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Biomedical subjects

A Sutton

Publications and source records attributed to A Sutton.

At least 19 recordsLinked to original sources
PubMed2000

Phosphorylation of Cdc28 and regulation of cell size by the protein kinase CKII in Saccharomyces cerevisiae.

The CDK (cyclin-dependent kinase) family of enzymes is required for the G(1)-to-S-phase and G(2)-to-M-phase transitions during the cell-division cycle of eukaryotes. We have shown previously that the protein kinase CKII catalyses the phosphorylation of Ser-39 in Cdc2 during the G(1) phase of the HeLa cell-division cycle [Russo, Vandenberg, Yu, Bae, Franza and Marshak (1992) J. Biol. Chem. 267, 20317-20325]. To identify a functional role for this phosphorylation, we have studied the homologous enzymes in the budding yeast Saccharomyces cerevisiae. The S. cerevisiae homologue of Cdc2, Cdc28, contains a consensus CKII site (Ser-46), which is homologous with that of human Cdc2. Using in vitro kinase assays, metabolic labelling, peptide mapping and phosphoamino acid analysis, we demonstrate that this site is phosphorylated in Cdc28 in vivo as well in vitro. In addition, S. cerevisiae cells in which Ser-46 has been mutated to alanine show a decrease in both cell volume and protein content of 33%, and this effect is most pronounced in the stationary phase. Because cell size in S. cerevisiae is regulated primarily at the G(1) stage, we suggest that CKII contributes to the regulation of the cell cycle in budding yeast by phosphorylation of Cdc28 as a checkpoint for G(1) progression.

Amino Acid Sequence↗
PubMed2000

The silencing protein SIR2 and its homologs are NAD-dependent protein deacetylases.

Homologs of the chromatin-bound yeast silent information regulator 2 (SIR2) protein are found in organisms from all biological kingdoms. SIR2 itself was originally discovered to influence mating-type control in haploid cells by locus-specific transcriptional silencing. Since then, SIR2 and its homologs have been suggested to play additional roles in suppression of recombination, chromosomal stability, metabolic regulation, meiosis, and aging. Considering the far-ranging nature of these functions, a major experimental goal has been to understand the molecular mechanism(s) by which this family of proteins acts. We report here that members of the SIR2 family catalyze an NAD-nicotinamide exchange reaction that requires the presence of acetylated lysines such as those found in the N termini of histones. Significantly, these enzymes also catalyze histone deacetylation in a reaction that absolutely requires NAD, thereby distinguishing them from previously characterized deacetylases. The enzymes are active on histone substrates that have been acetylated by both chromatin assembly-linked and transcription-related acetyltransferases. Contrary to a recent report, we find no evidence that these proteins ADP-ribosylate histones. Discovery of an intrinsic deacetylation activity for the conserved SIR2 family provides a mechanism for modifying histones and other proteins to regulate transcription and diverse biological processes.

Acetylation↗
PubMed2000

rpoS mutants in archival cultures of Salmonella enterica serovar typhimurium.

Long-term survival under limited growth conditions presents bacterial populations with unique environmental challenges. The existence of Salmonella enterica serovar Typhimurium cultures undisturbed in sealed nutrient agar stab vials for 34 to 45 years offered a unique opportunity to examine genetic variability under natural conditions. We have initiated a study of genetic changes in these archival cultures. We chose to start with examination of the rpoS gene since, among gram-negative bacteria, many genes needed for survival are regulated by RpoS, the stationary-phase sigma factor. In each of 27 vials examined, cells had the rpoS start codon UUG instead of the expected AUG of Salmonella and Escherichia coli strains recorded in GenBank. Ten of the 27 had additional mutations in the rpoS gene compared with the X77752 wild-type strain currently recorded in GenBank. The rpoS mutations in the 10 strains included two deletions as well as point mutations that altered amino acid sequences substantially. Since these stored strains were derived from ancestral cells inoculated decades ago and remained undisturbed, it is assumed that the 10 rpoS mutations occurred during storage. Since the remaining 17 sequences were wild type (other than in the start codon), it is obvious that rpoS remained relatively stable during decades of sealed storage.

Amino Acid Substitution↗
PubMed1999

The reproducibility of central aortic blood pressure measurements in healthy subjects using applanation tonometry and sphygmocardiography.

