulate the levels of their own mRNAs through the recently described transcription repression functions of the 289- and 243-amino acid E1a protein products and that pSVXL3 fails to autoregulate negatively.
Signals for transcription initiation and termination in the Saccharomyces cerevisiae plasmid 2 micron circle.
Authors: Sutton A , Broach J R
Published: September 2015

By S1 nuclease protection experiments and primer extension analysis, we determined precisely the cap and polyadenylation sites of transcripts from the four genes of the yeast 2 micron circle plasmid, as well as those of other plasmid transcripts of unknown function. In addition, we used deletion analysis to identify sequences necessary for polyadenylation in plasmid transcripts. Our results indicate that plasmid genes constitute independent transcription units and that plasmid mRNAs are not derived by extensive processing of precursor transcripts. In addition, we found that the D coding region of 2 micron circle is precisely encompassed by a polyadenylated transcript, suggesting that this coding region constitutes a functional plasmid gene. Our identification of the position of plasmid polyadenylation sites and of sequences necessary for polyadenylation provides support for a tripartite signal for polyadenylation as proposed by Zaret and Sherman (K.S. Zaret and F. Sherman, Cell 28:563-573, 1982). Finally, these data highlight salient features of the transcriptional regulatory circuitry that underlies the control of plasmid maintenance in the cell.
Tsp transposons: a heterogeneous family of mobile sequences in the genome of the sea urchin Strongylocentrotus purpuratus.
Authors: Cohen J B , Liebermann D , Kedes L
Published: September 2015

In the preceding paper (J.B. Cohen, B. Hoffman-Liebermann, and L. Kedes, Mol. Cell. Biol., 5:2804-2813, 1985), we described the nucleotide sequence of ISTU4, which is a member of a new family of repetitive sequences, the Tsp family, present in a higher eucaryote, the sea urchin Strongylocentrotus purpuratus. We provided evidence that individual members of this family can act as transposable elements. Here we describe our structural analysis of the Tsp element family, which numbers about 1,000 members per haploid genome. Hybridization and nucleotide sequence analysis of several genomic Tsp clones demonstrate that structurally most Tsp elements resemble ISTU4. Tsp elements range in size up to about 1.3 kilobase pairs, have terminal domains that are conserved between the various examples studied, and contain a central portion of varying size, which may be extensively diverged. Structurally, however, the central portions are very similar and consist of several approximately 150-base-pairs-long, tandemly arranged, imperfect repeats, which are followed by a truncated repeat. The structural analysis is consistent with the possibility that the individual Tsp elements differ by multiples of these 150-base-pair repeats. One variant genomic clone has a solitary repeat and lacks the truncated repeat. The nucleotide sequences of different repeats of a single Tsp element can diverge extensively. The truncated repeat is divergent from most of the repeats, but in one case it is almost identical to a repeat of the same element. Comparison of the sequences from different elements enabled us to determine the boundaries of each structural domain and allows us to propose that each of these domains may be independent units of genetic information. Analysis of the population of Tsp-related sequences in the S. purpuratus genome by genomic blot hybridization suggests that most Tsp family members share the same overall structure. In addition, there is a structural element, about 70 base pairs long, that appears to interrupt the tandem arrangement of the 150-base-pair repeats at regular intervals.
v-src inhibits differentiation via an extracellular intermediate(s).
Authors: Langer-Safer P R , Lehrman S R , Skalka A M
Published: September 2015

Murine 3T3L1 preadipocytes transformed by avian sarcoma virus were unable to differentiate in response to insulin or dexamethasone plus 1-methyl-3-isobutylxanthine, both potent inducers of differentiation of the nontransformed 3T3L1 parental line. Conditioned medium from transformed cells contained a relatively heat-stable factor(s) which inhibited the differentiation of untransformed parental 3T3L1 cells but did not induce any changes in their morphology. A protease-sensitive mitogen was also detected in the medium. The relationship between the two activities remains to be elucidated.
Activation of c-myb expression by phytohemagglutinin stimulation in normal human T lymphocytes.
Authors: Torelli G , Selleri L , Donelli A , Ferrari S , Emilia G , Venturelli D , Moretti L , Torelli U
Published: September 2015

The expression of c-myb in normal human T lymphocytes directly derived from a normal subject and not adapted to continuous growth in culture was found to be markedly increased after phytohemagglutinin stimulation. In the same cells, the expression of c-myc mRNA is a much earlier event compared with the appearance of c-myb mRNA, which takes place soon after that of histone H3 mRNA. The increase in c-myb expression was not due to a particular T-lymphocyte subset, as shown by in situ hybridization assays.
Identification and characterization of the centromere from chromosome XIV in Saccharomyces cerevisiae.
Authors: Neitz M , Carbon J
Published: September 2015

