bluuebunny/crossref_metadata_2025 huggingface dataset:
p53 Accumulation, defective cell proliferation, and early embryonic lethality in mice lacking
tsg101
Authors: Ruland Jürgen , Sirard Christian , Elia Andrew , MacPherson David , Wakeham Andrew , Li Limin , Luis de la Pompa José , Cohen Stanley N. , Mak Tak W.
Published: July 2002

Functional inactivation of the tumor susceptibility gene
tsg101
in NIH 3T3 fibroblasts results in cellular
transformation and the ability to form metastatic tumors in nude mice.
The N-terminal region of tsg101 protein is structurally similar to the
catalytic domain of ubiquitin-conjugating enzymes, suggesting a
potential role of tsg101 in ubiquitin-mediated protein degradation. The
C-terminal domain of TSG101 can function as a repressor of
transcription. To investigate the physiological function of
tsg101
, we generated a null mutation of the mouse gene
by gene targeting. Homozygous
tsg101
−/− embryos fail
to develop past day 6.5 of embryogenesis (E6.5), are reduced in size,
and do not form mesoderm. Mutant embryos show a decrease in cellular
proliferation
in vivo
and
in vitro
but no
increase in apoptosis. Although levels of
p53
transcripts were not affected in
tsg101
−/− embryos,
p53 protein accumulated dramatically, implying altered
posttranscriptional control of p53. In addition, transcription of the
p53 effector, cyclin-dependent kinase inhibitor
p21
WAF-1/CIP-1
, was increased 5- to 10-fold, whereas
activation of MDM2 transcription secondary to p53 elevation was not
observed. Introduction of a
p53
null mutation into
tsg101
−/− embryos rescued the gastrulation defect
and prolonged survival until E8.5. These results demonstrate that
tsg101
is essential for the proliferative burst before
the onset of gastrulation and establish a functional connection between
tsg101
and the p53 pathway
in vivo
.
Brain responses associated with consciousness of breathlessness (air hunger)
Authors: Liotti Mario , Brannan Stephen , Egan Gary , Shade Robert , Madden Lisa , Abplanalp Bart , Robillard Rachel , Lancaster Jack , Zamarripa Frank E. , Fox Peter T. , Denton Derek
Published: July 2002

Little is known about the physiological mechanisms subserving the
experience of air hunger and the affective control of breathing in
humans. Acute hunger for air after inhalation of CO
2
was
studied in nine healthy volunteers with positron emission tomography.
Subjective breathlessness was manipulated while end-tidal
CO
2
- was held constant. Subjects experienced a
significantly greater sense of air hunger breathing through a face mask
than through a mouthpiece. The statistical contrast between the two
conditions delineated a distributed network of primarily
limbic/paralimbic brain regions, including multiple foci in dorsal
anterior and middle cingulate gyrus, insula/claustrum,
amygdala/periamygdala, lingual and middle temporal gyrus,
hypothalamus, pulvinar, and midbrain. This pattern of activations was
confirmed by a correlational analysis with breathlessness ratings. The
commonality of regions of mesencephalon, diencephalon and
limbic/paralimbic areas involved in primal emotions engendered by the
basic vegetative systems including hunger for air, thirst, hunger,
pain, micturition, and sleep, is discussed with particular reference to
the cingulate gyrus. A theory that the phylogenetic origin of
consciousness came from primal emotions engendered by immediate threat
to the existence of the organism is discussed along with an alternative
hypothesis by Edelman that primary awareness emerged with processes of
ongoing perceptual categorization giving rise to a scene [Edelman,
G. M. (1992)
Bright Air, Brilliant Fire
(Penguin,
London)].
Receiving shadows: governance and liminality in the night‐time economy1
Authors: Hobbs Dick , Lister Stuart , Hadfield Philip , Winlow Simon , Hall Steve
Published: July 2002

This paper focuses upon the emergence of the night‐time economy both materially and culturally as a powerful manifestation of post‐industrial society. This emergence features two key processes: firstly a shift in economic development from the industrial to the post‐industrial; secondly a significant orientation of urban governance involving a move away from the traditional managerial functions of local service provision, towards an entrepreneurial stance primarily focused on the facilitation of economic growth. Central to this new economic era is the identification and promotion of liminality. The State's apparent inability to control these new leisure zones constitutes the creation of an urban frontier that is governed by commercial imperatives.
Intergroup Conflict
Authors: Hewstone Miles , Greenland Katy
Published: July 2002

This article outlines some of the main social‐psychological bases of intergroup conflict, illustrating the many ways in which social psychology can contribute to the study of conflict. The most convincing theoretical account is provided by perspectives that concentrate on the distinct nature of intergroup phenomena (especially social identity theory). Two of the most promising social‐psychological interventions to reduce intergroup conflict are also reviewed. The first is based on improving contact between members of previously hostile groups; the second attempts to change the structure of social categorizations (via decategorization, recategorization, and crossed categorization). This social‐psychological perspective is proposed as a key part of a necessary, multi‐disciplinary approach to intergroup conflict.
Determinants of Differential Acculturation of Developmental Timetables among Adolescent Immigrants to Germany
Authors: Schmitt‐Rodermund Eva , Silbereisen Rainer K.
Published: July 2002