AIM: Sphygmocardiography via applanation tonometry is a non-invasive, bedside technology which utilises tonometric analysis of the radial artery pulse wave and measurement of peripheral arterial blood pressure (BP) to derive a central arterial pulse wave, central arterial BP and related indices. The present study was designed to determine: (1) the inter-operator variability in measurements obtained using this technique; (2) the relationship between measured peripheral arterial BP and derived central arterial BP. METHOD: Multiple measurements were made from 25 healthy subjects (15 male), mean age 33 (s.d. 10.3) years, mean arterial BP 90 (s.d. 12) mm Hg by two trained observers at the same time of day on three separate occasions. RESULTS: The mean inter-operator difference was 0.1 (s.d. 1.7) mm Hg for derived systolic aortic BP and 0.1 (s.d. 0.7) mm Hg for derived diastolic aortic BP (Bland and Altman analysis). Pulse wave Augmentation Index (AIx) values, ranged from -22% to +40%, with the inter-operator measurement difference being only 0.4 (s.d. 6.4)%. Buckberg ratio measurements ranged from 119% to 254%, with the inter-operator measurement difference being only 2.7 (s.d. 15.4)%. The relationship between derived central systolic BP and peripheral systolic BP readings in individual patients was not constant and showed significant variance when compared on different days (ANOVA, P = 0.03). This was not explained by any significant variance in heart rate (ANOVA, P = 0.39). CONCLUSION: Applanation tonometry has excellent inter-observer reproducibility when used by trained observers. Moreover, the inconsistency in the relationship between peripheral and central aortic BP suggests that the former is not a perfect surrogate for the latter. Further prospective studies are required to define whether derived central aortic BP may be a better predictor of cardiovascular morbidity and mortality and the impact of different antihypertensive therapies on the relationship between peripheral and central arterial BP.

Adult↗
PubMed1999

Analyte comparisons between 2 clinical chemistry analyzers.

The purpose of this study was to assess agreement between a wet reagent and a dry reagent analyzer. Thirteen analytes (albumin, globulin, alkaline phosphatase, alanine aminotransferase, amylase, urea nitrogen, calcium, cholesterol, creatinine, glucose, potassium, total bilirubin, and total protein) for both canine and feline serum were evaluated. Concordance correlations, linear regression, and plots of difference against mean were used to analyze the data. Concordance correlations were excellent for 8 of 13 analytes (r > or = 0.90); the correlations for albumin, potassium, and calcium were clinically unreliable. The linear regression analysis revealed that several analytes had slopes significantly different from unity, which was likely related to methodological differences. Compared to the wet reagent analyzer, the dry reagent analyzer showed excellent agreement for alkaline phosphatase, alanine aminotransferase, amylase (feline), urea nitrogen, cholesterol, creatinine, glucose, total bilirubin (canine), and total protein. However, it showed only slight to substantial agreement for amylase (canine), calcium, albumin, potassium, and total bilirubin (feline).

Animals↗
PubMed1999

Nursing in genitourinary medicine: 10 years on from the Monks Report. The membership of GUNA. Genito-Urinary Nurses Association.

Various reports and surveys since 1988 have highlighted differences in gradings and diversity of roles and responsibilities of nurses in genitourinary medicine (GUM). In addition there was no method of defining how many nurses work in the speciality. Therefore the Genito-Urinary Nurses Association (GUNA) undertook a survey last year to establish how many nurses, of what grade, undertaking what roles are working in GUM clinics. Over half the clinics in the UK responded, from full-time busy London clinics to small part-time rural clinics. The results suggest approximately 2000 nurses working in GUM nationally, with a wide diversity of grades, roles and responsibilities still apparent 10 years after the Monks Report. Further analysis of the data obtained by the survey is given in the report.

Female Urogenital Diseases↗
PubMed1998

Scleroderma fibroblasts promote migration of mononuclear leucocytes across endothelial cell monolayers.

