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Biomedical subjects

S Fields

Publications and source records attributed to S Fields.

At least 109 records · Page 6Linked to original sources
PubMed1991

The two-hybrid system: a method to identify and clone genes for proteins that interact with a protein of interest.

We describe a method that detects proteins capable of interacting with a known protein and that results in the immediate availability of the cloned genes for these interacting proteins. Plasmids are constructed to encode two hybrid proteins. One hybrid consists of the DNA-binding domain of the yeast transcriptional activator protein GAL4 fused to the known protein; the other hybrid consists of the GAL4 activation domain fused to protein sequences encoded by a library of yeast genomic DNA fragments. Interaction between the known protein and a protein encoded by one of the library plasmids leads to transcriptional activation of a reporter gene containing a binding site for GAL4. We used this method with the yeast SIR4 protein, which is involved in the transcriptional repression of yeast mating type information. (i) We used the two-hybrid system to demonstrate that SIR4 can form homodimers. (ii) A small domain consisting of the C terminus of SIR4 was shown to be sufficient to mediate this interaction. (iii) We screened a library to detect hybrid proteins that could interact with the SIR4 C-terminal domain and identified SIR4 from this library. This approach could be readily extended to mammalian proteins by the construction of appropriate cDNA libraries in the activation domain plasmid.

Cloning, Molecular↗
PubMed1991

[CT-stereotactic determination of targets for thalamotomy for movement disorders].

The methodology of determining an invisible target by CT-stereotaxy, such as the ventrolateral nucleus of the thalamus, is described. This approach obviates the need for traditional ventriculographic determination of the third ventricle commissures and allows accurate measurement of third ventricular width. Morbidity and length of hospitalization are thereby reduced.

Adolescent↗
PubMed1991

Pheromone-dependent phosphorylation of the yeast STE12 protein correlates with transcriptional activation.

Haploid a and alpha cells of yeast respond to the pheromones alpha- and a-factor, respectively, by increasing the transcription of many genes whose products are essential for mating. The STE12 protein acts in this process by binding to the DNA sequence that mediates the increased transcription of pheromone-responsive genes. We show here that a hybrid protein containing STE12 fused to the DNA-binding domain of GAL4 can activate transcription of a reporter gene containing GAL4-binding sites but only after treatment of cells with pheromone. Thus, STE12 alone, when bound to DNA, is sufficient to mediate pheromone-induced transcription. By constructing hybrids of different STE12 regions with the GAL4 domain, we map the domain of STE12 necessary for this activation to the central third of the protein. Upon alpha-factor treatment, the hybrid of GAL4 with the complete STE12 sequence is rapidly phosphorylated, with kinetics consistent with the observed transcriptional induction of pheromone-responsive genes. The domain of STE12 necessary for this phosphorylation correlates with that involved in transcriptional activation. We propose that induction of pheromone-responsive genes is mediated by phosphorylation of STE12 to alter its activation function but not its DNA-binding ability.

Binding Sites↗
PubMed1991

Properties of the DNA-binding domain of the Saccharomyces cerevisiae STE12 protein.

The STE12 protein of the yeast Saccharomyces cerevisiae binds to the pheromone response element (PRE) present in the upstream region of genes whose transcription is induced by pheromone. Using DNase I footprinting assays with bacterially made STE12 fragments, we localized the DNA-binding domain to 164 amino acids near the amino terminus. Footprinting of oligonucleotide-derived sequences containing one PRE, or two PREs in head-to-tail or tail-to-tail orientation, showed that the N-terminal 215 amino acids of STE12 has similar binding affinity to either of the dimer sites and a binding affinity 5- to 10-fold lower for the monomer site. This binding cooperativity was also evident on a fragment from the MFA2 gene, which encodes the a-factor pheromone. On this fragment, the 215-amino-acid STE12 fragment protected both a consensus PRE as well as a degenerate PRE containing an additional residue. Mutation of the degenerate site led to a 5- to 10-fold decrease in binding; mutation of the consensus site led to a 25-fold decrease in binding. The ability of PREs to function as pheromone-inducible upstream activation sequences in yeast correlated with their ability to bind the STE12 domain in vitro. The sequence of the STE12 DNA-binding domain contains similarities to the homeodomain, although it is highly diverged from other known examples of this motif. Moreover, the alignment between STE12 and the homeodomain postulates loops after both the putative helix 1 and helix 2 of the STE12 sequence.

