bluuebunny/crossref_metadata_2025 huggingface dataset:
Responsible Research: Ways of being a researcher
Authors: Edwards Anne
Published: July 2002

Abstract Being an educational researcher is not an easy option. Educational researchers are practitioners in an engaged social science which makes particular demands on them. These include responsibility to the field of study. In this presidential address it is argued that close‐to‐the‐field research, that can do justice to the meaning making that occurs there, is an important part of the responsibility of the educational research community. Research of this kind, sometimes called interpretative or transformative research, calls for a form of engagement with the field which could be termed responsible agency. Because examinations of individual agency and responsibility must take into account contexts, their values and opportunities, the address also examines how the British Educational Research Association, as a learned society, can sustain the identities of engaged researchers and how their revelations from the field can inform educational policy and the methodologies which shape educational enquiry.
Emotional and Adrenocortical Responses of Infants to the Strange Situation: The Differential Function of Emotional Expression
Authors: Spangler Gottfried , Schieche Michael
Published: July 2002

The aim of the study was to investigate biobehavioural organisation in infants with different qualities of attachment. Quality of attachment (security and disorganisation), emotional expression, and adrenocortical stress reactivity were investigated in a sample of 106 infants observed during Ainsworth’s Strange Situation at the age of 12 months. In addition, behavioural inhibition was assessed from maternal reports. As expected, securely attached infants did not show an adrenocortical response. Regarding the traditionally defined insecurely attached groups, adrenocortical activation during the strange situation was found for the ambivalent group, but not for the avoidant one. Previous ndings of increased adrenocortical activity in disorganised infants could not be replicated. In line with previous ndings, adrenocortical activation was most prominent in insecure infants with high behavioural inhibition indicating the function of a secure attachment relationship as a social buffer against less adaptive temperamental dispositions. Additional analyses indicated that adrenocortical reactivity and behavioural distress were not based on common activation processes. Biobehavioural associations within the different attachment groups suggest that biobehavioural processes in securely attached infants may be different from those in insecurely attached and disorganised groups. Whereas a coping model may be applied to describe the biobehavioural organisation of secure infants, an arousal model explanation may be more appropriate for the other groups.
A Smithean Perspective on Increasing Returns
Authors: Buchanan James M. , Yoon Yong J.
Published: July 2002

Despite its recent re-emergence to analytical importance, the phenomenon of increasing returns remains outside the central core of neoclassical economics. The history of this idea (or set of ideas) might have been quite different if Adam Smith's explanation of the origins of trade had not been replaced by that of David Ricardo. To Adam Smith, mutually beneficial exchange emerges because of specialization, which, in its turn, implies the presence of increasing returns to the size of the exchange nexus. Even in a world of equals, trade offers mutuality of gain. There is no need for participants in the economic nexus to differ one from another. In the Ricardian logic, by contrast, trade presumably emerges because productive resources differ in their capacities to create economic value, at least among separate “goods.” Specialization is a “natural” feature of resource endowments—a feature that is exploited by trade. Comparative advantage ensures the mutuality of gain. But, in this explanation, there is no direct linkage between the size of the exchange network and the degree of specialization that is viable. There is no need to introduce increasing returns. Comparative advantage may be present even if there are constant returns to scale, both for the economy and for its separate productive sectors.
The Labor “Embodied” In Smith's Labor-Commanded Measure: A “Rationally Reconstructed” Legend
Authors: Hueckel Glenn
Published: July 2002

Nearly a half-century has passed since Ronald Meek (1956, p. 63) warned us that Adam Smith's notion of the labor commanded by a commodity in the marketplace is to be understood not as an expression of the “substance” of value, varying “directly with the quantity of social labor used to produce” the object, but rather as nothing more than a unit of value measure with no fixed relationship to the labor “embodied” in production. It was this distinction that he sought to fix in our minds with his memorable image of the magnet. Indeed, it is more than three times as long since John Stuart Mill (1848, p. 568) conveyed the same distinction with his particularly apt metaphor of “the thermometer and the fire.” Further, it is now forty years since Mark Blaug (1959), reminding us of that distinction, returned our attention to Smith's use of his measure as an expression of potential productive capacity (a view advanced earlier yet by Hla Myint 1948, pp. 20–21 and by Meek 1956, p. 65), but one that conveys a subjective dimension as well. Yet in spite of a now widespread concurrence in this reading, the “legend” that Smith's concept of labor commanded is to be understood as expressing, in some way, price ratios proportional to ratios of labor embodied in production remains remarkably resilient.
Substance Dependence and Other Psychiatric Disorders Among Drug Dependent Subjects: Race and Gender Correlates
Authors: Compton Wilson M. , Cottler Linda B. , Ben Abdallah Arbi , Phelps Deborah L. , Spitznagel Edward L. , Horton Joseph C.
Published: July 2002

