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Biomedical subjects

W Schaffner

Publications and source records attributed to W Schaffner.

At least 19 recordsLinked to original sources
PubMed1992

POU-specific domain of Oct-2 factor confers 'octamer' motif DNA binding specificity on heterologous Antennapedia homeodomain.

The bipartite DNA binding domain of the POU family of transcription factors contains a 'POU-specific' domain unique to this class of factors and a 'POU homeodomain' homologous to other homeodomains. We compared DNA binding of the Oct-2 factor POU domain and the Antennapedia (Antp) homeodomain with a chimeric Oct-2/Antp protein in which the distantly related Antp homeodomain was substituted for the Oct-2 POU homeodomain. The Oct-2/Antp chimeric protein bound both the octamer and the Antp sites efficiently, indicating that DNA binding specificity is contributed by both components of the POU domain.

Antennapedia Homeodomain Protein↗
PubMed1992

Different activation domains stimulate transcription from remote ('enhancer') and proximal ('promoter') positions.

We reported previously that the lymphocyte-derived octamer transcription factor 2A (Oct-2A or OTF-2A) activated both natural immunoglobulin promoters and synthetic promoters which contain the 'octamer' site, but was unable by itself to stimulate transcription from a remote enhancer position. Here we examine a larger set of transcription factors with respect to their proximal versus remote activation. Since a transcription factor may contain more than one activation domain, we have chosen to study the potential of individual activation domains in the context of fusion proteins that contain the DNA binding domain of GALA. We have identified at least two distinct functional classes of transcriptional activation domains. 'Proximal' activation domains, exemplified by glutamine-rich domains of Oct-1, Oct-2A and Sp1, stimulate transcription only from a position close to the TATA box, usually in response to a remote enhancer. 'General' activation domains, derived from VP16, GAL4, p65 (NF-chi B), TFE3, ITF-1 and ITF-2, can activate transcription from remote as well as proximal positions. These domains contain many acidic amino acids and/or other features such as clusters of serine and threonine. The proline-rich activation domains of AP-2 and CTF/NF1 may represent a third class with considerable promoter activity and low but significant enhancer activity. Furthermore, activation domains of both the acidic and glutamine-rich types seem to have a modular structure, since duplicated subdomains can substitute for the entire domain.

Amino Acid Sequence↗
PubMed1992

Rubella among the Amish: resurgent disease in a highly susceptible community.

Although the Amish make up less than 0.05% of the United States population, nearly all rubella reported in the United States in 1991 occurred in this population. In early 1991 a large rubella outbreak in a Tennessee Amish community that had experienced no rubella for 17 years afforded an opportunity to describe the epidemiology of rubella in this unique population. Structured interviews were conducted with 54 Amish families. Of 383 persons in the sample 85 (22%) had rubella. Illnesses were mild; 16% of cases lacked fever and 20% of cases reported no symptoms except rash. Children < 17 years of age were 7 times more likely than older individuals to be affected (77 of 214 vs. 8 of 165). All pregnant women in the community were > 20 years of age; none developed rubella. No congenital rubella syndrome was recognized. Although rubella is increasingly a disease of adolescents and young adults, in this outbreak, rubella was again a childhood disease. Illness in this community-based investigation was mild; rubella may be difficult to diagnose and report. Immunity after remote natural infection was durable since the community's last outbreak. Pregnant women probably were protected by the age distribution of immunity; this age distribution may not occur in other Amish populations. If preventable morbidity from rubella and other vaccine preventable diseases is to be avoided in this group, increased attention should be directed to encouraging vaccinations among Amish persons.

Disease Outbreaks↗
PubMed1992

Focused microbiologic surveillance by specific hospital unit as a sensitive means of defining antimicrobial resistance problems.

