bluuebunny/crossref_metadata_2025 huggingface dataset:
Kainate receptor-mediated synaptic currents in cerebellar Golgi cells are not shaped by diffusion of glutamate
Authors: Bureau Ingrid , Dieudonné Stéphane , Coussen Françoise , Mulle Christophe
Published: July 2002

We report the presence of kainate receptors (KARs) in cerebellar Golgi cells of wild-type but not GluR6-deficient mice. Parallel fiber stimulation activates KAR-mediated synaptic currents [KAR-excitatory postsynaptic currents (EPSCs)] of small amplitude. KAR-EPSCs greatly differ from synaptic currents mediated by α-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors (AMPAR-EPSCs) at the same synapse. KAR-EPSCs display slow rise and decay time and summate in response to a train of stimulations. By using PDA, a low-affinity competitive antagonist and agents that modify the clearance of glutamate, we show that these properties cannot be explained by diffusion of glutamate outside of the synaptic cleft and activation of extrasynaptic KARs. These data suggest that the slow kinetic of KAR-EPSCs is due to intrinsic properties of KARs being localized at postsynaptic sites. The contrasting properties of KAR- and AMPAR-EPSCs in terms of kinetics and summation offer the possibility for a glutamatergic synapse to integrate excitatory inputs over two different time scales.
Structure and function of pectic enzymes: Virulence factors of plant pathogens
Authors: Herron Steven R. , Benen Jacques A. E. , Scavetta Robert D. , Visser Jaap , Jurnak Frances
Published: July 2002

The structure and function of
Erwinia chrysanthemi
pectate lysase C, a plant virulence factor, is reviewed to illustrate
one mechanism of pathogenesis at the molecular level. Current
investigative topics are discussed in this paper.
Rice dwarf mutant
d1
, which is defective in the α subunit of the heterotrimeric G protein, affects gibberellin signal transduction
Authors: Ueguchi-Tanaka Miyako , Fujisawa Yukiko , Kobayashi Masatomo , Ashikari Motoyuki , Iwasaki Yukimoto , Kitano Hidemi , Matsuoka Makoto
Published: July 2002

Previously, we reported that the rice dwarf mutant,
d1
, is defective in the α subunit of the
heterotrimeric G protein (Gα). In the present study, gibberellin (GA)
signaling in
d1
and the role of the Gα protein in the
GA-signaling pathway were investigated. Compared with the wild type, GA
induction of α-amylase activity in aleurone cells of
d1
was greatly reduced. Relative to the wild type, the
GA
3
-treated aleurone layer of
d1
had lower
expression of
Ramy1A
, which encodes α-amylase, and
OsGAMYB
, which encodes a GA-inducible transcriptional
factor, and no increase in expression of
Ca
2
+
-ATPase
. However, in the presence of high GA
concentrations, α-amylase induction occurred even in
d1
. The GA sensitivity of second leaf sheath elongation
in
d1
was similar to that of the wild type in terms of
dose responsiveness, but the response of internode elongation to GA was
much lower in
d1
. Furthermore,
Os20ox
expression was up-regulated, and the GA content was elevated in the
stunted internodes of
d1
. All these results suggest that
d1
affects a part of the GA-signaling pathway, namely
the induction of α-amylase in the aleurone layer and internode
elongation. In addition, a double mutant between
d1
and
another GA-signaling mutant,
slr
, revealed that
SLR
is epistatic to the
D1
, supporting
that the Gα protein is involved in GA signaling. However, the data
also provide evidence for the presence of an alternative GA-signaling
pathway that does not involve the Gα protein. It is proposed that GA
signaling via the Gα protein may be more sensitive than that of the
alternative pathway, as indicated by the low GA responsiveness of this
Gα-independent pathway.
Role of Syk in B-cell development and antigen-receptor signaling
Authors: Cornall Richard J. , Cheng Alex M. , Pawson Tony , Goodnow Christopher C.
Published: July 2002

Antigen receptors (BCRs) on developing B lymphocytes play two opposing roles—promoting survival of cells that may later bind a foreign antigen and inhibiting survival of cells that bind too strongly to self-antigens. It is not known how these opposing outcomes are signaled by BCRs on immature B cells. Here we analyze the effect of a null mutation in the Syk tyrosine kinase on maturing B cells displaying a transgene-encoded BCR that binds hen egg lysozyme (HEL). In the absence of HEL antigen, HEL-specific BCRs are expressed normally on the surface of Syk-deficient immature B-lineage cells, but this fails to promote maturation beyond the earliest stages of B-lineage commitment. Binding of HEL antigen, nevertheless, triggers phosphorylation of CD79α/β BCR subunits and modulation of receptors from the surface in Syk-deficient cells, but it cannot induce an intracellular calcium response. Continuous binding of low- or high-avidity forms of HEL, expressed as self-antigens, fails to restore the signal needed for maturation. Compared with the effects in the same system of null mutations in other BCR signaling elements, such as CD45 and Lyn kinase, these results indicate that Syk is essential for transmitting a signal that initiates the program of B-lymphocyte maturation.
Cardiolipin is a normal component of human plasma lipoproteins
Authors: Deguchi Hiroshi , Fernández José A. , Hackeng Tilman M. , Banka Carole L. , Griffin John H.
Published: July 2002

