Showing papers by "University of Dundee published in 2000"
TL;DR: The p53 tumour-suppressor gene integrates numerous signals that control cell life and death, and the disruption of p53 has severe consequences when a highly connected node in the Internet breaks down.
Abstract: The p53 tumour-suppressor gene integrates numerous signals that control cell life and death. As when a highly connected node in the Internet breaks down, the disruption of p53 has severe consequences.
6,605 citations
Mark Raymond Adams1, Susan E. Celniker2, Robert A. Holt1, Cheryl A. Evans1 +191 more•Institutions (23)
TL;DR: The nucleotide sequence of nearly all of the approximately 120-megabase euchromatic portion of the Drosophila genome is determined using a whole-genome shotgun sequencing strategy supported by extensive clone-based sequence and a high-quality bacterial artificial chromosome physical map.
Abstract: The fly Drosophila melanogaster is one of the most intensively studied organisms in biology and serves as a model system for the investigation of many developmental and cellular processes common to higher eukaryotes, including humans. We have determined the nucleotide sequence of nearly all of the approximately 120-megabase euchromatic portion of the Drosophila genome using a whole-genome shotgun sequencing strategy supported by extensive clone-based sequence and a high-quality bacterial artificial chromosome physical map. Efforts are under way to close the remaining gaps; however, the sequence is of sufficient accuracy and contiguity to be declared substantially complete and to support an initial analysis of genome structure and preliminary gene annotation and interpretation. The genome encodes approximately 13,600 genes, somewhat fewer than the smaller Caenorhabditis elegans genome, but with comparable functional diversity.
6,180 citations
TL;DR: The results demonstrate that the specificities of protein kinase inhibitors cannot be assessed simply by studying their effect on kinases that are closely related in primary structure, and proposes guidelines for the use of protein Kinase inhibitors in cell-based assays.
Abstract: The specificities of 28 commercially available compounds reported to be relatively selective inhibitors of particular serine/threonine-specific protein kinases have been examined against a large panel of protein kinases. The compounds KT 5720, Rottlerin and quercetin were found to inhibit many protein kinases, sometimes much more potently than their presumed targets, and conclusions drawn from their use in cell-based experiments are likely to be erroneous. Ro 318220 and related bisindoylmaleimides, as well as H89, HA1077 and Y 27632, were more selective inhibitors, but still inhibited two or more protein kinases with similar potency. LY 294002 was found to inhibit casein kinase-2 with similar potency to phosphoinositide (phosphatidylinositol) 3-kinase. The compounds with the most impressive selectivity profiles were KN62, PD 98059, U0126, PD 184352, rapamycin, wortmannin, SB 203580 and SB 202190. U0126 and PD 184352, like PD 98059, were found to block the mitogen-activated protein kinase (MAPK) cascade in cell-based assays by preventing the activation of MAPK kinase (MKK1), and not by inhibiting MKK1 activity directly. Apart from rapamycin and PD 184352, even the most selective inhibitors affected at least one additional protein kinase. Our results demonstrate that the specificities of protein kinase inhibitors cannot be assessed simply by studying their effect on kinases that are closely related in primary structure. We propose guidelines for the use of protein kinase inhibitors in cell-based assays.
4,091 citations
TL;DR: The mechanism by which PKB is activated and the downstream actions of this multifunctional kinase are reviewed, as well as the evidence that PDK1 may be involved in the activation of protein kinases other than PKB, and the possibility that some of the currently postulated PKB substrates targets might in fact be phosphorylated byPDK1-regulated kinasesother than P KB.
Abstract: Phosphoinositide 3-kinases (PI3Ks) generate specific inositol lipids that have been implicated in the regulation of cell growth, proliferation, survival, differentiation and cytoskeletal changes One of the best characterized targets of PI3K lipid products is the protein kinase Akt or protein kinase B (PKB) In quiescent cells, PKB resides in the cytosol in a low-activity conformation Upon cellular stimulation, PKB is activated through recruitment to cellular membranes by PI3K lipid products and phosphorylation by 3'-phosphoinositide-dependent kinase-1 (PDK1) Here we review the mechanism by which PKB is activated and the downstream actions of this multifunctional kinase We also discuss the evidence that PDK1 may be involved in the activation of protein kinases other than PKB, the mechanisms by which this activity of PDK1 could be regulated and the possibility that some of the currently postulated PKB substrates targets might in fact be phosphorylated by PDK1-regulated kinases other than PKB
1,663 citations
TL;DR: The functional bioenergetics of isolated mitochondria are reviewed, with emphasis on the chemiosmotic proton circuit and the application (and occasional misapplication) of these principles to intact neurons.