Amino Acid Sequence↗
PubMed1990

Presence of a potent transcription activating sequence in the p53 protein.

The p53 gene is frequently mutated in a wide variety of human cancers. However, the role of the wild-type p53 gene in growth control is not known. Hybrid proteins that contain the DNA binding domain of yeast GAL4 and portions of p53 have been used to show that the p53 protein contains a transcription-activating sequence that functions in both yeast and mammalian cells. The NH2-terminal 73 residues of p53 activated transcription in mammalian cells as efficiently as the herpes virus protein VP16, which contains one of the strongest known activation domains. Combined with previous data that showed p53 is localized to the nucleus and can bind to DNA, these results support the idea that one function of p53 is to activate the transcription of genes that suppress cell proliferation.

Animals↗
PubMed1990

Architectural organization in the interphase nucleus of the protozoan Trypanosoma brucei: location of telomeres and mini-chromosomes.

We studied the spatial organization of chromatin in the interphase G1, S and G2 nucleus of the protozoan Trypanosoma brucei, applying in situ hybridization with conventional fluorescence and confocal scanning optical microscopy. The majority of the trypanosome telomere GGGTTA repeats from different chromosomes were found clustered together, either extending in a network through the nuclear interior or localized at the nuclear periphery. The population of one hundred mini-chromosomes was often asymmetrically located: either clustered in a narrow band in close association with the nuclear envelope or distributed into several clusters that segregated into roughly one half of the nucleus. The nuclear organization may undergo modifications during the cell cycle and development. We conclude that non-random spatial positioning of DNA exists in the nucleus of this protozoan. Finding a high level of structural organization in the interphase nucleus of T.brucei is an important first step towards understanding chromosome structure and functioning and its role in the control of gene expression.

Animals↗
PubMed1990

Pheromone response in yeast.

In response to peptide pheromones, yeast cells prepare themselves for mating; changes include arrest of the cell cycle and induction of transcription. Proteins involved in this signal transduction pathway include the pheromone receptors, subunits of a G protein, protein kinases and DNA-binding proteins. Understanding of this pathway has been facilitated by yeast genetics, which has allowed the genes encoding all of these proteins to be identified and characterized.

Cell Cycle↗
PubMed1990

Overproduction of the yeast STE12 protein leads to constitutive transcriptional induction.

Haploid a and alpha cells of the yeast Saccharomyces cerevisiae respond to the pheromones alpha- and a-factor, respectively, by increasing transcription of inducible genes, arresting cell division, and forming cell-surface projections. These responses are dependent on the activity of several genes, including STE12, whose product binds to the pheromone response element located within the regulatory DNA sequences of inducible genes. We assayed the effects of overproducing the STE12 protein in both STE+ cells, as well as ste2, ste7, and ste11 mutant cells. We find that overproduction leads to increased transcription of pheromone-inducible genes and is able to suppress the mating defect of the ste mutants. These results suggest that one effect of pheromone treatment may be to increase the ability of the STE12 protein to activate transcription. In addition, we observed that cells cannot tolerate very high levels of the STE12 protein, and many arrest in G1 with a large size and morphological changes. Thus, constitutively high-level transcription of pheromone-inducible genes causes cells to display some features similar to treatment with pheromone.

Fungal Proteins↗
PubMed1990

Clinical trial of remote radiologist services for a military installation.