The acculturation of developmental timetables for autonomy was studied in a sample of 220 ethnic German immigrants from Romania, Poland, and countries of the former Soviet Union. A distinction was made between the timing of autonomy from parental supervision and autonomy in social relationships. Four waves of semi‐annual asessments were conducted for two groups, newcomers (0–18 months of residence in Germany) and experienced immigrants (more than 18–36 months). From these assessments a gradual acculturation of timetables was observed, resulting, as expected, in considerably earlier ages at which the two facets of autonomy were accomplished. The rate of acculturation was predicted to be related to prior interindividual differences in parent‐adolescent interaction and frequency of adolescents' activities outside the home. As expected, higher levels of conflict and permissiveness reported by parents, and lower levels of monitoring and more frequent out‐of‐home activities, corresponded to a more accelerated acculturation to earlier timetables concerning autonomy from parental supervision—especially among the newcomers. With regard to autonomy in social relationships, the results were less clear, presumably due to the fact that prior interpersonal experiences were less associated with this facet of autonomy. Higher levels of the family variables studied encourage “escape” from the constraints of the newcomers' provisional homes, and thus provide opportunities to make contacts and adopt the lifestyle of adolescents from the general German population. In general, prediction was not possible concerning adolescents whose residence in Germany was longer than 18 months. For this group of immigrants, such escapes no longer serve the expected purpose because they have access to other opportunities, which were not measured in this study.
Metropolitan Development in a Transitional Socialist Economy: Spatial Restructuring in the Pearl River Delta, China
Authors: Lin George C. S.
Published: July 2002

The dynamic of globalisation and urban change in Europe and North America has been extensively documented. Relatively little is known about the processes and consequences of spatial restructuring in metropolitan regions within the context of a transitional socialist economy. This study investigates economic restructuring and spatial transformation in one of the most dynamic metropolitan regions in China. Deregulation of the post-reform socialist central state has allowed Chinese peasants to diversify agricultural production and to industrialise the rural economy according to various personal strengths and the changing market demand. Despite the rapid commercialisation and industrialisation of the regional economy, there has been no growing concentration of population and production facilities in large cities. The loci of accelerated economic growth, increased population mobility and massive land-use transformation have been in the intermediate zones surrounding and between metropolitan centres. Rapid expansion of the extended metropolitan zone has been driven primarily by forces of rural industrialisation at the grassroots level rather than a result of urban sprawl. The on-going processes and evolving patterns of 'urban-rural integration' ( chengxiang yitifa) in Chinese extended metropolitan regions demonstrate the complexity of the relationship between industrialisation and urbanisation in different political economies and question the adequacy of the widely accepted urban-rural dichotomy. The intrusion of global forces has not homogenised local particularities. Global capitalism has to seek shelter from locally specific conditions in order to take root in socialist soil.
Rescuing Aunt Sally: Taking Institutional Theory Seriously in Urban Politics
Authors: Lowndes Vivien
Published: July 2002

The sub-discipline of 'urban politics' has been constructed in opposition to a traditional version of 'institutional theory'-an approach that collapsed the political processes affecting urban communities with the workings of elected local government. Attention has shifted towards the broader influences on local decision-making and to the growing fragmentation of urban government and the rise of 'partnerships'. The article argues that recent developments, far from signifying the last gasp of 'institutionalism', call for a reformulated theory of the (diverse) institutional constraints within which urban political processes operate. Drawing on insights from the 'new institutionalism', the article discusses processes of institutional change and differentiation ; the underlying shift from strong to weak forms of institutional constraint; and the challenges involved in redesigning local political institutions.
Bulgarian Housing Reform and Forms of Housing Provision
Authors: Tsenkova Sasha
Published: July 2002

The objective of this paper is to explore the extent to which state intervention and market forces affect the housing provision system. Existing Bulgarian housing policy reform will be analysed with particular emphasis on the dynamics of change, the formation of a new market-oriented system and its considerable impact on innovation in the structures of housing provision. Relevant issues will be brought together to present the wide range of factors which contribute to the formation of nationally specific private and public forms of housing provision. Relevant issues will be brought together to present the wide range of factors which contribute to the formation of nationally specific private and public forms of housing provision, identifying their potential and constraints, and outlining major characteristics. Of primary concern is the extent to which a radical change of relationship between the state and the market, as occurred in Bulgaria, can modify the evolution of different forms of housing provision and the performance of key actors.
Government Policies and Private Housing Prices in Singapore
Authors: Phang Sock-Yong , Wong Wing-Keung
Published: July 2002

Regression models are used to show that interest rates, income growth rates and the supply of housing have not played a statistically significant role in the determination of private housing prices in Singapore between 1975 and 1994. Instead, private housing prices in Singapore were highly correlated with the prices for public-sector-built housing. Moreover, the timing of government policies relating to the use of compulsory savings for private housing finance purposes, the liberalisation of rules on public housing ownership criterion as well as for housing finance had a significant impact on private housing prices.
Segregation in Vienna: Impacts of Market Barriers and Rent Regulations
Authors: Giffinger Rudolf
Published: July 2002