An annual summary of susceptibility patterns for the predominant clinical isolates from hospitalized patients can be of considerable assistance in selecting antimicrobial agents for sepsis of unclear etiology, as well as for guiding empiric therapy for other serious infections. Yearly summaries of the susceptibility patterns of the predominant clinical isolates from all patients hospitalized at Vanderbilt University Hospital (VUH) from July 1987 through June 1991 revealed only minor differences over time in susceptibility patterns. However, the clinical impression of physicians treating patients in various intensive care units (ICUs) was that there were serious resistance problems in some units. To better define the prevalence of clinical isolates and their susceptibility patterns for patients within ICUs at VUH, we utilized a "focused microbiologic surveillance" technique that addressed each unit separately. Both the predominant clinical isolates and their susceptibility patterns were determined and compared with those from the hospital as a whole. Because susceptibility patterns of clinical isolates by site of infection within these units were considered important, we also reviewed the summaries of susceptibility patterns for blood, sputum, and urine isolates from patients in ICUs and compared these with the summaries from each ICU and from the hospital. No major resistance problems were identified on a hospital-wide basis. In contrast, focused microbiologic surveillance by specific hospital ICU revealed important differences in the prevalence of pathogens among units and at different times. In 1987, Pseudomonas aeruginosa was the single most common Gram-negative organism isolated in the neonatal unit, while Acinetobacter spp. were rarely isolated. By 1991, this trend was completely reversed.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Therapeutic and epidemiologic recommendations to reduce the spread of type-I beta-lactamase resistance.

The objectives of this United States Consensus Panel meeting were to evaluate the effectiveness of current surveillance systems for the detection of bacterial resistance as well as to formulate recommendations that can assist hospitals in determining actions that should be taken when a resistance problem is detected. These recommendations may be particularly helpful in controlling the emergence and spread of type-I beta-lactamase resistance. Numerous case reports of antimicrobial resistance among Enterobacter species, Pseudomonas aeruginosa, and other Gram-negative nosocomial pathogens known to produce type-I beta-lactamases have appeared in the literature since the introduction of the newer "third-generation" cephalosporins. The widespread use of these newer antimicrobial agents, often selected as standard therapy for serious hospital-acquired infections, has been associated with a corresponding increase in resistance to them. The failure of hospitalwide surveillance methods to describe the scope of this problem, especially among the most critically ill patients, may have resulted in a false sense of security among some infectious disease specialists and clinicians prescribing these antimicrobials as empiric therapy. High-level resistance in individual hospital units may be masked in hospitalwide antibiograms. A variety of conclusions and recommendations were formulated based on the collective experiences of the Consensus Panel members. Microbiology laboratories must make it a high priority to identify markers that will assist in rapidly identifying resistant organisms. Cooperative efforts are needed among users of commercial and automated microbiology test instruments to standardize results and to improve quality control, thereby making the data more directly comparable between laboratories.(ABSTRACT TRUNCATED AT 250 WORDS)

Anti-Bacterial Agents↗
PubMed1991

Expression in mammalian cells of a cloned gene encoding murine DNA methyltransferase.

Mammalian DNA cytosine-5-methyltransferase (MTase, EC 2.1.1.37) is an essential component for establishing and maintaining cell-type specific methylation patterns in the genome. The cDNA for the murine enzyme was previously cloned in segments. We have reconstructed the entire gene, encoding a protein of 1517 amino acids, from a set of overlapping cDNA clones. We report the assembly of two expression constructs in bacterial/mammalian shuttle vectors. Transcription in the first construct (pEMT) is driven by the cytomegalovirus enhancer/promoter and encodes a fusion protein with 15 additional aa at the N terminus, while the second construct (pJMT) is driven by the simian virus 40 early promoter/enhancer upstream from the natural ATG codon. Immunofluorescence microscopy and immunoblot analysis have shown that both constructs direct the synthesis of MTase in COS-1 cells. Enzyme activity in whole-cell lysates of transfected COS-1 cells transfected with pEMT and pJMT are on average tenfold and fivefold higher than in controls, respectively. The specific activities of the recombinant and endogenous mouse-cell enzyme are similar. These expression constructs will be of use in studies of DNA methylation in mammals.

Animals↗
PubMed1991

Upstream box/TATA box order is the major determinant of the direction of transcription.