Anticardiolipin (anti-CL) antibodies, diagnostic for antiphospholipid antibody syndrome, are associated with increased risks of venous and arterial thrombosis. Because CL selectively enhances activated protein C/protein S-dependent anticoagulant activities in purified systems and because CL is not known to be a normal plasma component, we searched for CL in plasma. Plasma lipid extracts [chloroform/methanol (2:1, vol/vol)] were subjected to analyses by using TLC, analytical HPLC, and MS. A plasma lipid component was purified that was indistinguishable from reference CL (M:1448). When CL in 40 fasting plasma lipid extracts (20 males, 20 females) was quantitated by using HPLC, CL (mean ± SD) was 14.9 ± 3.7 μg/ml (range 9.1 to 24.2) and CL was not correlated with phosphatidylserine (3.8 ± 1.7 μg/ml), phosphatidylethanolamine (64 ± 20 μg/ml), or choline-containing phospholipid (1,580 ± 280 μg/ml). Based on studies of fasting blood donors, CL (≥94%) was recovered in very low density, low density, and high density lipoproteins (11 ± 5.3%, 67 ± 11.0%, and 17 ± 10%, respectively), showing that the majority of plasma CL (67%) is in low density lipoprotein. Analysis of relative phospholipid contents of lipoproteins indicated that high density lipoprotein is selectively enriched in CL and phosphatidylethanolamine. These results shows that CL is a normal plasma component and suggest that the epitopes of antiphospholipid antibodies could include CL or oxidized CL in lipoproteins or in complexes with plasma proteins (e.g., β
2
-glycoprotein I, prothrombin, protein C, or protein S) or with platelet or endothelial surface proteins.
Deregulated expression of insulin-like growth factor 1 in prostate epithelium leads to neoplasia in transgenic mice
Authors: DiGiovanni John , Kiguchi Kaoru , Frijhoff Anita , Wilker Eric , Bol David K. , Beltrán Linda , Moats Samantha , Ramirez Angel , Jorcano José , Conti Claudio
Published: July 2002

Transgenic mice expressing human insulin-like growth factor 1 (IGF-1) in basal epithelial cells of prostate have been characterized. Transgene expression led to activation of the IGF-1 receptor and spontaneous tumorigenesis in prostate epithelium. Hyperplasia was evident in these mice by 2–3 months of age. Atypical hyperplasias and prostatic intraepithelial neoplasia were evident by 6–7 months of age. Well differentiated adenocarcinomas appeared in mice 6 months or older. Less differentiated tumors, diagnosed as small cell carcinomas, were also observed in two of the older mice. Both lobes of the mouse prostate gland (dorsolateral and ventral) presented preneoplastic and neoplastic changes. The incidence of tumors in mice ≥6 months of age (38 mice total) was 50%. The development of neoplasia in these transgenic mice appeared to follow a stepwise progression through early preneoplastic changes that ultimately culminated in frank neoplasia. These mice offer an animal model for prostate cancer that will allow study of the stepwise development of this disease and the mechanism(s) whereby IGF-1 mediates this process.
Distinct roles of the homeotic genes Ubx and abd-A in beetle embryonic abdominal appendage development
Authors: Lewis David L. , DeCamillis Mark , Bennett Randy L.
Published: July 2002

Insects are easily distinguishable by the absence of legs on the
adult abdomen. Studies performed on the Dipteran,
Drosophila
melanogaster
, indicate that this is because of the repressive
effects of the homeotic genes
Ultrabithorax
(
Ubx
) and
abdominal-A
(
abd-A
) on the limb promoting gene
Distal-less
(
Dll
) during embryonic
development. However, in many species appendage-like structures are
present on abdominal segments in embryonic and juvenile stages. Here,
by using classical genetics and double-stranded RNA-mediated gene
silencing in the red flour beetle,
Tribolium castaneum
,
a species that develops an appendage on the first abdominal segment, we
investigate the roles of Ubx and Abd-A in abdominal limb development.
We find that in
Tribolium
, Abd-A, but not Ubx, represses
early expression of Dll in the embryonic abdomen. Ubx appears to modify
the A1 appendage. This difference in the activities of Abd-A and Ubx is
critical for proper development of this appendage. We suggest that an
ancestral role of Abd-A in insect abdominal appendage development was
in the repression of Dll initiation and that of Ubx was in modulation
of abdominal appendage morphology.
Identification of two short internal ribosome entry sites selected from libraries of random oligonucleotides
Authors: Owens Geoffrey C. , Chappell Stephen A. , Mauro Vincent P. , Edelman Gerald M.
Published: July 2002

Sequences that control translation of mRNA may play critical roles
in regulating protein levels. One such element is the internal ribosome
entry site (IRES). We previously showed that a 9-nt segment in the 5′
leader sequence of the mRNA encoding Gtx homeodomain protein could
function as an IRES. To identify other short sequences with similar
properties, we designed a selection procedure that uses a retroviral
vector to express dicistronic mRNAs encoding enhanced green and cyan
fluorescent proteins as the first and second cistrons, respectively.
Expression of the second cistron was dependent upon the intercistronic
sequences and was indicative of IRES activity. B104 cells were infected
with two retroviral libraries that contained random sequences of 9 or
18 nt in the intercistronic region. Cells expressing both cistrons were
sorted, and sequences recovered from selected cells were reassayed for
IRES activity in a dual luciferase dicistronic mRNA. Two novel IRESes
were identified by this procedure, and both contained segments with
complementarity to 18S rRNA. When multiple copies of either segment
were linked together, IRES activities were dramatically enhanced.
Moreover, these synthetic IRESes were differentially active in various
cell types. These properties are similar to those of the previously
identified 9-nt IRES module from Gtx mRNA. These results provide
further evidence that short nucleotide sequences can function as IRESes
and support the idea that some cellular IRESes may be composed of
shorter functional modules. The ability to identify IRES modules with
specific expression properties may be useful in the design of vectors
for biotechnology and gene therapy.
Cross‐national changes in time‐use: some sociological (hi) stories re‐examined
Authors: Sullivan Oriel , Gershuny Jonathan
Published: July 2002