Market barriers may have an enormous influence on the spatial segregation patterns of immigrant groups. This is specifically the case in Vienna, Austria, where accessibility rules with respect to different segments of the housing stock can be seen as one of the most important factors in explaining the residential patterns of Turks and Yugoslavs. Describing and explaining the patterns of these groups is the central issue in this paper. Conclusions are drawn with a reference to recent economic and social trends and the ongoing deregulation of the housing market.
Evaluation of Investor Behaviour in Urban Regeneration
Authors: Adair Alastair , Berry Jim , McGreal Stanley , Deddis Bill , Hirst Suzanne
Published: July 2002

The financing of urban regeneration and levering of private-sector investment remains a major policy issue. This article, utilising the results of a behavioural survey, analyses the motivations of investors and their participation across a range of regeneration initiatives. The pattern of investment activity over the market cycle, reasons for holding an urban regeneration portfolio, evaluative factors and perspectives concerning the attraction of private finance into urban regeneration are considered. The findings indicate that regeneration initiatives provide a significant cushioning effect but that, in terms of holding projects in a property portfolio or evaluating schemes, market-based factors relating to return and risk are core influences.
Growth-pole Strategies in Regional Economic Planning: A Retrospective View
Authors: Parr John B.
Published: July 2002

The paper continues from Part 1 which appeared in the previous issue of the journal. The primary concern is with neglected aspects of the growth-pole strategy, particularly as these relate to its implementation. Of importance here are the spatial configuration of the planned poles, the economic activity to be located within these, the spillover effects of a planned pole, and the presence of a pole within an existing urban system. Consideration is also given to the failure, abandonment and non-adoption of the strategy and to the reasons for this. It is argued that growth-pole strategy has never been evaluated in terms of an adequate conceptual framework, and the rudiments of one such framework are outlined.
Cultural Activities: Source of Competitiveness and Prosperity in Urban Regions
Authors: Dziembowska-Kowalska Jolanta , Funck Rolf H.
Published: July 2002

Over the past two decades, the potential of cultural activities to contribute to regional economic development has been increasingly recognised. This paper provides an analysis of the importance of the cultural sector for the economic prosperity of an urban region using the example of the city of Karlsruhe in Germany. It argues that the evaluation of the role of this sector involves the assessment of its direct contribution to regional performance and its impact on the competitiveness of other economic activities located there. The paper contains an empirical part where an effort is made to measure these effects. Finally, the role played by cultural initiatives in the regional development of the urban area of Karlsruhe is discussed.
Growth-pole Strategies in Regional Economic Planning: A Retrospective View
Authors: Parr John B.
Published: July 2002

The paper undertakes a detailed examination of growth-pole strategies, an emphasis in regional economic planning during the 1960s which never lived up to its early promise. The initial concern is with the origins of the strategy, particularly the manner in which the work of Perroux (on dominance and economic space) became modified to form a normative concept in regional economic planning. Consideration is given to the various regional-problem settings in which the growth-pole strategy has been advocated. These settings reflected such policy concerns as depressed-area revival, the encouragement of regional deconcentration, the modification of a national urban system, the pursuit of interregional balance, etc. Attention then turns to the fundamental nature and underlying rationale of the strategy. The paper is continued in Part 2 which appears in the next issue of the journal.
Ego identity status research in the new millennium
Authors: Kroger Jane
Published: July 2002

The present brief report overviews developments in the field of ego identity status research over the past 35 years and suggests directions for identity status research as we enter the new millenium. In the mid 1960s, James Marcia (1966, 1967) expanded Erik Erikson’s (1968) concept of ego identity to suggest four qualitatively different styles by which late adolescents undertake identity-deifning psychosocial commitments. Over the ensuing decades, identity status research focused primarily on validating the identity statuses and finding associated personality correlates, examining patterns of change over time, and investigating familial communication patterns associated with each of the identity positions. Research in the new millennium might fruitfully address the course and contents of identity beyond late adolescence as well as predictors of developmental arrest. A greater range of developmental contexts in which identity formation occurs is also in need of examination. Ultimately, an individual’s interpretation of context and further mediating events that may be associated with identity status resolutions are critical to a more complete understanding of the identity formation process.
Mothers’ empathic understanding of their preschoolers’ internal experience: Relations with early attachment
Authors: Oppenheim David , Koren-Karie Nina , Sagi Abraham
Published: July 2002

This study examined the links between mothers’ empathic understanding of their preschoolers’ internal experience and early infant-mother attachment. The empathic understanding of 118 mothers of 4.5-year-olds was assessed by showing them three videotaped segments of observations of their children and themselves and interviewing them regarding their children’s and their own thoughts and feelings. Interviews were rated and then classi” ed into one empathic and three nonempathic categories, and mothers’ misperceptions of the observations were coded as well. Infant-mother attachment classifications obtained using the Strange Situation when infants were 12 months old were also available. Results showed associations between mothers’ empathic understanding classifications and children’s attachment classifications as well as differences between mothers of secure and insecure children on one of the two interview composite scores. Also, mothers of insecurely attached children had more misperceptions than those of securely attached children. The contributions of this study to the work on mothers’ representations of their children are discussed.
The Value of Historical Control Data—Scientific Advantages for Pathologists, Industry and Agencies
Authors: Deschl Ulrich , Kittel Birgit , Rittinghausen Susanne , Morawietz Gerd , Kohler Manfred , Mohr Ulrich , Keenan Charlotte
Published: July 2002

