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C S Parker

Publications and source records attributed to C S Parker.

At least 19 recordsLinked to original sources
PubMed2001

Developmental regulation of the heat shock response by nuclear transport factor karyopherin-alpha3.

During early stages of Drosophila development the heat-shock response cannot be induced. It is reasoned that the adverse effects on cell cycle and cell growth brought about by Hsp70 induction must outweigh the beneficial aspects of Hsp70 induction in the early embryo. Although the Drosophila heat shock transcription factor (dHSF) is abundant in the early embryo it does not enter the nucleus in response to heat shock. In older embryos and in cultured cells the factor is localized within the nucleus in an apparent trimeric structure that binds DNA with high affinity. The domain responsible for nuclear localization upon stress resides between residues 390 and 420 of the dHSF. Using that domain as bait in a yeast two-hybrid system we now report the identification and cloning of a Drosophila nuclear transport protein karyopherin-alpha3 (dKap-alpha3). Biochemical methods demonstrate that the dKap-alpha3 protein binds specifically to the dHSF's nuclear localization sequence (NLS). Furthermore, the dKap-alpha3 protein does not associate with NLSs that contain point mutations, which are not transported in vivo. Nuclear docking studies also demonstrate specific nuclear targeting of the NLS substrate by dKap-alpha3. Consistant with previous studies demonstrating that early Drosophila embryos are refractory to heat shock as a result of dHSF nuclear exclusion, we demonstrate that the early embryo is deficient in dKap-alpha3 protein through cycle 12. From cycle 13 onward the transport factor is present and the dHSF is localized within the nucleus thus allowing the embryo to respond to heat shock.

Active Transport, Cell Nucleus↗
PubMed1999

Site-specific inhibition of transcription factor binding to DNA by a metallointercalator.

The metallointercalator Lambda-1-Rh(MGP)2phi5+ binds tightly and specifically to the site 5'-CATATG-3' in the major groove of double helical DNA by a combination of direct readout and shape selection. To examine competitive interactions between this small metal complex and a DNA-binding transcription factor, the preferred binding site for Lambda-1-Rh(MGP)2phi5+ was engineered into the AP-1 recognition element (ARE) of the major-groove binding bZIP transcription factor yAP-1, the yeast analogue of mammalian AP-1. Binding experiments confirmed that the modified ARE retained normal yAP-1 binding affinity. Photocleavage experiments demonstrated that the modified ARE contained a high-affinity binding site for Lambda-1-Rh(MGP)2phi5+, whereas the native ARE showed no interaction. Competition experiments using gel shift mobility assays demonstrated that Lambda-1-Rh(MGP)2phi5+ at 120 nM competes 50% of yAP-1 binding to the 5'-CATATG-3' containing oligonucleotide. In contrast, competitive disruption of protein binding to the native ARE requires 3 microM Lambda-1-Rh(MGP)2phi5+. Metallointercalator derivatives, including geometric isomers of Lambda-1-Rh(MGP)2phi5+, show no specific binding to the target site and show no inhibition of yAP-1/DNA complexes at concentrations as high as 20 microM. Thus, metallointercalators can be tuned to show selectivity for major groove sites on DNA comparable to transcription factors and indeed can inhibit transcription factor binding site selectively.

Binding Sites↗
PubMed1997

Nuclear entry, oligomerization, and DNA binding of the Drosophila heat shock transcription factor are regulated by a unique nuclear localization sequence.

In normally growing Drosophila cultured cells the Drosophila heat shock transcription factor (dHSF) is localized in the cytosol and translocates into the nucleus after heat shock. In the cytosol of nonshocked cells, the dHSF is present as a monomer that cannot bind DNA. Upon stress, the dHSF enters the nucleus where it is observed to be a trimer. A novel nuclear localization sequence (NLS) in the dHSF was found to be responsible for stress-dependent nuclear entry. Deletion of the NLS prevents nuclear entry, as expected, yet surprisingly also allows constitutive oligomerization and DNA binding in the cytosol. Further analysis of the NLS by mutagenesis suggests that the two functions of nuclear entry and oligomerization are separable in that distinct residues present in the NLS are responsible for each. Mutations in certain basic residues completely block nuclear entry, as expected for a constitutive NLS. In addition, two residues were found in the NLS that, when altered, allowed constitutive nuclear entry of dHSF independent of stress. These residues may interact with a putative cellular component or possibly other domains of the HSF to prevent nuclear entry in normally growing cells. The NLS can also function autonomously to target a beta-galactosidase fusion protein into the nucleus in a heat shock-dependent fashion.