The paper addresses some macro‐sociological questions about changes in broad categories of time‐use. The focus is on large‐scale cross‐national time trends from developed countries in paid and unpaid work, and leisure. Reference is made to some well‐known sociological and historical accounts of such change, and to the fact that time‐use diary data has only relatively recently become available for analysing trends over time. The data used are drawn from a comparative cross‐time data archive held by the Institute for Social and Economic Research at Essex University, comprising successive time‐use diary surveys from a range of industrialized countries collected from the 1960s to the 1990s. The time use evidence suggests relative stability in the balance between work and leisure time over the period covered by the analyses. Some alternative explanations are advanced for why there seems to be a gap between this evidence and, on the one hand, the burgeoning literature in both academic and popular media addressing the ‘time famine’ and, on the other, people's professed experience of what is happening to their time.
Divided attention in music
Authors: Bigand E. , McAdams S. , Forêt S.
Published: July 2002

Two models have been advanced to account for the apparent ease with which attention can be divided in music: a “divided attention” model postulates that listeners effectively manage to follow two or more melodic lines played simultaneously. According to a “figure‐ground model,” the harmonic coherence of Western polyphonies allows a listener to focus on one melody while staying aware of the other melody, which acts as a background. This figure‐ground processing compensates for the inability to divide attention. The present study was designed to further investigate these two models. Participants were required to detect melodic errors in two familiar nursery tunes played simultaneously an octave apart. The divided‐attention model predicted that this task would be easily performed by participants, irrespective of the key of the nursery tune. The figure‐ground model predicted better performance when the keys of the tunes were identical or closely related. None of these predictions was fully supported by the data, leading us to propose a new “integrative model” of listening to polyphonic music.
Phonological Short‐term Memory and Foreign Language Learning
Authors: Masoura Elvira V. , Gathercole Susan E.
Published: July 2002

This study investigated links between short‐term memory skills and children's abilities to learn the vocabulary of a foreign language taught in school. Forty‐five Greek children who were learning English as a foreign language were assessed on their short‐term memory in both languages, and on their knowledge of both native and foreign vocabulary. Knowledge of native and foreign vocabulary shared highly significant associations with the phonological short‐term memory measures. However, vocabulary scores in the two languages shared a close relationship that could not be explained exclusively in terms of phonological loop capacity. Implications of these findings for theoretical accounts of how words are learned in the native and foreign language are considered.
Knowledge and Urban Economic Development: An Evolutionary Perspective
Authors: Lambooy Jan G.
Published: July 2002

Knowledge is considered as a main source of economic growth for nations and regions. It is the basis for technological development and for increasing the productivity of capital and labour. This paper explores the question whether urban regions offer effective contexts for the development of knowledge. Actors and selection environments have an interactive relation in which actors learn in a cumulative way. In this paper, the primary focus is on entrepreneurs as actors in an evolutionary process where three kinds of competency enable them to compete successfully: cognitive, innovative and organisational competencies. These are influenced by three kinds of selection environment: institutions, markets and the spatial structure. Not only entrepreneurs, but also governments and scientific organisations can influence the regional innovation system. Although there is a certain path-dependency, in economic development, regional economic growth cannot be completely determined by policy, because it rests on a continuous learning process and on the strategic behaviour of actors. Regional policy, however, can nurture this process, using the opportunities of specialisation.
Rhetoric and Research in Family Literacy
Authors: Hannon Peter
Published: July 2002

ABSTRACT The term ‘family literacy’ now figures prominently in the discourses of early childhood education, literacy and adult education in several English speaking countries. It can refer to a focus for research or to a kind of educational programme. This article distinguishes family literacy programmes which combine adult basic education for parents with early literacy education and parental involvement from other kinds of family literacy programmes and terms the former ‘restricted’ programmes. The rhetoric concerning restricted programmes, and relevant research, is examined in relation to five issues: the usage of the term ‘family literacy’; the targeting of restricted programmes for selected families; the accessibility and take‐up of such programmes; their educational effects; and their socio‐economic effects. Drawing on evidence from Britain and the USA, it is argued that, although rhetoric has sometimes been informed by research, it has also obscured, misinterpreted, ignored and exaggerated research findings. Some implications for policy, practice and research are identified.
Cognitive, Emotional and Behavioural Responses to Students with Emotional and Behavioural Difficulties: A model of decision‐making
Authors: Poulou Maria , Norwich Brahm
Published: July 2002