Historical control tumor data are useful in the interpretation of long-term rodent carcinogenicity bioassays, especially to assess the occurrence of rare tumors and marginally increased tumor incidences. The major prerequisites to compare historical control data with studies under evaluation are the validity and consistency of the respective databases. The RITA (Registry of Industrial Toxicology Animal-data) database for historical data of tumors and pre-neoplastic lesions collects data according to highly standardized procedures including tissue sampling and trimming, histopathology according to internationally harmonized nomenclature and diagnostic criteria, and peer review. All lesions that are entered are unanimously diagnosed according to IARC (International Agency for Research on Cancer)/WHO criteria. The validity of data is additionally confirmed by a complete peer review performed by a database pathologist. Equivocal diagnoses and selected cases are additionally submitted to a panel of RITA pathologists. In the RITA database, there are currently 10,896 rats from 106 studies with more than 17,604 primary tumors and 16,551 pre-neoplastic lesions. The RITA database for historical control data for Wistar and Sprague Dawley rats as well as for different mouse strains is briefl ydescribed. Based upon RITA background data, the survival rate of Wistar rats has been consistent over a period of 10 years. The occurrence of tumor-bearing animals also shows a stable percentage over a decade. Additionally, examples of how historical control data may support carcinogenic risk assessment in cases of rare tumors or marginally increased incidences of tumors and pre-neoplastic lesions are given.
Segmentation and selection contribute to local processing in hierarchical analysis
Authors: Han Shihui , Humphreys Glyn W.
Published: July 2002

The present study examined the role of segmentation and selection processes when we respond to local elements in hierarchical stimuli. The ease of segmentation and selection of an individual local element from hierarchical patterns was manipulated by making one local element substantially distinct from the others in colour. Experiment 1 showed that, when attention was spread across the global and local levels in a divided attention task, the introduction of the local red element speeded responses to local targets but slowed responses when targets appeared at the global level. Experiment 2 used a selective attention task in which subjects responded only to the local or the global shapes across a block of trials. Under these circumstances, the local red element reduced global-to-local interference in addition to speeding local responses. The results suggest that the efficiency with which local elements are segmented and selected affects responses to local aspects of hierarchical patterns; furthermore, the effect of local pop-out on global processing is contingent on top-down attentional control settings.
The health effects of ecstasy: a literature review
Authors: GOWING LINDA R. , HENRY‐EDWARDS SUSAN M. , IRVINE RODNEY J. , ALI ROBERT L.
Published: July 2002

Ecstasy (3,4‐methylenedioxymethamphetamine, MDMA) is the third most used illicit drug, after cannabis and amphetamines. There has been considerable interest in the adverse effects of use, with particular attention given to a small number of deaths related to ecstasy use, and the neurotoxic effects of MDMA. This paper reviews case reports of adverse effects attributed to ecstasy use, and the findings of animal and human studies, so as to identify the health effects of ecstasy use, and factors contributing to their occurrence. The incidence of serious acute adverse events related to ecstasy is low. It is the unpredictability of those adverse events and the risk of mortality and substantial morbidity that makes the health consequences of ecstasy significant. Hyperthermia and hyponatraemia are the most significant acute adverse effects, and can occur even when MDMA is the only drug used. Ecstasy users should be aware of the importance of controlling body temperature and fluid intake, early signs of adverse effects, and the need to seek medical assistance promptly. Neurotoxicity is potentially the most significant long‐term effect of ecstasy. The clinical implications of neurotoxicity are uncertain at this time, but short‐term memory impairment may be significant.
Genetics of the Risk for Alcoholism
Authors: Schuckit Marc A.
Published: July 2002

This paper reviews the literature on the importance of genetic influences in the development of alcohol abuse and dependence (alcoholism). The alcohol use disorders are fairly typical of most complex genetic conditions in that multiple genetic influences combine together to explain approximately 40% to 60% of the risk. One useful approach for identifying specific genes related to alcoholism involves identifying a population in which known genetic factors are controlled and using genome scan and/or case‐control, association approaches to search for specific genes. Several characteristics, or endophenotypes, have been identified as both genetically influenced and contributing toward the risk for alcoholism, including alcohol‐metabolizing enzymes, the low level of response to alcohol, and electrophysiological measures. The potential importance of each of these characteristics is reviewed, and data relating to the search for specific genetic material for each endophenotype are presented. These findings are placed in the perspective of the impact that they are likely to have on both prevention and treatment efforts in the alcohol field.
Appropriate Exposure Routes and Doses in Studies Designed to Assess Developmental Toxicity: A Case Study of Inorganic Arsenic
Authors: Jacobson Catherine F. , Stump Donald G. , Nemec Mark D. , Holson Joseph F. , DeSesso John M.
Published: July 2002

