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M Carey

Publications and source records attributed to M Carey.

At least 109 records · Page 6Linked to original sources
PubMed1993

Comparing regions of the Epstein-Barr virus ZEBRA protein which function as transcriptional activating sequences in Saccharomyces cerevisiae and in B cells.

The ZEBRA protein activates expression of Epstein-Barr virus early-lytic-cycle genes in human B lymphocytes. Here it is shown that ZEBRA also behaves as a sequence-specific transcriptional activator in Saccharomyces cerevisiae. Deletional mutagenesis defined three regions of ZEBRA that participate in activation in S. cerevisiae. These regions are designated YI (amino acids [aa] 1 to 25), YII (aa 51 to 102), and YIII (aa 228 to 245). Two of the three regions of the native ZEBRA protein act together to mediate activation when assayed on ZEBRA binding sites. However, when fused to the DNA binding domain of GAL4 and assayed on GAL4 binding sites, regions YII and YIII were each sufficient to confer activation in S. cerevisiae. Regions of ZEBRA which affected activation in S. cerevisiae were also required in human B lymphocytes. The amino-terminal region of ZEBRA (aa 1 to 98) was required for activation both in S. cerevisiae and in human B cells; deletion of the carboxy-terminal 18 aa also significantly reduced activation in both cell types. Thus, the behavior of ZEBRA in human B cells and S. cerevisiae suggests that the protein contains universal activation motifs that interact with conserved components of the transcription machinery. However, certain deletion mutants of ZEBRA containing mutations in the N-terminal region exhibited discordant behaviors in S. cerevisiae and in B cells. For example, deletion of ZEBRA aa 26 to 51 impaired activation to a great extent in B cells but had little or no effect in S. cerevisiae. The discordant mutants may reflect interactions with a variable domain of a conserved component or unique interactions with specialized components of the basal transcription apparatus in different cells.

B-Lymphocytes↗
PubMed1993

The ZEBRA activation domain: modular organization and mechanism of action.

An RNA polymerase II activator often contains several regions that contribute to its potency, an organization ostensibly analogous to the modular architecture of promoters and enhancers. The regulatory significance of this parallel organization has not been systematically explored. We considered this problem by examining the activation domain of the Epstein-Barr virus transactivator ZEBRA. We performed our experiments in vitro so that the activator concentrations, stabilities, and affinities for DNA could be monitored. ZEBRA and various amino-terminal deletion derivatives, expressed in and purified from Escherichia coli, were assayed in a HeLa cell nuclear extract for the ability to activate model reporter templates bearing one, three, five, and seven upstream ZEBRA binding sites. Our data show that ZEBRA contains four modules that contribute to its potency in vitro. The modules operate interchangeably with promoter sites to determine the transcriptional response such that the loss of modules can be compensated for by increasing promoter sites. Potassium permanganate footprinting was used to show that transcriptional stimulation is a consequence of the activator's ability to promote preinitiation complex assembly. Kinetic measurements of transcription complex assembly in a reconstituted system indicate that ZEBRA promotes formation of a subcomplex requiring the TFIIA and TFIID fractions, where TFIIA acts as an antirepressor. We propose a model in which the concentration of DNA-bound activation modules in the vicinity of the gene initiates synergistic transcription complex assembly.

Cell Nucleus↗
PubMed1993

Fungal biotransformation of organophosphines.

1. The biotransformations of triphenyl phosphite and of triphenyl-, tri-n.-butyl-, diethylphenyl-, and ethylmethylphenyl-phosphines by the fungi Mortierella isabellina ATCC 42613, Helminthosporium species NRRL 4671, and Aspergillus foetidus ATCC 10254, have been examined. 2. The triaryl and tri-n.-butyl substrates underwent oxidation at phosphorus in low yield, a process attributed largely to auto-oxidation under the experimental conditions used. 3. The alkyl aryl phosphines were enzymically oxidized at phosphorus, but concurrent auto-oxidation also occurred. 4. Incubation of ethylmethylphenyl phosphine with M. isabellina gave the corresponding S(-) phosphine oxide in a 92% yield with an enantiomeric purity of 13%.

Aspergillus↗
PubMed1993

Necrotic pseudoxanthomatous nodules of the omentum and peritoneum: a peculiar reaction to endometriotic cyst contents.

BACKGROUND: Necrotic pseudoxanthomatous nodules of the ovary, omentum, and peritoneum are rare, and have been described as a late manifestation of endometriosis. CASE: A 39-year-old woman presented with acute abdominal pain and an abdominopelvic mass. Emergency laparotomy revealed a ruptured ovarian neoplasm with several liters of old blood in the abdominal cavity. Left adnexectomy was performed, and exploration of the abdominal cavity revealed no other lesions. Pathologic examination of the neoplasm showed a large endometrioma with an associated endometrioid adenocarcinoma. At surgery 7 weeks later, multiple pseudoxanthomatous peritoneal and omental nodules were found. CONCLUSION: This case suggests that pseudoxanthomatous nodules of the peritoneum and omentum may arise from a peculiar reaction to endometriotic cyst contents and clinically can mimic metastatic ovarian carcinoma.