It is often difficult to provide continuity of professional radiological services at U.S. military facilities located at a distance from major metropolitan areas. To address this problem, we have designed and evaluated a program to send x-ray films from a U.S. Army hospital to a remote site for expert radiological interpretation. Nine hundred and forty-two x-ray studies were sent by U.S. Express Mail from the Radiology Department of a U.S. Army Hospital over a 2-month period. The typed reports were sent back to the hospital through the use of a computer-to-computer telephone link within 72 hours of film transport. No studies were lost and security of all studies was maintained. This program offers a cost-effective, otherwise unavailable means to provide high quality professional radiological services to U.S. military facilities both inside the United States and in foreign countries.

Computer Communication Networks↗
PubMed1989

A novel genetic system to detect protein-protein interactions.

Protein-protein interactions between two proteins have generally been studied using biochemical techniques such as crosslinking, co-immunoprecipitation and co-fractionation by chromatography. We have generated a novel genetic system to study these interactions by taking advantage of the properties of the GAL4 protein of the yeast Saccharomyces cerevisiae. This protein is a transcriptional activator required for the expression of genes encoding enzymes of galactose utilization. It consists of two separable and functionally essential domains: an N-terminal domain which binds to specific DNA sequences (UASG); and a C-terminal domain containing acidic regions, which is necessary to activate transcription. We have generated a system of two hybrid proteins containing parts of GAL4: the GAL4 DNA-binding domain fused to a protein 'X' and a GAL4 activating region fused to a protein 'Y'. If X and Y can form a protein-protein complex and reconstitute proximity of the GAL4 domains, transcription of a gene regulated by UASG occurs. We have tested this system using two yeast proteins that are known to interact--SNF1 and SNF4. High transcriptional activity is obtained only when both hybrids are present in a cell. This system may be applicable as a general method to identify proteins that interact with a known protein by the use of a simple galactose selection.

DNA Restriction Enzymes↗
PubMed1989

[Limb length discrepancy determined by computerized tomography and radiography].

A reliable radiographic limb length measurement is necessary for evaluating and treating discrepancies in limb length, a most difficult problem in pediatric orthopedics. The accuracy and efficiency of conventional orthoroentgenography was compared to that of computerized tomographic (CT) scanography in 17 patients. There was no significant difference in measurements between the techniques. CT scanography with a 66% reduction in radiation dose is now our routine examination for leg length discrepancy.

Evaluation Studies as Topic↗
PubMed1989

Intra-abdominal lymphangioma.

The historical background, aetiology, clinical features, radiographic findings and treatment of abdominal lymphangiomas are reviewed. The condition may give rise to the acute surgical abdomen.

Humans↗
PubMed1989

The yeast STE12 protein binds to the DNA sequence mediating pheromone induction.

The STE12 gene product of Saccharomyces cerevisiae is required for the transcription of two sets of cell-type-specific genes: the a-specific genes (active only in a cells) and the alpha-specific genes (active only in alpha cells). We show that radiolabeled STE12 protein, prepared by in vitro transcription and translation, is capable of forming complexes with unlabeled DNA fragments from two a-specific genes. Wild-type yeast, but not a ste12 mutant, produce a factor that forms complexes with labeled DNA from these same two genes. We use assays with yeast extracts to localize the binding site for the STE12-dependent activity. This site corresponds to the sequence identified as the pheromone induction element, which is responsible for increased transcription of genes when cells are exposed to alpha-factor or a-factor. Thus the STE12 protein may be an ultimate effector in the signal transduction pathway triggered by pheromone.

Amino Acid Sequence↗
PubMed1989

Solitary nonparasitic cyst of the liver: a rare cause of abdominal distension.

Solitary nonparasitic liver cysts are clinically rare, and fewer than 900 patients have been documented over the years. A case is described of a female patient with a large solitary cyst containing 7 L of fluid, which was treated successfully by unroofing. The literature relating to the clinical features, incidence, pathogenesis, histopathology, diagnostic imaging, and surgical management of such cysts is reviewed.

Abdomen↗