Abstract: Mitochondria play a central role in the survival and death of neurons. The detailed bioenergetic mechanisms by which isolated mitochondria generate ATP, sequester Ca2+, generate reactive oxygen species, and undergo Ca2+-dependent permeabilization of their inner membrane are currently being applied to the function of mitochondria in situ within neurons under physiological and pathophysiological conditions. Here we review the functional bioenergetics of isolated mitochondria, with emphasis on the chemiosmotic proton circuit and the application (and occasional misapplication) of these principles to intact neurons. Mitochondria play an integral role in both necrotic and apoptotic neuronal cell death, and the bioenergetic principles underlying current studies are reviewed.
1,200 citations
TL;DR: The phosphorylation of a protein can alter its behaviour in almost every conceivable way, including modulation of its intrinsic biological activity, subcellular location, half-life and docking with other proteins.
1,094 citations
TL;DR: The results support prevention policies based on the classic risk factors but suggest potential for prevention beyond these, and changes in theclassic risk factors seem to partly explain the variation in population trends in CHD.
972 citations
TL;DR: Two supergene families encode proteins with glutathione S-transferase (GST) activity that detoxify a variety of electrophilic compounds, including oxidized lipid, DNA and catechol products generated by reactive oxygen species-induced damage to intracellular molecules.
Abstract: Two supergene families encode proteins with glutathione S-transferase (GST) activity: the family of soluble enzymes comprises at least 16 genes; the separate family of microsomal enzymes comprises at
932 citations
TL;DR: Findings indicate that LAMP-2 is critical for autophagy, and this theory is further substantiated by the finding that human Lamp-2 deficiency causing Danon's disease is associated with the accumulation of autophagic material in striated myocytes.
Abstract: Lysosome-associated membrane protein-2 (LAMP-2) is a highly glycosylated protein and an important constituent of the lysosomal membrane Here we show that LAMP-2 deficiency in mice increases mortality between 20 and 40 days of age The surviving mice are fertile and have an almost normal life span Ultrastructurally, there is extensive accumulation of autophagic vacuoles in many tissues including liver, pancreas, spleen, kidney and skeletal and heart muscle In hepatocytes, the autophagic degradation of long-lived proteins is severely impaired Cardiac myocytes are ultrastructurally abnormal and heart contractility is severely reduced These findings indicate that LAMP-2 is critical for autophagy This theory is further substantiated by the finding that human LAMP-2 deficiency causing Danon's disease is associated with the accumulation of autophagic material in striated myocytes
860 citations
TL;DR: The role model framework is of use in the assessment of the needs for staff to implement a curriculum, in the appointment and promotion of teachers and in the organization of a staff development programme.
Abstract: Teaching is a demanding and complex task. This guide looks at teaching and what it involves. Implicit in the widely accepted and far-reaching changes in medical education is a changing role for the medical teacher. Twelve roles have been identified and these can be grouped in six areas in the model presented: (1) the information provider in the lecture, and in the clinical context; (2) the role model on-the-job, and in more formal teaching settings; (3) the facilitator as a mentor and learning facilitator; (4) the student assessor and curriculum evaluator; (5) the curriculum and course planner; and (6) the resource material creator, and study guide producer. As presented in the model, some roles require more medical expertise and others more educational expertise. Some roles have more direct face-to-face contact with students and others less. The roles are presented in a 'competing values' framework-they may convey conflicting messages, e.g. providing information or encouraging independent learning, helpi...
826 citations
TL;DR: A computer vision system for tracking multiple people in relatively unconstrained environments is described and should provide a useful mechanism for bootstrapping and reinitialization of tracking using more specific but less robust human models.
TL;DR: Labelling studies, using the reactive AMP analogue 8-azido-[(32)P]AMP, indicate that the gamma subunit may participate directly in the binding of AMP within the complex.