Abstract Teachers' perceptions and thoughts on educational matters reflect on their teaching decisions and actions to their students in general, and their students with emotional and behavioural difficulties, in particular. A study based on the theoretical assumptions of Weiner's model of attributional theory of motivation, Ajzen & Fishbein's planned behaviour theory and Bandura's social cognitive theory aimed to construct a portrayal of teachers' causal attributions, emotional, cognitive and behavioural responses to students with emotional and behavioural difficulties. The distribution of an Attribution Inventory to 391 elementary teachers revealed that teachers' causal attributions predicted their emotional and cognitive responses, which in turn predicted their intentional behaviour. Intentional behaviour in conjunction with perceptions of effective coping strategies finally predicted their actual behaviour. This study contributes to our understanding of teachers' perceptions of and decisions about emotional and behavioural difficulties, with relevance to teacher trainers and policy‐makers.
Joint attention by mothers and children while using plans
Authors: de la Ossa Jennifer L. , Gauvain Mary
Published: July 2002

This paper reports on the role of joint attentional processes in the development of children’s skill at using pictorial plans to construct objects. Efforts to establish joint attentional focus between mother and child were identified, and the nature and extent of maternal assistance and child involvement during planning were examined. Sixteen 4 to 5-year-old and sixteen 6 to 7-year-old children and their mothers participated in three problem-solving sessions (i.e., child-only pre-test and post-test, and mother-child interaction) that involved constructing a toy from multiple pieces using a pictorial, step-by-step plan. Older children were more planful than younger children during all the planning sessions. Mothers planning with younger children assumed greater responsibility for establishing joint attentional episodes than mothers planning with older children. Results indicate that mothers tailor their guidance on joint planning tasks in relation to developmental needs, and that an important aspect of these efforts is the establishment and maintenance of joint attention.
The quest for quality in early day care and preschool experience continues
Authors: Melhuish Edward C.
Published: July 2002

In research in the areas of day care for young children and preschool education at the end of the second millennium, common themes can be recognised. Early research was primarily concerned with whether children attending institutions (day care or preschool centre), developed differently from those not attending such centres. Later work recognised that day care or preschool experience is not unitary and that the quality or characteristic of experience matters. Yet further research drew attention to the importance of the interaction between home and out of home experience. These have been referred to as the three waves of research. In the 1980s, the proposition emerged that infant day care may be a risk factor for insecure attachment to the mother. In an ideologically and politically sensitive ” eld, the concern raised by this proposition that day care might be bad for infants, led to the funding of one of the largest studies of day care, the NICHD study. The results of this study so far indicate that quality of care is an important aspect of child care experience. This study is likely to be a watershed in that the sample size and detail of data are far greater than preceding studies. The conclusion that quality of experience for young children matters however, sets the agenda for research in the new millennium. Currently, approaches to this issue generally adopt the strategy of using a measure of child care quality and investigating associations with child development outcomes. An alternative approach derives from school effectiveness research. Children from specific centres are followed longitudinally. Their developmental progress is then considered in terms of family factors, type amount and quality of centre experience, and the specific centre attended. In this approach the presence of specific centre effects can be detected so that a specific centre can be identi” ed as associated with a quantifiable positive or negative effect on development. The resulting incongruence between traditional measures of quality and measures derived from developmental effects will require a reformulation of the links between child care characteristics, child experience, and developmental outcomes. As measures of quality become more ”rmly related to developmental outcomes child care research can become more integrated within developmental psychology.
Perspectives on gender development
Authors: Maccoby Eleanor E.
Published: July 2002

Two traditional perspectives on gender development—the socialisation and cognitive perspectives— are reviewed. It is noted that although they deal quite well with individual differences within each sex with regard to degree of sex-typing, they do not offer satisfactory explanations for some of the most robust gender dimorphisms: namely, gender segregation and the divergent patterns of interaction within all-male as compared with all-female dyads or groups. These patterns are briefly summarised, and their similarity to those found in nonhuman primates and other mammals is noted. It is argued that an ethological perspective, and its modern successor the psychobiological perspective, are needed, along with the more traditional perspectives, to provide a comprehensive account of gender development as it occurs in dyads and groups as well as within individual children.
Older and Wiser? Integrating Results on the Relationship between Age and Wisdom-related Performance
Authors: Staudinger Ursula M.
Published: July 2002

Evidence from a psychological approach to the study of wisdom is reviewed with regard to the relationship between age and wisdom. Between 20 and 75 years, age has been demonstrated to show a zero relation with wisdom-related knowledge and judgement. A complex pattern of person characteristics and experiential features have to coalesce in order for wisdom to emerge. However, it is not only growing experience that (on average) comes with age but also decreases in basic intellectual functioning and changes in the personality make-up that seem to undermine rather than facilitate the development of wisdom-related knowledge and judgement. However, there is also some evidence indicating that under certain supportive conditions it may indeed be older people who hold the greatest potential for wisdom.
Genetically Engineered Rodent Models of Mammary Gland Carcinogenesis: An Overview
Authors: Blackshear Pamela E.
Published: July 2002