A functional centromere located on a small DNA restriction fragment from Saccharomyces cerevisiae was identified as CEN14 by integrating centromere-adjacent DNA plus the URA3 gene by homologous recombination into the yeast genome and then by localizing the URA3 gene to chromosome XIV by standard tetrad analysis. DNA sequence analysis revealed that CEN14 possesses sequences (elements I, II, and III) that are characteristic of other yeast centromeres. Mitotic and meiotic analyses indicated that the CEN14 function resides on a 259-base-pair (bp) RsaI-EcoRV restriction fragment, containing sequences that extend only 27 bp to the right of the element I to III region. In conjunction with previous findings on CEN3 and CEN11, these results indicate that the specific DNA sequences required in cis for yeast centromere function are contained within a region about 150 bp in length.
Sequence-dependent DNA replication in preimplantation mouse embryos.
Authors: Wirak D O , Chalifour L E , Wassarman P M , Muller W J , Hassell J A , DePamphilis M L
Published: September 2015

Circular, double-stranded DNA molecules were injected into nuclei of mouse oocytes and one- or two-cell embryos to determine whether specific sequences were required to replicate DNA during mouse development. Although all of the injected DNAs were stable, replication of plasmid pML-1 DNA was not detected unless it contained either polyomavirus (PyV) or simian virus 40 (SV40) DNA sequences. Replication occurred in embryos, but not in oocytes. PyV DNA, either alone or recombined with pML-1, underwent multiple rounds of replication to produce superhelical and relaxed circular monomers after injection into one- or two-cell embryos. SV40 DNA also replicated, but only 3% as well as PyV DNA. Coinjection of PyV DNA with either pML-1 or SV40 had no effect on the replicating properties of the three DNAs. These results are consistent with a requirement for specific cis-acting sequences to replicate DNA in mammalian embryos, in contrast to sequence-independent replication of DNA injected into Xenopus eggs. Furthermore, PyV DNA replication in mouse embryos required PyV large T-antigen and either the alpha-beta-core or beta-core configuration of the PyV origin of replication. Although the alpha-core configuration replicated in differentiated mouse cells, it failed to replicate in mouse embryos, demonstrating cell-specific activation of an origin of replication. Replication or expression of PyV DNA interfered with normal embryonic development. These results reveal that mouse embryos are permissive for PyV DNA replication, in contrast to the absence of PyV DNA replication and gene expression in mouse embryonal carcinoma cells.
Isolation and DNA sequence of ADH3, a nuclear gene encoding the mitochondrial isozyme of alcohol dehydrogenase in Saccharomyces cerevisiae.
Authors: Young E T , Pilgrim D
Published: September 2015

The Saccharomyces cerevisiae nuclear gene, ADH3, that encodes the mitochondrial alcohol dehydrogenase isozyme ADH III was cloned by virtue of its nucleotide homology to ADH1 and ADH2. Both chromosomal and plasmid-encoded ADH III isozymes were repressed by glucose and migrated heterogeneously on nondenaturing gels. Nucleotide sequence analysis indicated 73 and 74% identity for ADH3 with ADH1 and ADH2, respectively. The amino acid identity between the predicted ADH III polypeptide and ADH I and ADH II was 79 and 80%, respectively. The open reading frame encoding ADH III has a highly basic 27-amino-acid amino-terminal extension relative to ADH I and ADH II. The nucleotide sequence of the presumed leader peptide has a high degree of identity with the untranslated leader regions of ADH1 and ADH2 mRNAs. A strain containing a null allele of ADH3 did not have a detectably altered phenotype. The cloned gene integrated at the ADH3 locus, indicating that this is the structural gene for ADH III.
Simian virus 40 minichromosomes contain torsionally strained DNA molecules.
Authors: Barsoum J , Berg P
Published: September 2015

Sundin and Varshavsky (J. Mol. Biol. 132:535-546, 1979) found that nearly two-thirds of simian virus 40 (SV40) minichromosomes obtained from nuclei of SV40-infected cells become singly nicked or cleaved across both strands after digestion with staphylococcal nuclease at 0 degrees C. The same treatment of SV40 DNA causes complete digestion rather than the limited cleavages produced in minichromosomal DNA. We have explored this novel behavior of the minichromosome and found that the nuclease sensitivity is dependent upon the topology of the DNA. Thus, if minichromosomes are pretreated with wheat germ DNA topoisomerase I, the minichromosomal DNA is completely resistant to subsequent digestion with staphylococcal nuclease at 0 degrees C. If the minichromosome-associated topoisomerase is removed, virtually all of the minichromosomes are cleaved to nicked or linear structures by the nuclease treatment. The cleavage sites are nonrandomly located; instead they occur at discrete loci throughout the SV40 genome. SV40 minichromosomal DNA is also cleaved to nicked circles and full-length linear fragments after treatment with the single strand-specific endonuclease S1; this cleavage is also inhibited by pretreatment with topoisomerase I. Thus, it may be that the nuclease sensitivity of minichromosomes is due to the transient or permanent unwinding of discrete regions of their DNA. Direct comparisons of the extent of negative supercoiling of native and topoisomerase-treated SV40 minichromosomes revealed that approximately two superhelical turns were removed by the topoisomerase treatment. The loss of these extra negative supercoils from the DNA probably accounts for the resistance of the topoisomerase-treated minichromosomes to the staphylococcal and S1 nucleases. These findings suggest that the DNA in SV40 intranuclear minichromosomes is torsionally strained. The functional significance of this finding is discussed.
Characterization of the myosin light-chain-2 gene of Drosophila melanogaster.
Authors: Parker V P , Falkenthal S , Davidson N
Published: September 2015