Abstract: The AMP-activated protein kinase (AMPK) cascade plays an important role in the regulation of energy homeostasis within the cell. AMPK is a heterotrimer composed of a catalytic subunit (alpha) and two regulatory subunits (beta and gamma). We have isolated and characterized two isoforms of the gamma subunit, termed gamma2 and gamma3. Both gamma2 (569 amino acids) and gamma3 (492 amino acids) have a long N-terminal domain which is not present in the previously characterized isoform, gamma1. As with gamma1, mRNA encoding gamma2 is widely expressed in human tissues, whereas significant expression of gamma3 mRNA was only detected in skeletal muscle. Using isoform-specific antibodies, we determined the AMPK activity associated with the different gamma isoforms in a number of rat tissues. In most tissues examined more than 80% of total AMPK activity was associated with the gamma1 isoform, with the remaining activity being accounted for mainly by the gamma2 isoform. Exceptions to this were testis and, more notably, brain where all three isoforms contributed approximately equally to activity. There was no evidence for any selective association between the alpha1 and alpha2isoforms and the various gamma isoforms. However, the AMP-dependence of the kinase complex is markedly affected by the identity of the gamma isoform present, with gamma2-containing complexes having the greatest AMP-dependence, gamma3 the lowest, and gamma1 having an intermediate effect. Labelling studies, using the reactive AMP analogue 8-azido-[(32)P]AMP, indicate that the gamma subunit may participate directly in the binding of AMP within the complex.
TL;DR: Monitoring of the major component of Deltap, the mitochondrial membrane potential Deltapsim, in intact neurones exposed to excitotoxic stimuli, in the hope of establishing the causal relationships between cell death and mitochondrial dysfunction.
TL;DR: Variations in mortality may be partly explained by excess ICU workload, and this methodology may have implications for planning and clinical governance.
TL;DR: It is shown that the major Pro-directed phosphatase PP2A is conformation-specific and effectively dephosphorylates only the trans pSer/Thr-Pro isomer, and prolyl isomerase activity of Pin1 is essential for cell division in vivo.
TL;DR: The fact that Phb1/2 is a large multimeric complex, which provides protection of native peptides against proteolysis, suggests a functional homology with protein chaperones with respect to their ability to hold and prevent misfolding of newly synthesized proteins.
Abstract: Prohibitins are ubiquitous, abundant and evolutionarily strongly conserved proteins that play a role in important cellular processes. Using blue native electrophoresis we have demonstrated that human prohibitin and Bap37 together form a large complex in the mitochondrial inner membrane. This complex is similar in size to the yeast complex formed by the homologues Phb1p and Phb2p. In yeast, levels of this complex are increased on co-overexpression of both Phb1p and Phb2p, suggesting that these two proteins are the only components of the complex. Pulse–chase experiments with mitochondria isolated from phb1/phb2-null and PHB1/2 overexpressing cells show that the Phb1/2 complex is able to stabilize newly synthesized mitochondrial translation products. This stabilization probably occurs through a direct interaction because association of mitochondrial translation products with the Phb1/2 complex could be demonstrated. The fact that Phb1/2 is a large multimeric complex, which provides protection of native peptides against proteolysis, suggests a functional homology with protein chaperones with respect to their ability to hold and prevent misfolding of newly synthesized proteins.
TL;DR: This study lays the foundation for future work to establish the phospholipid-binding specificities of these proteins in vivo, and their physiological role(s).