Mammalian gene promoters for transcription by RNA polymerase II are typically organized in the following order: upstream sequence motif(s)/TATA box/initiation site. Here we report studies in which the order, orientation and DNA sequences of these three elements are varied to determine how these affect polarity of transcription. We have constructed promoters with an 'octamer' upstream sequence ATTTGCAT (or its complement ATGCAAAT) in combination with several different TATA boxes and initiation (cap) sites, and tested these promoters in transfection experiments with cultured cells. TATA boxes derived from the adenovirus major late promoter (TATAAAA), immunoglobulin kappa light chain (TTATATA) and heavy chain (TAAATATA) promoter functioned equally well or even better when inverted. Only the beta-globin TATA box (CATAAAA) was poorly active when inverted. In addition, a symmetrical TATA box (TATATATA) derived from a casein gene was very active. Our results suggest that the asymmetry of most TATA boxes (consensus TATAAAA) is not a primary determinant of the polarity of transcription. We also found that the initiation (cap) site, which usually consists of an adenine embedded in a pyrimidine-rich region (PyPyCAPyPyPyPyPy), was permissive towards sequence alterations; even a randomly composed sequence worked well. However, an inverted, hence purine-rich, cap site reduced transcript levels to 1/7th, as did an oligo G sequence. Irrespective of the presence of a cap site, the configuration: 'TATA box/octamer' yielded a strong leftward, rather than rightward transcription. From this, we conclude that the polarity of transcription is primarily determined by the linear order of an upstream sequence relative to a TATA box, rather than by the individual orientations of either of these two elements.

Adenoviridae↗
PubMed1991

Salmonella enteritidis gastroenteritis transmitted by intact chicken eggs.

OBJECTIVE: To determine the source and to describe the clinical importance of a large outbreak of Salmonella enteritidis gastroenteritis in Tennessee, which is outside the geographic focus of the S. enteritidis pandemic. DESIGN: A case-control study and tracing of the source eggs. SETTING: A Tennessee community and a large layer farm in Indiana. PATIENTS: Case patients ate at the implicated restaurant and subsequently developed S. enteritidis gastroenteritis; controls ate with the case patients, but did not develop gastroenteritis. MEASUREMENTS: Eighty-one case patients were identified; 73 (90%) had eaten egg-containing sauces at a local restaurant on a given evening. The eggs were traced to their farm of origin in Indiana. The farm was inspected 5 weeks after the outbreak. MAIN RESULTS: Of 24 patients with culture-proved cases, 11 were hospitalized. Hollandaise and bernaise sauces prepared with intact, extra-large, grade-A eggs were strongly associated with illness (P less than 0.001). Salmonella enteritidis was isolated from specimens collected from chickens and the farm. Antimicrobial susceptibility patterns, phage typing, and plasmid profiles of isolates from the farm and from patients were indistinguishable. CONCLUSIONS: Salmonella enteritidis infection is a large and growing public health problem that is spreading beyond the northeastern United States. This study shows a direct link between infected poultry flocks and an outbreak of human illness.

Adult↗
PubMed1991

Thionein (apometallothionein) can modulate DNA binding and transcription activation by zinc finger containing factor Sp1.

A number of transcription factors contain so-called zinc finger domains for the interaction with their cognate DNA sequence. It has been shown that removal of the zinc ions complexed in these zinc fingers abrogates DNA binding and transcription activation. Therefore we wanted to test the hypothesis that the activity of transcription factors could be regulated by physiological chelators of zinc. A prominent candidate for such a chelator is the Cys-rich protein thionein (apometallothionein) that is inducible by heavy metal loads, and by other environmental stimuli. Here we show with DNA binding and in vitro transcription assays that thionein indeed can inactivate the zinc finger-containing Sp1 in a reversible manner. By contrast, transcription factor Oct-1, which binds DNA via a homeo-domain, i.e. a helix-turn-helix motif not involving zinc ions, is refractory to thionein action. We propose that modulation of intracellular thionein concentration is used for the coordinated regulation of a large subset of genes whose transcription depends on zinc finger proteins.

Animals↗
PubMed1991

Promoters with the octamer DNA motif (ATGCAAAT) can be ubiquitous or cell type-specific depending on binding affinity of the octamer site and Oct-factor concentration.

Immunoglobulin (Ig) gene promoters contain the octamer sequence motif ATGCAAAT which is recognized by cellular transcription factors (Oct factors). Besides the ubiquitous Oct-1 factor, there is also a group of related factors (Oct-2 factors) encoded by a separate gene. The Oct-2 gene is regulated in a cell-type specific manner, and the protein is present in large amounts in B lymphocytes. We have previously shown that simple composite promoters of an octamer/TATA box type are poorly active in non-B cells but are strongly responsive to ectopic expression of Oct-2A factor, a major representative of the lymphocyte Oct-2 factors. In the present study we have tested the activity of a number of composite promoters and natural Ig promoters, and their response to Oct-1 and Oct-2 factors. Unexpectedly, we find that octamer/TATA promoters with a high affinity octamer site direct ubiquitous expression. By contrast, promoter constructions that behave in a B cell-specific manner tend to have a weak octamer binding site. These promoters are responsive to ectopic expression of additional Oct-factor, irrespective of whether it is Oct-1 or Oct-2. Using natural Ig promoters rather than composite promoters, we find that an IgH promoter is well transcribed in non-B cells via the ubiquitous Oct-1 factor, while Ig kappa and Ig lambda light chain promoters require additional Oct factor for maximal expression. It seems therefore likely that during B cell differentiation, Ig heavy chain promoters can be activated by Oct-1, before the appearance of Oct-2 factors. Oct-2 factors then would serve to boost the expression from Ig light chain promoters, which are known to be activated only after successful heavy chain gene rearrangement.