Breast cancer is a multifactorial disease that develops as a result of interactions among genetic, environmental, and hormonal factors. Multiple genetic derangements are believed to be involved in the pathogenesis of breast cancer, including the inactivation of tumor suppressor genes and/or the disregulation of proto-oncogenes. Age, hormones, and environmental factors further influence these genetic derangements. Spontaneous and chemically induced animal models of breast cancer have been limited in their usefulness. The advent of targeted gene mutations has allowed for a more specifi cexploration of the pathogenesis of breast cancer by creating mouse models that mimic single or multiple gene alterations found in human mammary tumors. The genes targeted in these models include mouse mammary tumor integration sites and genes that encode for growth regulators, signal transduction proteins, cell cycle proteins, and cell matrix proteinases. In this review, I summarize tumor morphology and the relevance of each model to the pathogenesis and progression of human breast cancer. These models have great potential for elucidating the multistep process of mammary gland carcinogenesis and for contributing to the identification of novel therapeutic targets.
Protective Effect of rhIL-11 in a Murine Model of Acetaminophen-Induced Hepatotoxicity
Authors: Trepicchio William L. , Bozza Mary , Bouchard Page , Dorner Andrew J.
Published: July 2002

Acetaminophen intoxication results in hepatotoxicity associated with increased serum concentrations of hepatocellular leakage enzymes such as aspartate aminotransferase, lactate dehydrogenase, and alanine aminotransferase, centrilobular degeneration and necrosis, and activation of Kupffer cells. Recombinant human Interleukin-11 (rhIL-11) downregulates the production of proinfl ammatory mediators from activated macrophages and has direct effects on hepatocyte gene expression. Based on these biological activities of rhIL-11, the effect of pretreatment with rhIL-11 in a murine model of acetaminophen-induce d hepatotoxicity was examined. Administration of 500 ug/kg acetaminophen to B6C3F1 mice resulted in progressive hepatotoxicity as demonstrated by elevated serum concentration s of hepatocellular leakage enzymes and TNF α and histopathology. Pretreatment with 250 or 500 ug/kg of subcutaneously administered rhIL-11 2 hours before acetaminophen administration reduced serum concentrations of hepatocellular leakage enzyme s and TNF α by 40—50%. This was associated with a statistically significant decreas e in mean severity score for centrilobular hemorrhag e and necrosis from grade 3 to grade 2 for rhIL-11-treated animals compared to vehicle. These results indicate that treatment with rhIL-11 has a protective effect in a model of acetaminophen-induce d liver damage.
Toxicogenomics, Drug Discovery, and the Pathologist
Authors: Boorman Gary A. , Anderson Steven P. , Casey Warren M. , Brown Roger H. , Crosby Lynn M. , Gottschalk K. , Easton Marilyn , Hong Ni  , Morgan Kevin T.
Published: July 2002

The field of toxicogenomics, which currently focuses on the application of large-scale differential gene expression (DGE) data to toxicology, is starting to influence drug discovery and development in the pharmaceutical industry. Toxicological pathologists, who play key roles in the development of therapeutic agents, have much to contribute to DGE studies, especially in the experimental design and interpretation phases. The intelligent application of DGE to drug discovery can reveal the potential for both desired (therapeutic) and undesired (toxic) responses. The pathologist's understanding of anatomic, physiologic, biochemical, immune, and other underlying factors that drive mechanisms of tissue responses to noxious agents turns a bewildering array of gene expression data into focused research programs. The latter process is critical for the successful application of DGE to toxicology. Pattern recognition is a useful first step, but mechanistically based DGE interpretation is where the long-term future of these new technologies lies. Pathologists trained to carry out such interpretations will become important members of the research teams needed to successfully apply these technologies to drug discovery and safety assessment. As a pathologist using DGE, you will need to learn to read DGE data in the same way you learned to read glass slides, patiently and with a desire to learn and, later, to teach. In return, you will gain a greater depth of understanding of cell and tissue function, both in health and disease.
Quality Review Procedures Necessary for Rodent Pathology Databases and Toxicogenomic Studies: The National Toxicology Program Experience
Authors: Boorman Gary A. , Haseman Joseph K. , Waters Michael D. , Hardisty Jerry F. , Sills Robert C.
Published: July 2002

Accuracy of the pathology data is crucial since rodent studies often provide critical data used for setting human chemical exposure standards. Diagnoses represent a judgment on the expected biological behavior of a lesion and peer review can improve diagnostic accuracy and consistency. With the conduct of 500 2-year rodent studies, the National Toxicology Program (NTP) has refined its process for comprehensive review of the pathology data and diagnoses. We have found that careful judgment can improve and simplify the review, whereas simply applying a set review procedure may not assure study quality. The use of reviewing pathologists and pathology peer review groups is a very effective procedure to increase study quality with minimal time and cost. New genomic technology to assess differential gene expression is being used to predict morphological phenotypes such as necrosis, hyperplasia, and neoplasia. The challenge for pathologists is to provide uniform pathology phenotypes that can be correlated with the gene expression changes. The lessons learned in assuring data quality in standard rodent studies also applies to the emerging field of toxicogenomics.
Pulmonary Sequestration with Ectopic Pancreatic Tissue in a Wistar Hannover GALAS Rat
Authors: Takeuchi Yukiko , Yoshida Toshinori , Chiba Yuko , Kuwahara Maki , Maita Keizo , Harada Takanori
Published: July 2002

A multicystic mass of the lung was found in a male 11-week-ol d Wistar Hannover GALAS rat. The cystic mass was located in the region of the right caudal lobe and had no direct communication with the tracheobranchia l tree. Histologically, the pulmonary mass was composed of variably sized cysts containing mucinous material, cellular debris, erythrocytes, and infl ammatory cells. In the septal stroma between the cysts, well-developed elastic and muscular arteries were present and in some areas, ectopic pancreatic tissue was observed. In the rest of the lung lobes, there was no evidence of previous pneumonic events or other respiratory diseases. Based on these findings, the present case was diagnosed as a congenital pulmonary sequestration with ectopic pancreatic tissue.
Causes and correlates of adolescent drug abuse and implications for treatment
Authors: SPOONER CATHERINE
Published: July 2002