Recombinant DNA clones encoding the Drosophila melanogaster homolog of the vertebrate myosin light-chain-2 (MLC-2) gene have been isolated. This single-copy gene maps to the chromosomal locus 99E. The nucleotide sequence was determined for a 3.4-kilobase genomic fragment containing the gene and for two MLC-2 cDNA clones generated from late pupal mRNA. Comparison of these sequences shows that the gene contains two introns, the positions of which are conserved in the corresponding rat sequence. Extension of a primer homologous to the mRNA reveals two start sites for transcription 12 nucleotides apart. The sequence TATA is not present ahead of the mRNA cap site. There are two major sites of poly(A) addition separated by 356 nucleotides. The protein sequence derived from translation of the cDNA sequence shows a high degree of homology with that for the DTNB myosin light chain (MLC-2) of chicken. A lower degree of sequence homology was seen in comparisons with other evolutionarily related calcium-binding proteins. RNA blots show high levels of expression of several transcripts during the developmental time stages when muscle is being produced. In vitro translation of hybrid-selected RNA produces two polypeptides which comigrate on two-dimensional gels with proteins from Drosophila actomyosin, although the cDNA sequence reveals only one 26-kilodalton primary translation product.
Reversion analysis of mutations induced by 5-bromodeoxyuridine mutagenesis in mammalian cells.
Authors: Kaufman E R
Published: September 2015

Two protocols have been developed, both of which utilize the thymidine analog 5-bromodeoxyuridine (BrdUrd) to induce mutations in mammalian cells in culture (E. R. Kaufman and R. L. Davidson, Proc. Natl. Acad. Sci. USA 75:4982-4986, 1978; E. R. Kaufman, Mol. Cell. Biol. 4:2449-2454, 1984). The first protocol, termed incorporational (INC) mutagenesis, utilizes high concentrations of BrdUrd in the culture medium to generate a high intracellular ratio of BrdUTP/dCTP. The second protocol, termed replicational (REP) mutagenesis, entails the incorporation of BrdUrd into DNA under nonmutagenic conditions, the removal of all BrdUrd from the culture medium, and the subsequent replication of the bromouracil-containing DNA in the presence of high intracellular levels of dTTP and dGTP. Genetic studies using reversion analysis at the hypoxanthine-guanine phosphoribosyltransferase locus were used to determine whether the mechanisms of these two BrdUrd mutagenesis protocols had enough specificity to be distinguishable by their ability to revert various mutants. The results of these studies indicated that (i) mutants induced by INC mutagenesis were induced to revert only by REP mutagenesis and not by INC mutagenesis, (ii) mutants induced by REP mutagenesis were more efficiently reverted by INC mutagenesis than by REP mutagenesis, and (iii) both spontaneous mutants and mutants induced by the chemical mutagen ethyl methanesulfonate showed a high degree of specificity when tested for reversion by the BrdUrd mutagenesis protocols.
Regulation of cellular morphology by the Rous sarcoma virus src gene: analysis of fusiform mutants.
Authors: Rohrschneider L , Reynolds S
Published: September 2015

We have been interested in how Rous sarcoma virus (RSV) influences transformed cell morphology and compared the molecular properties of chicken embryo cells (CEC) infected with mutants of RSV that induce the fusiform transformed cell morphology with those of CEC infected by wild-type RSV, which induces the more normal round transformed cell morphology. We looked for properties shared by all fusiform mutant-infected cells, because these may be responsible for maintaining the fusiform morphology. Five different fusiform mutants, two wild-type RSVs, and one wild-type back revertant of a fusiform mutant were studied. In the fusiform mutant-infected cells, the localization and myristylation of pp60src were determined and the extent of expression of the extracellular matrix protein fibronectin was examined at both the mRNA and protein levels. The phosphorylation of vinculin on tyrosine also was examined in the same CEC. Within all fusiform mutant-transformed CEC, pp60src was dramatically absent from the adhesion plaque sites normally seen in cells transformed with wild-type RSV, and these transformed CEC all expressed more fibronectin mRNA and protein in the extracellular matrix than did the wild-type RSV-transformed CEC. The absence of pp60src from the adhesion plaques was not due to lack of myristylation of the src protein, and tyrosine phosphorylation of vinculin was not related to fibronectin expression. These results suggest that the inverse relationship between pp60src in the adhesion plaques and fibronectin expression in the extracellular matrix may be interconnected phenomena and could be related to the maintenance of the fusiform transformed morphology.
Alternative model for chromatin organization of the Saccharomyces cerevisiae chromosomal DNA plasmid TRP1 RI circle (YARp1).
Authors: Long C M , Brajkovich C M , Scott J F
Published: September 2015