Assessment of risks to human health from chemical agents is a complex process that requires the assembly, careful analysis, and integration of human and animal data collected from studies performed at different times, for disparate purposes, and under varying conditions. The application of risk assessment methods to data without consideration of the relevance of critical experimental parameters such as route of exposure or magnitude of dose can lead to specious determinations of the risk posed by exposure to environmental agents. A case study of the purported risk of developmental toxicity from inorganic arsenic is presented to illustrate (1) the nature of the problem, (2) how extant data from all studies are useful, (3) how appropriately designed modern studies can clarify the situation, and (4) how conflicted data should be evaluated in terms of appropriateness for use in risk assessment.
Mdm12p, a Component Required for Mitochondrial Inheritance That Is Conserved between Budding and Fission Yeast
Authors: Berger Karen H. , Sogo L. Farah , Yaffe Michael P.
Published: July 2002

Saccharomyces cerevisiae cells lacking the MDM12 gene product display temperature-sensitive growth and possess abnormally large, round mitochondria that are defective for inheritance by daughter buds. Analysis of the wild-type MDM12 gene revealed its product to be a 31-kD polypeptide that is homologous to a protein of the fission yeast Schizosaccharomyces pombe. When expressed in S. cerevisiae, the S. pombe Mdm12p homolog conferred a dominant-negative phenotype of giant mitochondria and aberrant mitochondrial distribution, suggesting partial functional conservation of Mdm12p activity between budding and fission yeast. The S. cerevisiae Mdm12p was localized by indirect immunofluorescence microscopy and by subcellular fractionation and immunodetection to the mitochondrial outer membrane and displayed biochemical properties of an integral membrane protein. Mdm12p is the third mitochondrial outer membrane protein required for normal mitochondrial morphology and distribution to be identified in S. cerevisiae and the first such mitochondrial component that is conserved between two different species.
Cross-Talk between Adherens Junctions and Desmosomes Depends on Plakoglobin
Authors: Lewis Jani E. , Wahl James K. , Sass Kristin M. , Jensen Pamela J. , Johnson Keith R. , Wheelock Margaret J.
Published: July 2002

Squamous epithelial cells have both adherens junctions and desmosomes. The ability of these cells to organize the desmosomal proteins into a functional structure depends upon their ability first to organize an adherens junction. Since the adherens junction and the desmosome are separate structures with different molecular make up, it is not immediately obvious why formation of an adherens junction is a prerequisite for the formation of a desmosome. The adherens junction is composed of a transmembrane classical cadherin (E-cadherin and/or P-cadherin in squamous epithelial cells) linked to either β-catenin or plakoglobin, which is linked to α-catenin, which is linked to the actin cytoskeleton. The desmosome is composed of transmembrane proteins of the broad cadherin family (desmogleins and desmocollins) that are linked to the intermediate filament cytoskeleton, presumably through plakoglobin and desmoplakin. To begin to study the role of adherens junctions in the assembly of desmosomes, we produced an epithelial cell line that does not express classical cadherins and hence is unable to organize desmosomes, even though it retains the requisite desmosomal components. Transfection of E-cadherin and/or P-cadherin into this cell line did not restore the ability to organize desmosomes; however, overexpression of plakoglobin, along with E-cadherin, did permit desmosome organization. These data suggest that plakoglobin, which is the only known common component to both adherens junctions and desmosomes, must be linked to E-cadherin in the adherens junction before the cell can begin to assemble desmosomal components at regions of cell–cell contact. Although adherens junctions can form in the absence of plakoglobin, making use only of β-catenin, such junctions cannot support the formation of desmosomes. Thus, we speculate that plakoglobin plays a signaling role in desmosome organization.
High Mobility Group 1 Protein Is Not Stably Associated with the Chromosomes of Somatic Cells
Authors: Falciola Luca , Spada Fabio , Calogero Sabina , Längst Gernot , Voit Renate , Grummt Ingrid , Bianchi Marco E.
Published: July 2002

High mobility group 1 (HMG1) protein is an abundant and conserved component of vertebrate nuclei and has been proposed to play a structural role in chromatin organization, possibly similar to that of histone H1. However, a high abundance of HMG1 had also been reported in the cytoplasm and on the surface of mammalian cells. We conclusively show that HMG1 is a nuclear protein, since several different anti-HMG1 antibodies stain the nucleoplasm of cultured cells, and epitope-tagged HMG1 is localized in the nucleus only. The protein is excluded from nucleoli and is not associated to specific nuclear structures but rather appears to be uniformly distributed. HMG1 can bind in vitro to reconstituted core nucleosomes but is not stably associated to chromatin in live cells. At metaphase, HMG1 is detached from condensed chromosomes, contrary to histone H1. During interphase, HMG1 readily diffuses out of nuclei after permeabilization of the nuclear membranes with detergents, whereas histone H1 remains associated to chromatin. These properties exclude a shared function for HMG1 and H1 in differentiated cells, in spite of their similar biochemical properties. HMG1 may be stably associated only to a very minor population of nucleosomes or may interact transiently with nucleosomes during dynamic processes of chromatin remodeling.
p53/58 Binds COPI and Is Required for Selective Transport through the Early Secretory Pathway
Authors: Tisdale Ellen J. , Plutner Helen , Matteson Jeanne , Balch William E.
Published: July 2002