Amino Acid Sequence↗
PubMed1997

Charting by exception.

Documentation takes up a significant portion of a home care nurse's daily workload. As cost containment measures reduce the amount of time nurses can spend with each patient, they need to find ways to make documentation more efficient to avoid compromising patient care. One agency found that charting by exception reduces documentation time so that nurses can devote more hours to hands-on care.

Community Health Nursing↗
PubMed1996

Allied health faculty attitudes toward rural clinical practice.

Research findings of allied health practitioners' attitudes toward rural practice are limited. The purpose of this study was to identify attitudes of faculty members in a school of allied health toward rural vs. urban living, clinical education, and practice. A survey consisting of demographic and attitudinal questions was mailed to 233 faculty representing five professions. The response rate was 63.5%. The majority viewed rural living as having both positive and negative aspects. Placement of clinical students in rural areas was seen as enhancing rural recruitment. Rural professional issues were viewed as mixed with the most positive aspect being greater intellectual challenge. There were a few significant attitude differences by gender, age, years of experience, profession, hometown location, and practice location site. The findings of this study generally support previous research and contribute additional knowledge regarding attitudes toward rural practice. Further studies of allied health professionals appear warranted.

Allied Health Personnel↗
PubMed1995

The essential yeast NLT1 gene encodes the 64 kDa glycoprotein subunit of the oligosaccharyl transferase.

The yeast oligosaccharyl transferase catalyzes the glycosylation of asparagine residues in secreted, vesicular, and membrane proteins. A complex of at least four membrane-bound polypeptides is responsible for oligosaccharyl transferase activity. Amino acid sequences from the 64 kDa glycoprotein subunit of the complex were used to clone the essential NLT1 (N-linked oligosaccharyl transferase) gene. The Nlt1p gene product is a processed, multiply glycosylated type I membrane protein; it has an extensive amino-terminal soluble domain, a potential hydrophobic transmembrane domain, and a short carboxy-terminal soluble domain. The Nlt1p is significantly similar than the mammalian ribophorin I, a component of the mammalian oligosaccharyl transferase complex, and the enzyme is conserved throughout eukaryotic evolution.

Amino Acid Sequence↗
PubMed1994

Transcriptional activation mediated by the yeast AP-1 protein is required for normal cadmium tolerance.

The yeast YAP1 gene encodes a transcriptional regulatory protein that utilizes a basic region-leucine zipper (bZip) DNA-binding domain to recognize its cognate DNA element. A synthetic reporter gene containing a SV40 AP-1 response element (ARE) cloned upstream of a TRP5 promoter-lacZ gene fusion shows yAP-1-dependent transactivation in vivo. Recent work has shown that changes in the gene dosage of this factor can dramatically alter the ability of a cell to tolerate a host of toxic agents including cadmium, cycloheximide, and sulfometuron methyl. We have focused on the YAP1-dependent cadmium resistance as cells that lack a functional YAP1 gene are hypersensitive to this metal. Deletion mapping experiments define two domains in the carboxyl-terminal region of the yAP-1 protein that are required for normal cadmium tolerance and ARE-TRP5-lacZ expression. Single amino acid substitutions in the bZip domain of yAP-1 indicate that this region is required for normal DNA binding and in vivo function of the protein. Replacement of a non-canonical asparagine with leucine in the yAP-1 leucine zipper leads to production of a defective protein. A substitution mutation in the basic domain converts this mutant protein into a dominant negative factor. The ability of yAP-1 to act as a positive regulator of transcription is required for its biological action.