On the basis of the literature, the following risk factors for drug abuse by adolescents were identified: biological predisposition to drug abuse; personality traits that reflect a lack of social bonding; a history of low quality and consistency of family management, family communication, family relationships and parental role‐modelling, a history of being abused or neglected; low socio‐economic status; emotional or psychiatric problems; significant stressors and/or inadequate coping skills and social supports; inadequate social skills; history of associating with drug‐using peers, rejection by prosocial peers due to poor social skills; a history of low commitment to education, failure at school; a history of anti‐social behaviour and delinquency and early initiation to drug use. Models for conceptualizing the aetiology of drug abuse are discussed, namely Jessor's problem behaviour syndrome and Rhodes & Jason's Social Stress Model. It is clear that the path to drug abuse is complex, so simple solutions to the problem are unlikely to be effective. [Spooner C. Causes and correlates of adolescent drug abuse and implications for treatment. Drug Alcohol Rev 1999;18:453–475]
Targeting of U2AF65 to Sites of Active Splicing in the Nucleus
Authors: Gama-Carvalho Margarida , Krauss Randy D. , Chiang Lijian , Valcárcel Juan , Green Michael R. , Carmo-Fonseca Maria
Published: July 2002

U2AF65 is an essential splicing factor that promotes binding of U2 small nuclear (sn)RNP at the pre-mRNA branchpoint. Here we describe a novel monoclonal antibody that reacts specifically with U2AF65. Using this antibody, we show that U2AF65 is diffusely distributed in the nucleoplasm with additional concentration in nuclear speckles, which represent subnuclear compartments enriched in splicing snRNPs and other splicing factors. Furthermore, transient expression assays using epitope-tagged deletion mutants of U2AF65 indicate that targeting of the protein to nuclear speckles is not affected by removing either the RNA binding domain, the RS domain, or the region required for interaction with U2AF35. The association of U2AF65 with speckles persists during mitosis, when transcription and splicing are downregulated. Moreover, U2AF65 is localized to nuclear speckles in early G1 cells that were treated with transcription inhibitors during mitosis, suggesting that the localization of U2AF65 in speckles is independent of the presence of pre-mRNA in the nucleus, which is consistent with the idea that speckles represent storage sites for inactive splicing factors. After adenovirus infection, U2AF65 redistributes from the speckles and is prefferentially detected at sites of viral transcription. By combining adenoviral infection with transient expression of deletion mutants, we show a specific requirement of the RS domain for recruitment of U2AF65 to sites of active splicing in the nucleus. This suggests that interactions involving the RS region of U2AF65 may play an important role in targeting this protein to spliceosomes in vivo.
A Metastable Intermediate State of Microtubule Dynamic Instability That Differs Significantly between Plus and Minus Ends
Authors: Tran P.T. , Walker R.A. , Salmon E.D.
Published: July 2002

The current two-state GTP cap model of microtubule dynamic instability proposes that a terminal crown of GTP-tubulin stabilizes the microtubule lattice and promotes elongation while loss of this GTP-tubulin cap converts the microtubule end to shortening. However, when this model was directly tested by using a UV microbeam to sever axoneme-nucleated microtubules and thereby remove the microtubule's GTP cap, severed plus ends rapidly shortened, but severed minus ends immediately resumed elongation (Walker, R.A., S. Inoué, and E.D. Salmon. 1989. J. Cell Biol. 108: 931–937).

To determine if these previous results were dependent on the use of axonemes as seeds or were due to UV damage, or if they instead indicate an intermediate state in cap dynamics, we performed UV cutting of self-assembled microtubules and mechanical cutting of axoneme-nucleated microtubules. These independent methods yielded results consistent with the original work: a significant percentage of severed minus ends are stable after cutting. In additional experiments, we found that the stability of both severed plus and minus ends could be increased by increasing the free tubulin concentration, the solution GTP concentration, or by assembling microtubules with guanylyl-(α,β)-methylene-diphosphonate (GMPCPP).

Our results show that stability of severed ends, particularly minus ends, is not an artifact, but instead reveals the existence of a metastable kinetic intermediate state between the elongation and shortening states of dynamic instability. The kinetic properties of this intermediate state differ between plus and minus ends. We propose a three-state conformational cap model of dynamic instability, which has three structural states and four transition rate constants, and which uses the asymmetry of the tubulin heterodimer to explain many of the differences in dynamic instability at plus and minus ends.
ICAP-1, a Novel β1 Integrin Cytoplasmic Domain–associated Protein, Binds to a Conserved and Functionally Important NPXY Sequence Motif of β1 Integrin
Authors: Chang David D. , Wong Carol , Smith Healy , Liu Jenny
Published: July 2002