p53/58 is a transmembrane protein that continuously recycles between the ER and pre-Golgi intermediates composed of vesicular-tubular clusters (VTCs) found in the cell periphery and at the cis face of the Golgi complex. We have generated an antibody that uniquely recognizes the p53/58 cytoplasmic tail. Here we present evidence that this antibody arrests the anterograde transport of vesicular stomatitis virus glycoprotein and leads to the accumulation of p58 in preGolgi intermediates. Consistent with a role for the KKXX retrieval motif found at the cytoplasmic carboxyl terminus of p53/58 in retrograde traffic, inhibition of transport through VTCs correlates with the ability of the antibody to block recruitment of COPI coats to the p53/58 cytoplasmic tail and to p53/58-containing membranes. We suggest that p53/58 function may be required for the coupled exchange of COPII for COPI coats during segregation of anterograde and retrograde transported proteins.
The Small GTPases Rho and Rac Are Required for the Establishment of Cadherin-dependent Cell–Cell Contacts
Authors: Braga Vania M.M. , Machesky Laura M. , Hall Alan , Hotchin Neil A.
Published: July 2002

Cadherins are calcium-dependent cell–cell adhesion molecules that require the interaction of the cytoplasmic tail with the actin cytoskeleton for adhesive activity. Because of the functional relationship between cadherin receptors and actin filament organization, we investigated whether members of the Rho family of small GTPases are necessary for cadherin adhesion. In fibroblasts, the Rho family members Rho and Rac regulate actin polymerization to produce stress fibers and lamellipodia, respectively. In epithelial cells, we demonstrate that Rho and Rac are required for the establishment of cadherin-mediated cell–cell adhesion and the actin reorganization necessary to stabilize the receptors at sites of intercellular junctions. Blocking endogenous Rho or Rac selectively removed cadherin complexes from junctions induced for up to 3 h, while desmosomes were not perturbed. In addition, withdrawal of cadherins from intercellular junctions temporally precedes the removal of CD44 and integrins, other microfilament-associated receptors. Our data showed that the concerted action of Rho and Rac modulate the establishment of cadherin adhesion: a constitutively active form of Rac was not sufficient to stabilize cadherindependent cell–cell contacts when endogenous Rho was inhibited. Upon induction of calcium-dependent intercellular adhesion, there was a rapid accumulation of actin at sites of cell–cell contacts, which was prevented by blocking cadherin function, Rho or Rac activity. However, if cadherin complexes are clustered by specific antibodies attached to beads, actin recruitment to the receptors was perturbed by inhibiting Rac but not Rho. Our results provide new insights into the role of the small GTPases in the cadherin-dependent cell– cell contact formation and the remodelling of actin filaments in epithelial cells.
Human Bcl-2 Reverses Survival Defects in Yeast Lacking Superoxide Dismutase and Delays Death of Wild-Type Yeast
Authors: Longo Valter D. , Ellerby Lisa M. , Bredesen Dale E. , Valentine Joan S. , Gralla Edith B.
Published: July 2002

We expressed the human anti-apoptotic protein, Bcl-2, in Saccharomyces cerevisiae to investigate its effects on antioxidant protection and stationary phase survival. Yeast lacking copper-zinc superoxide dismutase (sod1Δ) show a profound defect in entry into and survival during stationary phase even under conditions optimal for survival of wild-type strains (incubation in water after stationary phase is reached). Expression of Bcl-2 in the sod1Δ strain caused a large improvement in viability at entry into stationary phase, as well as increased resistance to 100% oxygen and increased catalase activity. In addition, Bcl-2 expression reduced mutation frequency in both wild-type and sod1Δ strains. In another set of experiments, wild-type yeast incubated in expired minimal medium instead of water lost viability quickly; expression of Bcl-2 significantly delayed this stationary phase death. Our results demonstrate that Bcl-2 has activities in yeast that are similar to activities it is known to possess in mammalian cells: (a) stimulation of antioxidant protection and (b) delay of processes leading to cell death.
A New Function for the LDL Receptor: Transcytosis of LDL across the Blood–Brain Barrier
Authors: Dehouck Bénédicte , Fenart Laurence , Dehouck Marie-Pierre , Pierce Annick , Torpier Gérard , Cecchelli Roméo
Published: July 2002