TRP1 RI circle (now designated YARp1, yeast acentric ring plasmid 1) is a 1,453-base-pair artificial plasmid composed exclusively of Saccharomyces cerevisiae chromosomal DNA. It contains both the TRP1 gene and ARS1 (a DNA sequence that permits extrachromosomal maintenance of recombinant plasmids). This high-copy-number, relatively stable plasmid was shown to be organized into nucleosomes comparable to typical yeast chromatin, containing a possible maximum of nine nucleosomes per circle. Therefore, YARp1 can be used to examine the structure of chromatin of both a chromosomally derived replicator and a functional gene. By mapping regions of micrococcal nuclease cleavage in chromatin versus purified DNA, we located the positions of protected regions on the circle with reference to six unique restriction sites. Measurements made on patterns of early digestion products indicated that a region of approximately 300 base pairs in the vicinity of ARS1 was strongly resistant to micrococcal nuclease. The remainder of the plasmid appeared to be associated with five positioned nucleosomes and two nonnucleosomal, partially protected regions on the bulk of the molecules. After similar extents of digestion, naked DNA did not exhibit an equivalent pattern, although some hypersensitive cleavage sites matched sites found in the chromatin. These results are consistent with the interpretation that the protected domains are aligned with respect to a specific site or sites on the small circular chromatin.
ATTAAA as well as downstream sequences are required for RNA 3'-end formation in the E3 complex transcription unit of adenovirus.
Authors: Bhat B M , Wold W S
Published: September 2015

We mapped the location of the E3A RNA 3' end site in the E3 transcription unit of adenovirus 2. The procedure used was nuclease-gel analysis with 32P-labeled RNA probes. The poly(A) addition sites were microheterogeneous and were located approximately 17 to 29 nucleotides downstream from an ATTAAA sequence. To identify the sequences that make up the E3A RNA 3' end signal, we constructed five viable virus mutants with deletions in or near the E3A RNA 3' end site. The mutants were analyzed for E3A RNA 3' end formation in vivo. No effect was observed from a 47-base-pair (bp) deletion (dl716) or a 72-bp deletion (dl714) located 22 and 19 nucleotides, respectively, upstream of the ATTAAA. In contrast, E3A RNA 3' end formation was abolished by a 554-bp deletion (dl708) that removes both the ATTAAA and the poly(A) addition sites, a 124-bp deletion (dl713) that removes the ATTAAA but leaves the poly(A) addition sites, and a 65-bp deletion (dl719) that leaves the ATTAAA but removes the poly(A) addition sites. These results indicate that the ATTAAA, as well as downstream sequences, including the poly(A) addition sites, are required for E3A RNA 3' end formation.
Sequence-independent autoregulation of the adenovirus type 5 E1A transcription unit.
Authors: Hearing P , Shenk T
Published: September 2015

The adenovirus E1A gene is known to be autoregulated at the level of transcription. Autoregulation was found to be mediated by products of the E1A 13S mRNA, which induced a fivefold increase in E1A transcription rate. Deletion analysis suggested that the autoregulation did not require any specific sequence in the E1A transcriptional control region. This conclusion was reinforced by the demonstration that a cellular alpha-globin gene substituted for the E1A gene on the adenovirus chromosome was also positively regulated by E1A gene products.
The bovine papillomavirus distal "enhancer" is not cis essential for transformation or for plasmid maintenance.
Authors: Howley P M , Schenborn E T , Lund E , Byrne J C , Dahlberg J E
Published: September 2015

We constructed a mutant of bovine papillomavirus type 1 (BPV-1) DNA that lacked a transcriptional enhancer located 3' to the polyadenylation site of the early viral RNAs expressed in transformed cells. This mutant DNA, when separated from the procaryotic sequences, transforms mouse cells with an efficiency comparable to that of the full BPV-1 genome, and it exists as a stable multicopy plasmid in transformed cells. The BPV-1 distal enhancer suppresses the effects of a cis-inhibitory element in pML2 sequences but is not essential for the expression of the viral genes involved in cellular transformation or plasmid maintenance.