Adenocarcinoma↗
PubMed1992

DNA recognition by GAL4: structure of a protein-DNA complex.

A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.

Amino Acid Sequence↗
PubMed1992

Polymerase II promoter activation: closed complex formation and ATP-driven start site opening.

Studies on bacterial RNA polymerases have divided the initiation pathway into three steps, namely (i) promoter binding to form the closed complex; (ii) DNA melting to form an open complex, and (iii) messenger RNA initiation. Potassium permanganate was used to detect DNA melting by mammalian RNA polymerase II in vitro. Closed complexes formed in a rate-limiting step that was stimulated by the activator GAL4-VP16. Adenosine triphosphate was then hydrolyzed to rapidly melt the DNA within the closed complex to form an open complex. Addition of nucleoside triphosphates resulted in the melted bubble moving away from the start site, completing initiation.

Adenosine Triphosphate↗
PubMed1992

A synergistic increase in potency of a multimerized VP16 transcriptional activation domain.

Transcriptional synergy in eucaryotes provides a means to control both the level and diversity of gene expression. The mechanism by which multiple activators elicit such effects is unknown. To address this problem we considered whether multimerizing an activation domain was equivalent to oligomerizing an activator's binding sites on DNA. Synthetic activators bearing one, two or four VP16 'core' activation domains, fused to the GAL4 DNA binding domain, were co-transfected into Cos-1 cells with CAT gene reporter templates containing one, two or five upstream GAL4 binding sites. Our results demonstrate that all of the activators elicit synergistic effects when comparing the amounts of transcription on multiple sites versus a single site. In contrast, the multimerized activation domains did not stimulate transcription significantly on a template bearing a single site; a synergistic increase in potency was, however, apparent on a template bearing two sites. Introducing the flexible lambda repressor linker region in between the activation domains increased the ability of activators bearing two or four VP16 domains to stimulate transcription from the single-site template. We discuss the mechanistic implications of this study on gene activation and synergy.

Binding Sites↗
PubMed1992

Relations among self-reported depressive symptoms in clinic-referred children versus adolescents.

Although there is consensus regarding the existence of childhood depression, disagreement remains as to whether symptoms are developmentally isomorphic. Previous studies focused on developmental differences in symptom levels; analyses of relations among symptoms may be more appropriate, however. Here both approaches were used to compare the Children's Depression Inventory responses from 1,030 clinic-referred children and adolescents. Four of nine symptom categories showed significant developmental differences in their correlations with total score. Externalizing behavior and guilt were more strongly related to depression in children than adolescents; affective symptoms and concerns about the future showed the reverse pattern. Results illustrate the importance of considering relations among symptoms as well as differences in symptom levels when evaluating theoretical claims about developmental differences in the nature of clinical syndromes.

Adolescent↗
PubMed1992

The acidic activator GAL4-AH can stimulate polymerase II transcription by promoting assembly of a closed complex requiring TFIID and TFIIA.

The assembly of activated RNA polymerase II (pol II) transcription complexes has been investigated by assaying whether pre-assembly of intermediate complexes reduces the extended time required for start-site melting. The results show that a closed complex requiring factors IIA, IID, and the acidic activator GAL4-AH forms in a rate-limiting step. This directs the templates into a productive assembly pathway. Factor TFIIB is then added rapidly, affording further protection against diversion into nonproductive pathways. These events are followed by a series of rapid steps in which the remaining general factors are assembled onto the template, which is then melted using the energy of ATP hydrolysis.

Base Sequence↗
PubMed1992

Transcriptional synergy by the Epstein-Barr virus transactivator ZEBRA.

ZEBRA is an Epstein-Barr virus (EBV) transcriptional activator that mediates a genetic switch between the latent and lytic states of the virus by binding to the promoters of genes involved in lytic DNA replication and activating their transcription. A computer survey revealed that 9 of 23 potential or known ZEBRA-responsive EBV genes contained two or more upstream binding sites; this suggested that ZEBRA can stimulate transcription synergistically. By using a series of synthetic promoters bearing one, two, three, five, and seven upstream recognition sites, we showed that ZEBRA activates transcription synergistically when templates bearing multiple sites were compared with a template bearing a single site. This phenomenon was observed in both uninfected and EBV-infected B-lymphoid cells and in vitro in a HeLa cell nuclear extract. DNase I footprinting was used to show that the synergy was not due to cooperative DNA binding mediated by direct contact between ZEBRA dimers. The in vitro experiments revealed two manifestations of synergy. One was seen when the levels of transcription observed with the same amounts of ZEBRA added to templates bearing different numbers of sites were compared. The other was observed when the two lowest concentrations of ZEBRA that stimulated measurable transcription from any given template were compared. On the basis of both the number of sites and the calculated Kd of ZEBRA for a single site, we estimated that the critical concentration of ZEBRA needed to elicit transcriptional synergy corresponds to a site occupancy of two or three bound ZEBRA dimers. Our results have biologic implications for both the EBV lytic cycle and other processes in which the concentration of an activator changes either temporally or spatially.