Lipoprotein transport across the blood–brain barrier (BBB) is of critical importance for the delivery of essential lipids to the brain cells. The occurrence of a low density lipoprotein (LDL) receptor on the BBB has recently been demonstrated. To examine further the function of this receptor, we have shown using an in vitro model of the BBB, that in contrast to acetylated LDL, which does not cross the BBB, LDL is specifically transcytosed across the monolayer. The C7 monoclonal antibody, known to interact with the LDL receptor-binding domain, totally blocked the transcytosis of LDL, suggesting that the transcytosis is mediated by the receptor. Furthermore, we have shown that cholesterol-depleted astrocytes upregulate the expression of the LDL receptor at the BBB. Under these conditions, we observed that the LDL transcytosis parallels the increase in the LDL receptor, indicating once more that the LDL is transcytosed by a receptor-mediated mechanism. The nondegradation of the LDL during the transcytosis indicates that the transcytotic pathway in brain capillary endothelial cells is different from the LDL receptor classical pathway. The switch between a recycling receptor to a transcytotic receptor cannot be explained by a modification of the internalization signals of the cytoplasmic domain of the receptor, since we have shown that LDL receptor messengers in growing brain capillary ECs (recycling LDL receptor) or differentiated cells (transcytotic receptor) are 100% identical, but we cannot exclude posttranslational modifications of the cytoplasmic domain, as demonstrated for the polymeric immunoglobulin receptor. Preliminary studies suggest that caveolae are likely to be involved in the potential transport of LDL from the blood to the brain.
A Septin-based Hierarchy of Proteins Required for Localized Deposition of Chitin in the Saccharomyces cerevisiae Cell Wall
Authors: DeMarini Douglas J. , Adams Alison E.M. , Fares Hanna , Virgilio Claudio De , Valle Giorgio , Chuang John S. , Pringle John R.
Published: July 2002

Just before bud emergence, a Saccharomyces cerevisiae cell forms a ring of chitin in its cell wall; this ring remains at the base of the bud as the bud grows and ultimately forms part of the bud scar marking the division site on the mother cell. The chitin ring seems to be formed largely or entirely by chitin synthase III, one of the three known chitin synthases in S. cerevisiae. The chitin ring does not form normally in temperature-sensitive mutants defective in any of four septins, a family of proteins that are constituents of the “neck filaments” that lie immediately subjacent to the plasma membrane in the mother-bud neck. In addition, a synthetic-lethal interaction was found between cdc12-5, a temperature-sensitive septin mutation, and a mutant allele of CHS4, which encodes an activator of chitin synthase III. Two-hybrid analysis revealed no direct interaction between the septins and Chs4p but identified a novel gene, BNI4, whose product interacts both with Chs4p and Cdc10p and with one of the septins, Cdc10p; this analysis also revealed an interaction between Chs4p and Chs3p, the catalytic subunit of chitin synthase III. Bni4p has no known homologues; it contains a predicted coiled-coil domain, but no other recognizable motifs. Deletion of BNI4 is not lethal, but causes delocalization of chitin deposition and aberrant cellular morphology. Overexpression of Bni4p also causes delocalization of chitin deposition and produces a cellular morphology similar to that of septin mutants. Immunolocalization experiments show that Bni4p localizes to a ring at the mother-bud neck that lies predominantly on the mother-cell side (corresponding to the predominant site of chitin deposition). This localization depends on the septins but not on Chs4p or Chs3p. A GFP-Chs4p fusion protein also localizes to a ring at the mother-bud neck on the mother-cell side. This localization is dependent on the septins, Bni4p, and Chs3p. Chs3p, whose normal localization is similar to that of Chs4p, does not localize properly in bni4, chs4, or septin mutant strains or in strains that accumulate excess Bni4p. In contrast, localization of the septins is essentially normal in bni4, chs4, and chs3 mutant strains and in strains that accumulate excess Bni4p. Taken together, these results suggest that the normal localization of chitin synthase III activity is achieved by assembly of a complex in which Chs3p is linked to the septins via Chs4p and Bni4p.
In Vitro Degradation of Endothelial Catenins by a Neutrophil Protease
Authors: Moll Thomas , Dejana Elisabetta , Vestweber Dietmar
Published: July 2002

It has been recently proposed that adhesion of polymorphonuclear cells (PMNs) to human umbilical vein endothelial cells leads to the disorganization of the vascular endothelial cadherin–dependent endothelial adherens junctions. Combined immunofluorescence and biochemical data suggested that after adhesion of PMNs to the endothelial cell surface, β-catenin, as well as plakoglobin was lost from the cadherin/catenin complex and from total cell lysates. In this study we present data that strongly suggest that the adhesion-dependent disappearance of endothelial catenins is not mediated by a leukocyte to endothelium signaling event, but is due to the activity of a neutrophil protease that is released upon detergent lysis of the cells.
Mice That Lack Thrombospondin 2 Display Connective Tissue Abnormalities That Are Associated with Disordered Collagen Fibrillogenesis, an Increased Vascular Density, and a Bleeding Diathesis
Authors: Kyriakides Themis R. , Zhu Yu-Hong , Smith Lynne T. , Bain Steven D. , Yang Zhantao , Lin Ming T. , Danielson Keith G. , Iozzo Renato V. , LaMarca Mary , McKinney Cindy E. , Ginns Edward I. , Bornstein Paul
Published: July 2002