Abstract: The second messenger phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P(3)] is generated by the action of phosphoinositide 3-kinase (PI 3-kinase), and regulates a plethora of cellular processes. An approach for dissecting the mechanisms by which these processes are regulated is to identify proteins that interact specifically with PtdIns(3,4,5)P(3). The pleckstrin homology (PH) domain has become recognized as the specialized module used by many proteins to interact with PtdIns(3,4,5)P(3). Recent work has led to the identification of a putative phosphatidylinositol 3,4,5-trisphosphate-binding motif (PPBM) at the N-terminal regions of PH domains that interact with this lipid. We have searched expressed sequence tag databases for novel proteins containing PH domains possessing a PPBM. Surprisingly, many of the PH domains that we identified do not bind PtdIns(3,4,5)P(3), but instead possess unexpected and novel phosphoinositide-binding specificities in vitro. These include proteins possessing PH domains that interact specifically with PtdIns(3,4)P(2) [TAPP1 (tandem PH-domain-containing protein-1) and TAPP2], PtdIns4P [FAPP1 (phosphatidylinositol-four-phosphate adaptor protein-1)], PtdIns3P [PEPP1 (phosphatidylinositol-three-phosphate-binding PH-domain protein-1) and AtPH1] and PtdIns(3,5)P(2) (centaurin-beta2). We have also identified two related homologues of PEPP1, termed PEPP2 and PEPP3, that may also interact with PtdIns3P. This study lays the foundation for future work to establish the phospholipid-binding specificities of these proteins in vivo, and their physiological role(s).
TL;DR: Microorganisms play important roles in the environmental fate of toxic metals and radionuclides with a multiplicity of mechanisms effecting transformations between soluble and insoluble forms and are of potential for both in situ and ex situ bioremedial treatment processes for solid and liquid wastes.
TL;DR: Changes in coronary care and secondary prevention were strongly linked with declining coronary endpoints.
TL;DR: In this paper, the results of a test programme to study the use of recycled concrete aggregate (RCA) in high-strength, 50 N/mm2 or greater, concrete are described.
Abstract: The results of a test programme to study the use of recycled concrete aggregate (RCA) in high-strength, 50 N/mm2 or greater, concrete are described. The effects of coarse RCA content on the ceiling strength, bulk engineering and durability properties of such concretes have been established. The results showed that up to 30% coarse RCA had no effect on concrete strength, but therafter there was a gradual reduction as the RCA content increased. A method of accommodating the effects of high RCA content, involving simple adjustment to water/cement ratio of the mix is given. It is shown that high-strength RCA concrete will have equivalent engineering and durability performance to concrete made with natural aggregates, for corresponding 28-day design strengths. The practical implications of the study for concrete construction are discussed.
TL;DR: PDK1 mediates activation of PKB, p70 S6 kinase and p90 Rsk in vivo, but is not rate-limiting foractivation of PKA, MSK1 and AMPK.
TL;DR: The authors conducted a qualitative formative assessment of academic writing by 12 postgraduate students of educational psychology and found that most found the process time consuming, intellectually challenging and socially uncomfortable, but effective in improving the quality of their own subsequent written work and developing other transferable skills.
Abstract: Reciprocal paired qualitative formative peer assessment of academic writing was undertaken by 12 postgraduate students of educational psychology. Overall, staff and peer assessments showed a very similar balance between positive and negative statements, but this varied according to assessment criterion. However, only half of all detailed formative assessment statements made showed some degree of correspondence between staff and peers. Nevertheless, there was very little evidence of conflict between the views of staff and peers-rather, they focused on different details. Subjective feedback from students indicated that most found the process time consuming, intellectually challenging and socially uncomfortable, but effective in improving the quality of their own subsequent written work and developing other transferable skills. The reliability and validity of this type of peer assessment thus appeared adequate, and the partiality of overlap in detail between staff and peer assessments suggested that the tria...
TL;DR: Evidence is provided that cyclic nucleotides are physiological substrates for MRP5 and may represent a novel pharmacological target for the enhancement of tissue levels of cGMP.
TL;DR: A review of the occurrence of cyanobacterial toxins in waterbodies with examples from national and local surveys is presented in this article, where properties of the known cyanobacteria toxins, associated health problems, and the likelihood of further toxins being discovered are discussed.
TL;DR: In this article, the results of two U.K. surveys of activity-based costing (ABC) adoption status of companies over a recent 5-year period are reviewed. But the authors focus on the non-users and not-users.
TL;DR: Results from bacteriophages and archaea show that the structural basis for the interaction of this motif with PCNA is extremely ancient, which helps to understand how these complex systems arose from ancestral organisms.