Perivascular infiltrates of inflammatory cells are a hallmark of lesional skin in scleroderma. We have explored the potential for scleroderma fibroblasts to modulate mononuclear leucocyte migration across endothelial cell monolayers in tissue culture, and to regulate expression of endothelial cell adhesion molecules. Fibroblasts were grown from skin biopsies of eight patients with active diffuse cutaneous scleroderma and from four healthy controls. Co-culture and conditioned medium transfer experiments examined the effect of soluble fibroblast products on mononuclear leucocyte (U937) cell migration across endothelial cell (1E-7) monolayers grown on tissue culture inserts. Co-culture of scleroderma, but not control fibroblasts, promoted transendothelial migration of U937 cells. Scleroderma fibroblast-conditioned medium had qualitatively similar effects and equivalent results were obtained using Jurkat-6 (T lymphocyte) cells, and with peripheral blood mononuclear cells from a patient with diffuse cutaneous scleroderma. Promotion of leucocyte migration does not appear to result from increased endothelial adhesion molecule expression, since fibroblast-conditioned medium did not up-regulate endothelial cell expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) or E-selectin. Moreover, leucocyte migration across cytokine-activated endothelial cell layers in co-culture with fibroblasts was less than across resting cells, although the selective effect of scleroderma fibroblast co-culture persisted. Recombinant monocyte chemoattractant protein-1 (MCP-1) or IL-8 increased passage of mononuclear leucocytes across endothelial cell monolayers, whilst anti-MCP-1, but not anti-IL-8 antibodies, significantly reduced the effect of fibroblast conditioned medium. These data suggest that systemic sclerosis (SSc) fibroblasts promote leucocyte migration across endothelial cell monolayers in tissue culture via an MCP-1-dependent mechanism. These findings may be relevant to the perivascular mononuclear leucocyte infiltrates characteristic of early SSc lesions.

Cell Line, Transformed↗
PubMed1997

The Cak1p protein kinase is required at G1/S and G2/M in the budding yeast cell cycle.

The CAK1 gene encodes the major CDK-activating kinase (CAK) in budding yeast and is required for activation of Cdc28p for cell cycle progression from G2 to M phase. Here we describe the isolation of a mutant allele of CAK1 in a synthetic lethal screen with the Sit4 protein phosphatase. Analysis of several different cak1 mutants shows that although the G2 to M transition appears most sensitive to loss of Cak1p function, Cak1p is also required for activation of Cdc28p for progression from G1 into S phase. Further characterization of these mutants suggests that, unlike the CAK identified from higher eukaryotes, Cak1p of budding yeast may not play a role in general transcription. Finally, although Cak1 protein levels and in vitro protein kinase activity do not fluctuate during the cell cycle, at least a fraction of Cak1p associates with higher molecular weight proteins, which may be important for its in vivo function.

CDC28 Protein Kinase, S cerevisiae↗
PubMed1997

A comparison of the effects of tramadol and morphine on gastric emptying in man.

In a previous study using an electrical bioimpedance technique and the paracetamol absorption test, we demonstrated that 0.09 mg.kg-1 of morphine delayed gastric emptying in healthy human volunteers. The aim of this study was to investigate whether analgesic doses of tramadol would cause a delay in gastric emptying similar to conventional opioids. Using the same volunteers and techniques as in our previous study, placebo or tramadol (1 mg.kg-1) was given in a randomised, double-blinded, cross-over placebo-controlled study. Gastric emptying was measured concurrently by a noninvasive epigastric bioimpedance technique and by the paracetamol absorption test. After the ingestion of 500 ml of deionised water plus paracetamol 1.5 g, the mean (SEM) time taken for gastric volume to decrease to 50% (t0.5) was recorded. No difference in gastric emptying rates (t0.5) between placebo, 7.7 (1 min), and tramadol, 9.5 (2 min), was noted. In our previous study, morphine prolonged t0.5 to 21 (3) min (p < 0.03). The maximum concentration and area under the curve of serum paracetamol concentrations following morphine were significantly different from placebo (p < 0.05) and tramadol (p < 0.05). We conclude that tramadol at a dose of 1 mg.kg-1 does not delay gastric emptying in humans.

Absorption↗
PubMed1997

Authority, autonomy, responsibility and authorisation: with specific reference to adolescent mental health practice.

Standards for professional training and practice are defined by accrediting organisations or statutory bodies. These describe the arena in which the practitioner may speak with authority. The sphere of authorised practice is further delineated by the external resources available. Within this explicit framework, unconscious mental processes can affect the professional response in potentially adverse ways. This is particularly important in mental health practice. Professionals must be prepared to examine their own responses on this basis in order to enhance their knowledge of the patient and minimise the possibilities of the patient becoming the victim of the professional's own psychopathology. The maintenance of such a position in an institution or organisation requires a similar process within its structure in order to provide the necessary setting and define the limits of good practice. In this paper, the field of adolescent mental health is specifically examined.

Adolescent↗
PubMed1996

The Cdk-activating kinase (CAK) from budding yeast.