The cytoplasmic domains of integrins are essential for cell adhesion. We report identification of a novel protein, ICAP-1 (integrin cytoplasmic domain– associated protein-1), which binds to the β1 integrin cytoplasmic domain. The interaction between ICAP-1 and β1 integrins is highly specific, as demonstrated by the lack of interaction between ICAP-1 and the cytoplasmic domains of other β integrins, and requires a conserved and functionally important NPXY sequence motif found in the COOH-terminal region of the β1 integrin cytoplasmic domain. Mutational studies reveal that Asn and Tyr of the NPXY motif and a Val residue located NH2-terminal to this motif are critical for the ICAP-1 binding. Two isoforms of ICAP-1, a 200–amino acid protein (ICAP-1α) and a shorter 150–amino acid protein (ICAP-1β), derived from alternatively spliced mRNA, are expressed in most cells. ICAP-1α is a phosphoprotein and the extent of its phosphorylation is regulated by the cell–matrix interaction. First, an enhancement of ICAP-1α phosphorylation is observed when cells were plated on fibronectin-coated but not on nonspecific poly-l-lysine–coated surface. Second, the expression of a constitutively activated RhoA protein that disrupts the cell–matrix interaction results in dephosphorylation of ICAP-1α. The regulation of ICAP-1α phosphorylation by the cell–matrix interaction suggests an important role of ICAP-1 during integrin-dependent cell adhesion.
Animal Models for Muscular Dystrophy Show Different Patterns of Sarcolemmal Disruption
Authors: Straub Volker , Rafael Jill A. , Chamberlain Jeffrey S. , Campbell Kevin P.
Published: July 2002

Genetic defects in a number of components of the dystrophin–glycoprotein complex (DGC) lead to distinct forms of muscular dystrophy. However, little is known about how alterations in the DGC are manifested in the pathophysiology present in dystrophic muscle tissue. One hypothesis is that the DGC protects the sarcolemma from contraction-induced damage. Using tracer molecules, we compared sarcolemmal integrity in animal models for muscular dystrophy and in muscular dystrophy patient samples. Evans blue, a low molecular weight diazo dye, does not cross into skeletal muscle fibers in normal mice. In contrast, mdx mice, a dystrophin-deficient animal model for Duchenne muscular dystrophy, showed significant Evans blue accumulation in skeletal muscle fibers. We also studied Evans blue dispersion in transgenic mice bearing different dystrophin mutations, and we demonstrated that cytoskeletal and sarcolemmal attachment of dystrophin might be a necessary requirement to prevent serious fiber damage. The extent of dye incorporation in transgenic mice correlated with the phenotypic severity of similar dystrophin mutations in humans. We furthermore assessed Evans blue incorporation in skeletal muscle of the dystrophia muscularis (dy/dy) mouse and its milder allelic variant, the dy2J/dy2J mouse, animal models for congenital muscular dystrophy. Surprisingly, these mice, which have defects in the laminin α2-chain, an extracellular ligand of the DGC, showed little Evans blue accumulation in their skeletal muscles. Taken together, these results suggest that the pathogenic mechanisms in congenital muscular dystrophy are different from those in Duchenne muscular dystrophy, although the primary defects originate in two components associated with the same protein complex.
Dismantling Cell–Cell Contacts during Apoptosis Is Coupled to a Caspase-dependent Proteolytic Cleavage of β-Catenin
Authors: Brancolini Claudio , Lazarevic Dean , Rodriguez Joe , Schneider Claudio
Published: July 2002

Cell death by apoptosis is a tightly regulated process that requires coordinated modification in cellular architecture. The caspase protease family has been shown to play a key role in apoptosis. Here we report that specific and ordered changes in the actin cytoskeleton take place during apoptosis.

In this context, we have dissected one of the first hallmarks in cell death, represented by the severing of contacts among neighboring cells. More specifically, we provide demonstration for the mechanism that could contribute to the disassembly of cytoskeletal organization at cell–cell adhesion. In fact, β-catenin, a known regulator of cell–cell adhesion, is proteolytically processed in different cell types after induction of apoptosis. Caspase-3 (cpp32/apopain/yama) cleaves in vitro translated β-catenin into a form which is similar in size to that observed in cells undergoing apoptosis. β-Catenin cleavage, during apoptosis in vivo and after caspase-3 treatment in vitro, removes the amino- and carboxy-terminal regions of the protein. The resulting β-catenin product is unable to bind α-catenin that is responsible for actin filament binding and organization. This evidence indicates that connection with actin filaments organized at cell–cell contacts could be dismantled during apoptosis. Our observations suggest that caspases orchestrate the specific and sequential changes in the actin cytoskeleton occurring during cell death via cleavage of different regulators of the microfilament system.
The Human DnaJ Homologue dj2 Facilitates Mitochondrial Protein Import and Luciferase Refolding
Authors: Terada Kazutoyo , Kanazawa Masaki , Bukau Bernd , Mori Masataka
Published: July 2002