Thrombospondin (TSP) 2, and its close relative TSP1, are extracellular proteins whose functions are complex, poorly understood, and controversial. In an attempt to determine the function of TSP2, we disrupted the Thbs2 gene by homologous recombination in embryonic stem cells, and generated TSP2-null mice by blastocyst injection and appropriate breeding of mutant animals. Thbs2−/− mice were produced with the expected Mendelian frequency, appeared overtly normal, and were fertile. However, on closer examination, these mice displayed a wide variety of abnormalities. Collagen fiber patterns in skin were disordered, and abnormally large fibrils with irregular contours were observed by electron microscopy in both skin and tendon. As a functional correlate of these findings, the skin was fragile and had reduced tensile strength, and the tail was unusually flexible. Mutant skin fibroblasts were defective in attachment to a substratum. An increase in total density and in cortical thickness of long bones was documented by histology and quantitative computer tomography. Mutant mice also manifested an abnormal bleeding time, and histologic surveys of mouse tissues, stained with an antibody to von Willebrand factor, showed a significant increase in blood vessels. The basis for the unusual phenotype of the TSP2-null mouse could derive from the structural role that TSP2 might play in collagen fibrillogenesis in skin and tendon. However, it seems likely that some of the diverse manifestations of this genetic disorder result from the ability of TSP2 to modulate the cell surface properties of mesenchymal cells, and thus, to affect cell functions such as adhesion and migration.
Selective Entrapment of Extrachromosomally Amplified DNA by Nuclear Budding and Micronucleation during S Phase
Authors: Shimizu Noriaki , Itoh Nobuo , Utiyama Hiroyasu , Wahl Geoffrey M.
Published: July 2002

Acentric, autonomously replicating extrachromosomal structures called double-minute chromosomes (DMs) frequently mediate oncogene amplification in human tumors. We show that DMs can be removed from the nucleus by a novel micronucleation mechanism that is initiated by budding of the nuclear membrane during S phase. DMs containing c-myc oncogenes in a colon cancer cell line localized to and replicated at the nuclear periphery. Replication inhibitors increased micronucleation; cell synchronization and bromodeoxyuridine–pulse labeling demonstrated de novo formation of buds and micronuclei during S phase. The frequencies of S-phase nuclear budding and micronucleation were increased dramatically in normal human cells by inactivating p53, suggesting that an S-phase function of p53 minimizes the probability of producing the broken chromosome fragments that induce budding and micronucleation. These data have implications for understanding the behavior of acentric DNA in interphase nuclei and for developing chemotherapeutic strategies based on this new mechanism for DM elimination.
Anaphase A Chromosome Movement and Poleward Spindle Microtubule Flux Occur At Similar Rates in Xenopus Extract Spindles
Authors: Desai Arshad , Maddox Paul S. , Mitchison Timothy J. , Salmon E.D.
Published: July 2002

We have used local fluorescence photoactivation to mark the lattice of spindle microtubules during anaphase A in Xenopus extract spindles. We find that both poleward spindle microtubule flux and anaphase A chromosome movement occur at similar rates (∼2 μm/min). This result suggests that poleward microtubule flux, coupled to microtubule depolymerization near the spindle poles, is the predominant mechanism for anaphase A in Xenopus egg extracts. In contrast, in vertebrate somatic cells a “Pacman” kinetochore mechanism, coupled to microtubule depolymerization near the kinetochore, predominates during anaphase A. Consistent with the conclusion from fluorescence photoactivation analysis, both anaphase A chromosome movement and poleward spindle microtubule flux respond similarly to pharmacological perturbations in Xenopus extracts. Furthermore, the pharmacological profile of anaphase A in Xenopus extracts differs from the previously established profile for anaphase A in vertebrate somatic cells. The difference between these profiles is consistent with poleward microtubule flux playing the predominant role in anaphase chromosome movement in Xenopus extracts, but not in vertebrate somatic cells. We discuss the possible biological implications of the existence of two distinct anaphase A mechanisms and their differential contributions to poleward chromosome movement in different cell types.
The Influence of Huntingtin Protein Size on Nuclear Localization and Cellular Toxicity
Authors: Hackam Abigail S. , Singaraja Roshni , Wellington Cheryl L. , Metzler Martina , McCutcheon Krista , Zhang Taiqi , Kalchman Michael , Hayden Michael R.
Published: July 2002

Huntington disease is an autosomal dominant neurodegenerative disorder caused by the pathological expansion of a polyglutamine tract. In this study we directly assess the influence of protein size on the formation and subcellular localization of huntingtin aggregates. We have created numerous deletion constructs expressing successively smaller fragments of huntingtin and show that these smaller proteins containing 128 glutamines form both intranuclear and perinuclear aggregates. In contrast, larger NH2-terminal fragments of huntingtin proteins with 128 glutamines form exclusively perinuclear aggregates. These aggregates can form in the absence of endogenous huntingtin. Furthermore, expression of mutant huntingtin results in increased susceptibility to apoptotic stress that is greater with decreasing protein length and increasing polyglutamine size. As both intranuclear and perinuclear aggregates are clearly associated with increased cellular toxicity, this supports an important role for toxic polyglutamine-containing fragments forming aggregates and playing a key role in the pathogenesis of Huntington disease.