Base Sequence↗
PubMed1994

The yeast and mammalian Ras pathways control transcription of heat shock genes independently of heat shock transcription factor.

Yeast strains in which the Ras-cyclic AMP (cAMP) pathway is constitutively active are sensitive to heat shock, whereas mutants in which the activity of this pathway is low are hyperresistant to heat shock. To determine the molecular basis for these differences, we examined the transcriptional induction of heat shock genes in various yeast strains. Activation of heat shock genes was attenuated in the strains in which the Ras-cAMP pathway is constitutively active. In contrast, in a strain deficient in cAMP production, several heat shock genes were induced by removal of cAMP from the medium. These results indicate that the Ras-cAMP pathway affects the induction of heat shock genes. In all of the mutants, heat shock transcription factor expression and activity were identical to those in wild-type cells. The response to heat shock in Ha-ras-transformed rat fibroblasts was also studied. While no induction of Hsp68 was observed in Ha-ras-transformed cells, proper regulation of heat shock transcription factor was found. Therefore, in mammals, as in Saccharomyces cerevisiae, the Ras pathway controls the transcription of heat shock genes via a mechanism not involving the heat shock transcription factor.

3T3 Cells↗
PubMed1993

BVA congress.

Explore the source record for details and available documents.

Congresses as Topic↗
PubMed1992

dOct2, a Drosophila Oct transcription factor that functions in yeast.

Oct factors are members of the POU family of transcription factors that are shown to play important roles during development in mammals. Here we report the cDNA cloning and expression of a Drosophila Oct transcription factor. Whole mount in situ hybridization experiments revealed that the spatial expression patterns of this gene during embryonic development have not yet been observed for any other gene. In early embryogenesis, its transcripts are transiently expressed as a wide uniform band from 20% to 40% of the egg length, very similar to that of gap genes. This pattern progressively resolves into a series of narrower stripes followed by expression in 14 stripes. Subsequently, transcripts from this gene are expressed in the central nervous system and the brain. When expressed in the yeast Saccharomyces cerevisiae, this Drosophila factor functions as a strong, octamer-dependent activator of transcription. Our data strongly suggest possible functions for the Oct factor in pattern formation in Drosophila that might transcend the boundaries of genetically defined segmentation genes.

Amino Acid Sequence↗
PubMed1991

The SV40 core sequence functions as a repressor element in yeast.

Our previous studies showed that the AP-1 recognition element (ARE) present within the SV40 72-base pair (bp) enhancer will activate transcription in yeast when placed upstream of a truncated CYC1 promoter. However, the AP-2/AP-3 recognition element (also known as the core sequence TGTGGAAAG) from the SV40 enhancer was not able to activate CYC1-dependent transcription. In this report, we show that the core sequence, when cloned next to a yeast UAS (upstream activation sequence), can inhibit the transcriptional stimulatory activity of the UAS. We refer to this sequence as the upstream repressor element (URE) in yeast. Repression occurs in an orientation-independent fashion and irrespective of the placement of the URE between the UAS and TATA box or upstream of both of these elements. Furthermore, repression is seen when the URE is separated from the UAS by up to 214 bp. Interestingly, multiple copies of an activator site can overcome this repression. Gel-shift analysis and URE-probed proteins blots indicate the presence of two polypeptide chains capable of binding the URE in yeast. The experimental evidence suggests that either the repression associated with the URE sequence is mediated by a direct, one-to-one interaction between the proteins recognizing the URE and GCRE, or alternatively, that there is a direct interaction between the activator and repressor for a general transcription factor.

Base Sequence↗
PubMed1991

Synthetic oligonucleotides recreate Drosophila fushi tarazu zebra-stripe expression.