Base Sequence↗
PubMed1992

The sinking bullet.

We report a case of a missile injury to the brain with an unusual complication. The bullet migrated by its mere weight to a distant location through the brain parenchyma after it initially lodged in a superficial site. Instances of similar phenomena reported in the literature are reviewed.

Adolescent↗
PubMed1992

Disability in Western Highlands Province.

During late 1987 and early 1988 the Mount Hagen Handicapped Children's Centre (as it was then known) set out to determine if substantial numbers of persons with disabling conditions lived within 15km of Mount Hagen town. The investigation located 1408 persons, deemed by themselves, family members or neighbours to have a disability.

Adolescent↗
PubMed1991

Some reasons for the lack of progress in the treatment of acute myelogenous leukemia: a review of three consecutive trials of the treatment of poor prognosis patients.

The failure of three consecutive treatment protocols to significantly increase the complete remission rate for poor prognosis newly diagnosed patients with acute myelocytic leukemia led to a detailed investigation of the causes of treatment failure. In the majority of cases treatment failure was attributable to "clinical resistance" to therapy. Upon close examination two types of "clinical resistance" were discernible: the failure of chemotherapy to produce adequate cytotoxic effects ("classical" drug resistance), and treatment failure attributed to the rapid regrowth of leukemia cells subsequent to the substantial killing of leukemia cells by cytotoxic therapy ("biological" resistance). Each form of resistance accounted for one-half of the treatment failures.

Aged↗
PubMed1991

The objective evaluation of alternative treatment plans: II. Score functions.

A series of six patients with adenocarcinoma of the prostate, Stages A2, B1, or B2, were planned for treatment using a four-field box technique at 25 MV. Plans were prepared by three techniques: composite, mid-plane, and conformal. The dose distributions at the central plane and at two planes offset by +/- 2 cm were evaluated by means of score functions which quantify the magnitude of regret for target dose gradient, target over- and under-dose, non-target tissue overdose, and for overdose to the rectum, bladder, and femoral heads. The score functions are normalized to give values in the range from 10 (ideal) to zero (limit of acceptability), with negative values indicating unacceptable deviations from the prescribed dose limits. The scores for off-axis conformal plans were found to be essentially the same as for mid-plane plans on the central plane. However, mid-plane planning was shown to be totally inadequate for off-axis planes, where the average target gradient and underdose scores were reduced by 10 units. Composite planning resulted in adequate target coverage on all planes, but at the expense of unacceptable overdose to non-target tissue. The effect of reducing the posterior beam weight to half that of the other three beams was to reduce the target gradient score by 1.6 +/- 0.5 units and to increase the rectal score by 0.9 +/- 0.3 units.

Adenocarcinoma↗
PubMed1991

Variable resistance loadings in anaerobic power testing.

The aim of this study was to establish optimal test conditions for maximal anaerobic testing on a frictionloaded bicycle ergometer. Power and work outputs of 15 subjects during a 60-s all-out effort were tested under five conditions on a specially modified Monark ergometer. Test conditions were produced by altering the commencement resistance loading and either holding it constant (0.065C, 0.075C kg/kg. BW) for the entire duration of the 60-s work test or by reducing it by a specific amount to a more manageable load if pedalling rate dropped below 90 rpm (0.075R, 0.085R, 0.095R kg/kg. BW). Resistance and pedalling rate were continuously monitored with power outputs being determined every 100 ms, breath by breath oxygen consumption (VO2) was measured and capillary blood lactate (HLa) analyzed before and 5 min post each test condition. The test condition with the highest initial load (0.095R) produced the greatest peak power (PP). The reduced test procedure with the higher initial loads resulted in the greatest mean power (MP) and total work outputs (0.095R and 0.085R). Post-exercise blood HLa was not significantly different in any test condition. Peak power and MP correlated highly with both body weight (r = 0.77-0.85 and r = 0.75-0.83) and lean body weight (r = 0.84-0.88 and r = 0.90). During the last 30 s of the test, subjects reached between 90-100% of their measured maximal aerobic capacity (VO2max). The results of this study indicate that for a 60-s maximal all-out effort, a reduced test condition with a high initial resistance setting is required to produce the highest anaerobic test parameters.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