Activation of the cyclin-dependent kinases to promote cell cycle progression requires their association with cyclins as well as phosphorylation of a threonine (residue 161 in human p34cdc2). This phosphorylation is carried out by CAK, the Cdk-activating kinase. We have purified and cloned CAK from S. cerevisiae. Unlike CAKs from other organisms, Cak1p is active as a monomer, has full activity when expressed in E. coli, and is not a component of the basal transcription factor, TFIIH. A temperature-sensitive mutation in CAK1 confers a G2 delay accompanied by low Cdc28p protein kinase activity and shows genetic interactions with altered expression of the gene for the major mitotic cyclin, CLB2. Our data raise the intriguing possibility that p40MO15-cyclin H-MAT1, identified as the predominant CAK in vertebrate cell extracts, may not function as a physiological CAK.

Amino Acid Sequence↗
PubMed1996

Evidence of sensitivity to structural contrasts in the literature on children's language comprehension.

This paper reviews the developmental literature on grammatical knowledge in language comprehension in the preschool years from the perspective of sensitivity to structural contrasts. This concept differs from mastery of individual grammatical structures. Structural sensitivity focuses on increments of partial grammatical knowledge that can be observed in distinctive response patterns to contrasting grammatical structures. Direct evidence of sensitivity to structural contrasts is found in comprehension studies that measured differential responding. Indirect evidence of sensitivity can also be discovered by detailed examination of the data presented in several additional studies. The evidence suggests that there may be a developmental sequence of increasing sensitivity with age to finer distinctions and to more detailed aspects of grammatical structure. The notion of sensitivity to structural contrasts has implications for future research.

Age Factors↗
PubMed1995

Protracted post-traumatic confusional state treated with physostigmine.

A case study is presented of confusion in a head-injured man, lasting for more than 2 years, when intermittent treatment with physostigmine resulted in progressive improvement in both confusion and usable cognitive functions. Aetiological mechanisms and implications for treatment plans are discussed.

Awareness↗
PubMed1992

SIT4 protein phosphatase is required for the normal accumulation of SWI4, CLN1, CLN2, and HCS26 RNAs during late G1.

In Saccharomyces cerevisiae, the RNA levels of the G1 cyclins CLN1, CLN2, and HCS26 increase dramatically during the late G1 phase of the cell cycle. The SIT4 gene, which encodes a serine/threonine protein phosphatase, is required for the normal accumulation of CLN1, CLN2, and HCS26 RNAs during late G1. This requirement for SIT4 in normal G1 cyclin RNA accumulation is at least partly via SWI4. Strains containing mutations in SIT4 are sensitive to the loss of either CLN2 or CLN3 function. At the nonpermissive temperature, temperature-sensitive sit4 strains are blocked for both bud emergence and DNA synthesis. Heterologous expression of CLN2 in the absence of SIT4 function results in DNA synthesis, but most of the cells are still blocked for bud emergence. Therefore, SIT4 is required for at least two late G1 or G1/S functions: the normal accumulation of G1 cyclin RNAs (which is required for DNA synthesis) and some additional function that is required for bud emergence or cell cycle progression through late G1 or G1/S.

Cyclins↗
PubMed1991

Characterization of SIS1, a Saccharomyces cerevisiae homologue of bacterial dnaJ proteins.

The Saccharomyces cerevisiae SIS1 gene was identified as a high copy number suppressor of the slow growth phenotype of strains containing mutations in the SIT4 gene, which encodes a predicted serine/threonine protein phosphatase. The SIS1 protein is similar to bacterial dnaJ proteins in the amino-terminal third and carboxyl-terminal third of the proteins. In contrast, the middle third of SIS1 is not similar to dnaJ proteins. This region of SIS1 contains a glycine/methionine-rich region which, along with more amino-terminal sequences, is required for SIS1 to associate with a protein of apparent molecular mass of 40 kD. The SIS1 gene is essential. Strains limited for the SIS1 protein accumulate cells that appear blocked for migration of the nucleus from the mother cell into the daughter cell. In addition, many of the cells become very large and contain a large vacuole. The SIS1 protein is localized throughout the cell but is more concentrated at the nucleus. About one-fourth of the SIS1 protein is released from a nuclear fraction upon treatment with RNase. We also show that overexpression of YDJ1, another yeast protein with similarity to bacterial dnaJ proteins, can not substitute for SIS1.

Amino Acid Sequence↗