A complex array of activator and repressor elements located within 669 bp proximal to the fushi tarazu (ftz) transcriptional start site is sufficient to generate the "zebra-stripe" expression pattern characteristic of the ftz gene. P-element-mediated transformation and ftz promoter/lacZ fusion genes were used to characterize, in detail, several of these transcriptional control elements. By reconstructing promoters with synthetic oligonucleotides containing cis-regulators of stripe expression, we show that these regulatory sites can function as independent units to direct position-specific transcription in the Drosophila embryo. In particular, we demonstrate that multiple copies of a positive regulatory site can mediate expression in both the odd- and even-numbered parasegments throughout most of the germ band and that negative regulatory sites can transform a continuous pattern of gene expression into discrete stripes. The reconstructed promoter system presented provides an effective means of studying molecular mechanisms governing spatially restricted transcription in the early embryo.

Animals↗
PubMed1990

The yeast heat shock transcription factor contains a transcriptional activation domain whose activity is repressed under nonshock conditions.

Transcription of heat shock genes is induced by exposure of cells to elevated temperatures or other stress conditions. In yeast, it is thought that induction of transcription is mediated by conversion of a DNA-bound transcriptionally inactive form of the heat shock transcription factor (HSTF) to a DNA-bound transcriptionally active form. We have identified domains in HSTF involved in transcriptional activation and in repression of transcriptional activation at non-shock temperatures. We present evidence that a temperature-regulated transcriptional activation domain exists in HSTF and that this domain is essential for survival of yeast cells at heat shock temperatures. We propose a model for temperature-regulated transcriptional activation by a derepression mechanism.

Amino Acid Sequence↗
PubMed1990

cDNA clone encoding Drosophila transcription factor TFIID.

Proper initiation of transcription by RNA polymerase II requires the TATA-consensus-binding transcription factor TFIID. A cDNA clone encoding the Drosophila TFIID protein has been isolated and characterized. The deduced amino acid sequence reveals an open reading frame of 353 residues. The carboxyl-terminal 180 amino acids are approximately 80% identical to yeast TFIID and 88% identical to human TFIID. The amino-terminal portions of the yeast and Drosophila TFIID proteins lack appreciable homology, whereas the Drosophila and human amino termini appear qualitatively similar. In addition, the amino-terminal region of the Drosophila TFIID contains several sequence motifs that are found in other Drosophila proteins which appear to regulate transcription.

Amino Acid Sequence↗
PubMed1989

The caudal gene product is a direct activator of fushi tarazu transcription during Drosophila embryogenesis.

A drosophila pair-rule segmentation gene, fushi tarazu (ftz), encodes a protein which is expressed in a characteristic seven-stripe pattern. The promoter sequences that are sufficient for generating this spatially restricted pattern of expression are located within 669 base pairs upstream of the transcription start site. Multiple transcriptional activators and repressors interact with this 'zebra-stripe' promoter unit to bring about the positional specificity of ftz transcription. Here we report that the homoeodomain-containing protein encoded by caudal (cad) is one such regulator. The cad gene product can increase the level of ftz transcription in the posterior half of the embryo by interacting with multiple copies of a TTTATG consensus sequence located in the zebra-stripe unit. This result demonstrates one pathway by which the product of a maternally expressed segmentation gene, expressed in an antero-posterior concentration gradient, can directly regulate the expression of a pair-rule gene.

Animals↗
PubMed1989

Yeast YAP1 encodes a novel form of the jun family of transcriptional activator proteins.

The jun family of transcriptional activators includes mammalian AP-1 as well as the yeast regulatory protein GCN4. Recently, an additional transcriptional activator has been found in yeast that recognizes the TGACTCA sequence element common in GCN4/AP-1 sites. This factor was designated yAP-1. The structural gene for yAP-1 has now been isolated and characterized. The deduced amino acid sequence predicts a protein of 650 residues, considerably larger than GCN4 or c-Jun. The amino terminus of yAP-1 is homologous to the carboxy-terminal DNA-binding domains of GCN4 and c-Jun. Disruption of the YAP1 gene demonstrates this gene is not essential but is required for AP-1 recognition element-dependent transcriptional activation. DNA-affinity blots of proteins from YAP1 cells suggest the presence of additional TGACTCA-binding proteins other than GCN4 and yAP-1. Furthermore, expression of at least one of these related DNA-binding proteins appears to be under control of yAP-1.

Amino Acid Sequence↗