Autoradiography↗
PubMed1991

Both Oct-1 and Oct-2A contain domains which can activate the ubiquitously expressed U2 snRNA genes.

The U2 snRNA genes, which are transcribed by RNA polymerase II at high levels in all tissues examined, require both a distal and a proximal sequence element for efficient expression. The distal sequence element which has many properties in common with transcriptional enhancers contains, in addition to Sp1 binding sites, an octamer binding site which mediates activation through interactions with the ubiquitous transcription factor Oct-1. In the present study we have attempted to answer the question whether Oct-1 contains a unique activating domain which is required for activation of snRNA genes or whether ubiquitously expressed and lymphoid specific octamer binding factors both have the capacity to activate snRNA transcription. Our results show that in the presence of Oct-1, overexpression of Oct-2A in HeLa or COS1 cells neither inhibits nor stimulates transcription of U2 constructions which contain octamer binding sites with or without an adjacent Sp1 binding site. Moreover, an Oct-2A--GAL4 fusion protein in which the DNA binding domain of Oct-2A was substituted for by the one of the yeast transcription activator GAL4 activates transcription of a human U2 snRNA gene in which the octamer binding site was replaced by a GAL4 binding site. From the results it is concluded that both Oct-1 and Oct-2A contain domains which can activate the ubiquitously expressed U2 snRNA genes.

Base Sequence↗
PubMed1991

Surgeons with HIV infection: the risk to patients.

The risk of human immunodeficiency virus (HIV) transmission from an infected surgeon to a patient is probably very low. In separate studies, the patients of four surgeons with acquired immunodeficiency syndrome (AIDS) have been followed up; no evidence of transmission was found. However, a recent report suggests that a dentist with AIDS may have transmitted HIV infection to a patient during the extraction of two molar teeth. Whether HIV-infected health care workers who perform invasive procedures ought to be obliged to discontinue or modify their practice continues to be debated.

Adult↗
PubMed1991

Every enhancer works with every promoter for all the combinations tested: could new regulatory pathways evolve by enhancer shuffling?

The promoters and enhancers of cell type-specific genes are often conserved in evolution, and hence one might expect that a given enhancer has evolved to work best with its own promoter. While this expectation may be realized in some cases, we have not found evidence for it. A total of 27 combinations of different promoters and enhancers were tested by transfection into cultured cells. We found that the relative efficiency of the enhancers is approximately the same, irrespective of the type of promoter used, i.e., there was no strong preference for any given enhancer/promoter combination. Notably, we do not see particularly strong transcription when the immunoglobulin kappa enhancer (or the immunoglobulin heavy chain enhancer) is used to activate a kappa gene promoter. We propose that a generally permissive enhancer/promoter interaction is of evolutionary benefit for higher eukaryotes: by enhancer shuffling, genes could be easily brought under a new type of inducibility/cell type specificity.

B-Lymphocytes↗
PubMed1991

Risk of seizures after measles-mumps-rubella immunization.

To evaluate the risks of seizures and other neurologic events following measles-mumps-rubella (MMR) or measles-rubella (MR) immunization, a retrospective cohort study was conducted among 18,364 Tennessee children enrolled in Medicaid who received MMR or MR immunizations in their first 3 years of life. One hundred children had seizures at some time between immunization and 36 months; there were no encephalopathies during this period. Four children had febrile seizures in the 7 through 14 days following MMR or MR immunization compared with 72 in the interval 30 or more days following MMR or MR immunization yielding a relative risk (95% confidence interval) of 2.1 (0.7 to 6.4). Although not statistically significant, this increase in febrile seizures in the 7- through 14-day interval following MMR immunization is coincident with the occurrence of fever following MMR immunization and is consistent with reports of other investigators.

Brain Diseases↗