DnaJ homologues function in cooperation with hsp70 family members in various cellular processes including intracellular protein trafficking and folding. Three human DnaJ homologues present in the cytosol have been identified: dj1 (hsp40/hdj-1), dj2 (HSDJ/hdj-2), and neuronal tissue-specific hsj1. dj1 is thought to be engaged in folding of nascent polypeptides, whereas functions of the other DnaJ homologues remain to be elucidated. To investigate roles of dj2 and dj1, we developed a system of chaperone depletion from and readdition to rabbit reticulocyte lysates. Using this system, we found that heat shock cognate 70 protein (hsc70) and dj2, but not dj1, are involved in mitochondrial import of preornithine transcarbamylase. Bacterial DnaJ could replace mammalian dj2 in mitochondrial protein import. We also tested the effects of these DnaJ homologues on folding of guanidine-denatured firefly luciferase. Unexpectedly, dj2, but not dj1, together with hsc70 refolded the protein efficiently. We propose that dj2 is the functional partner DnaJ homologue of hsc70 in the mammalian cytosol. Bacterial DnaJ protein could replace mammalian dj2 in the refolding of luciferase. Thus, the cytosolic chaperone system for mitochondrial protein import and for protein folding is highly conserved, involving DnaK and DnaJ in bacteria, Ssa1–4p and Ydj1p in yeast, and hsc70 and dj2 in mammals.
Ceramide Accumulation Uncovers a Cycling Pathway for the cis-Golgi Network Marker, Infectious Bronchitis Virus M Protein
Authors: Maceyka Michael , Machamer Carolyn E.
Published: July 2002

The M glycoprotein from the avian coronavirus, infectious bronchitis virus (IBV), contains information for localization to the cis-Golgi network in its first transmembrane domain. We hypothesize that localization to the Golgi complex may depend in part on specific interactions between protein transmembrane domains and membrane lipids. Because the site of sphingolipid synthesis overlaps the localization of IBV M, we asked whether perturbation of sphingolipids affected localization of IBV M. Short-term treatment with two inhibitors of sphingolipid synthesis had no effect on localization of IBV M or other Golgi markers. Thus, ongoing synthesis of these lipids was not required for proper localization. Surprisingly, a third inhibitor, d,l-threo-1-phenyl-2-decanoylamino-3-morpholino- 1-propanol (PDMP), shifted the steady-state distribution of IBV M from the Golgi complex to the ER. This effect was rapid and reversible and was also observed for ERGIC-53 but not for Golgi stack proteins. At the concentration of PDMP used, conversion of ceramide into both glucosylceramide and sphingomyelin was inhibited. Pretreatment with upstream inhibitors partially reversed the effects of PDMP, suggesting that ceramide accumulation mediates the PDMP-induced alterations. Indeed, an increase in cellular ceramide was measured in PDMP-treated cells. We propose that IBV M is at least in part localized by retrieval mechanisms. Further, ceramide accumulation reveals this cycle by upsetting the balance of anterograde and retrograde traffic and/ or disrupting retention by altering bilayer dynamics.
Expression of Rab3D N135I Inhibits Regulated Secretion of ACTH in AtT-20 Cells
Authors: Baldini Giulia , Baldini Giovanna , Wang Guangyi , Weber Mattew , Zweyer Marina , Bareggi Renato , Witkin Joan W. , Martelli Alberto M.
Published: July 2002

Rab proteins are small molecular weight GTPases that control vesicular traffic in eucaryotic cells. A subset of Rab proteins, the Rab3 proteins are thought to play an important role in regulated exocytosis of vesicles. In transfected AtT-20 cells expressing wild-type Rab3D, we find that a fraction of the protein is associated with dense core granules. In the same cells, expression of a mutated isoform of Rab3D, Rab3D N135I, inhibits positioning of dense core granules near the plasma membrane, blocks regulated secretion of mature ACTH, and impairs association of Rab3A to membranes. Expression of Rab3D N135I does not change the levels of ACTH precursor or the efficiency with which the precursor is processed into ACTH hormone and packaged into dense core granules. We also find that cells expressing mutated Rab3D differentiate to the same extent as untransfected AtT-20 cells. We conclude that expression of Rab3D N135I specifically impairs late membrane trafficking events necessary for ACTH hormone secretion.
Apoptotic Membrane Blebbing Is Regulated by Myosin Light Chain Phosphorylation
Authors: Mills Jason C. , Stone Nicole L. , Erhardt Joseph , Pittman Randall N.
Published: July 2002

The evolutionarily conserved execution phase of apoptosis is defined by characteristic changes occurring during the final stages of death; specifically cell shrinkage, dynamic membrane blebbing, condensation of chromatin, and DNA fragmentation. Mechanisms underlying these hallmark features of apoptosis have previously been elusive, largely because the execution phase is a rapid event whose onset is asynchronous across a population of cells. In the present study, a model system is described for using the caspase inhibitor, z-VAD-FMK, to block apoptosis and generate a synchronous population of cells actively extruding and retracting membrane blebs. This model system allowed us to determine signaling mechanisms underlying this characteristic feature of apoptosis. A screen of kinase inhibitors performed on synchronized blebbing cells indicated that only myosin light chain kinase (MLCK) inhibitors decreased blebbing. Immunoprecipitation of myosin II demonstrated that myosin regulatory light chain (MLC) phosphorylation was increased in blebbing cells and that MLC phosphorylation was prevented by inhibitors of MLCK. MLC phosphorylation is also mediated by the small G protein, Rho. C3 transferase inhibited apoptotic membrane blebbing, supporting a role for a Rho family member in this process. Finally, blebbing was also inhibited by disruption of the actin cytoskeleton. Based on these results, a working model is proposed for how actin/myosin II interactions cause cell contraction and membrane blebbing. Our results provide the first evidence that MLC phosphorylation is critical for apoptotic membrane blebbing and also implicate Rho signaling in these active morphological changes. The model system described here should facilitate future studies of MLCK, Rho, and other signal transduction pathways activated during the execution phase of apoptosis.