Abstract: The identification of proteins that interact with proliferating cell nuclear antigen (PCNA) has recently been a rapidly expanding field of discovery. PCNA is involved in many aspects of DNA replication and processing, forming a sliding platform that can mediate the interaction of proteins with DNA. It is striking that many proteins bind to PCNA through a small region containing a conserved motif; these include proteins involved in cell cycle regulation as well as those involved in DNA processing. Sequential and regulated binding of motif-containing proteins to PCNA may contribute to the ordering of events during DNA replication and repair. Results from bacteriophages and archaea show that the structural basis for the interaction of this motif with PCNA is extremely ancient. The analysis of how such functional motifs have been recruited to proteins in present day organisms helps us to understand how these complex systems arose from ancestral organisms.
TL;DR: Analysis of mammalian mtDNA by two-dimensional agarose gel electrophoresis indicates that two modes of mtDNA replication operate in mammalian cells and that changes in mtDNA copy number involve an alteration in the mode of mt DNA replication.
TL;DR: Only an integrated and multidisciplinary approach can determine if (a) N2-fixing symbioses with Gramineae really exist, and (b) if they are effective in agronomic terms.
TL;DR: It is suggested that p53 C-terminal lysine residues are the main sites of ubiquitin ligation, which target p53 for proteasome-mediated degradation.
Abstract: In normal cells, p53 is maintained at a low level by ubiquitin-mediated proteolysis, but after genotoxic insult this process is inhibited and p53 levels rise dramatically. Ubiquitination of p53 requires the ubiquitin-activating enzyme Ubc5 as a ubiquitin conjugation enzyme and Mdm2, which acts as a ubiquitin protein ligase. In addition to the N-terminal region, which is required for interaction with Mdm2, the C-terminal domain of p53 modulates the susceptibility of p53 to Mdm2-mediated degradation. To analyze the role of the C-terminal domain in p53 ubiquitination, we have generated p53 molecules containing single and multiple lysine-to-arginine changes between residues 370 and 386. Although wild-type (WT) and mutant molecules show similar subcellular distributions, the mutants display a higher transcriptional activity than WT p53. Simultaneous mutation of lysine residues 370, 372, 373, 381, 382, and 386 to arginine residues (6KR p53 mutant) generates a p53 molecule with potent transcriptional activity that is resistant to Mdm2-induced degradation and is refractory to Mdm2-mediated ubiquitination. In contrast to WT p53, transcriptional activity directed by the 6KR p53 mutant fails to be negatively regulated by Mdm2. Those differences are also manifest in HeLa cells which express the human papillomavirus E6 protein, suggesting that p53 C-terminal lysine residues are also implicated in E6-AP-mediated ubiquitination. These data suggest that p53 C-terminal lysine residues are the main sites of ubiquitin ligation, which target p53 for proteasome-mediated degradation.
TL;DR: Treatment of hepatoma cells with 5-aminoimidazole-4-carboxamide riboside (AICAR), an agent that activates AMP-activated protein kinase (AMPK), mimics the ability of insulin to repress PEPCK gene transcription and suggests activation of AMPK would inhibit hepatic gluconeogenesis in an insulin-independent manner and thus help to reverse the hyperglycemia associated with type 2 diabetes.
Abstract: Insulin regulates the rate of expression of many hepatic genes, including PEPCK, glucose-6-phosphatase (G6Pase), and glucose-6-phosphate dehydrogenase (G6PDHase). The expression of these genes is also abnormally regulated in type 2 diabetes. We demonstrate here that treatment of hepatoma cells with 5-aminoimidazole-4-carboxamide riboside (AICAR), an agent that activates AMP-activated protein kinase (AMPK), mimics the ability of insulin to repress PEPCK gene transcription. It also partially represses G6Pase gene transcription and yet has no effect on the expression of G6PDHase or the constitutively expressed genes cyclophilin or beta-actin. Several lines of evidence suggest that the insulin-mimetic effects of AICAR are mediated by activation of AMPK. Also, insulin does not activate AMPK in H4IIE cells, suggesting that this protein kinase does not link the insulin receptor to the PEPCK and G6Pase gene promoters. Instead, AMPK and insulin may lie on distinct pathways that converge at a point upstream of these 2 gene promoters. Investigation of the pathway by which AMPK acts may therefore give insight into the mechanism of action of insulin. Our results also suggest that activation of AMPK would inhibit hepatic gluconeogenesis in an insulin-independent manner and thus help to reverse the hyperglycemia associated with type 2 diabetes.