Persons in drug treatment with drug dependence were interviewed with the NIMH Diagnostic Interview Schedule to ascertain DSM‐III‐R disorders. Lifetime prevalence rates were 64% for alcohol dependence, 44% for antisocial personality disorder (ASPD), 39% for phobic disorders, 24% for major depression, 12% for dysthymia, 10% for generalized anxiety disorder, 3% for panic disorder, 3% for mania, 3% for obsessive compulsive disorder, 2% for bulimia, 1% for schizophrenia, and 1% for anorexia. When stratified by race and age, significant main effects were seen, but there were no significant interactions except in “any non‐substance disorder” and in the mean number of non‐substance use disorders. Caucasians had a higher mean number of drug dependence disorders and higher overall rates of “any other” disorder than African‐Americans, and Caucasians and males had higher mean numbers of non‐substance use disorders than African‐Americans and females, respectively. This was related to rates of alcohol, cannabis, and hallucinogen dependence, and ASPD rates that were higher among men than women and higher among Caucasian respondents than African‐American for alcohol, cannabis, hallucinogen, opiate and sedative dependence, major depression, dysthymia, and generalized anxiety disorder. In contrast, women had higher rates than men of amphetamine dependence, phobic disorder, major depression, dysthymia, panic disorder, obsessive compulsive disorder, and mania. African‐Americans had higher rates than Caucasians of amphetamine, cocaine, and phencyclidine dependence, but for no comorbid disorders were the rates higher among African‐Americans than Caucasians. The differences according to gender in rates of disorders among substance dependent persons are consistent with the results of general population surveys, but the differences in rates according to race are in contrast to these same community surveys. Limitations in the utility of the concept of race as a valid category diminish the generalizability of the findings; however, one possible explanation is differential treatment seeking in African‐American and Caucasian populations that would result in the differences seen.
The Effect of Stimulant and Sedative Use on Treatment Outcome of Patients Admitted to Methadone Maintenance Treatment
Authors: DeMaria Peter A. , Sterling Robert , Weinstein Stephen P.
Published: July 2002

While methadone maintenance treatment (MMT) has been demonstrated to be an effective treatment for opiate dependence, its impact on the treatment outcome of other illicit drug abuse is not as clear. Using the initial urine drug screen (UDS) and follow‐up UDS at 1, 6, 12, and 24 months, 167 patients consecutively admitted to MMT were evaluated for opiate, sedative (predominantly benzodiazepine), and stimulant (predominantly cocaine) use. Retention for the opiate only group was 97.32 days longer on average than for patients using opiates along with stimulants, sedatives, or both stimulants and sedatives. Patients abusing opiates only had the greatest decrease in drug use; however, MMT was also associated with decreases in cocaine and sedative use over the 24 month follow‐up period. There was no evidence that patients “switched” their drugs of abuse with time in treatment. The negative impact of non‐opiate drug use on outcome in MMT and its implications for treatment planning are discussed.
New Developments in the Pharmacotherapy of Alcohol Dependence
Authors: Myrick Hugh , Brady Kathleen T. , Malcolm Robert
Published: July 2002

Neuroscientific underpinnings and pharmacotherapeutic treatments of sub‐stance use disorders are rapidly developing areas of study. In particular, there have been exciting new developments in our understanding of the involvement of excitatory amino acid neurotransmitter systems and the opiate and serotonin systems in the pathophysiology of alcohol withdrawal, alcohol dependence, and in subtypes of individuals with alcoholism. In this article, new developments in the pharmacotherapy of alcohol dependence will be reviewed. In particular, the use of anticonvulsants in alcohol withdrawal and protracted abstinence syndromes will be discussed. New data on opiate antagonists and acamprosate, an agent that exerts actions through excitatory amino acid systems in relapse prevention, will be reviewed. Finally, there will be a review of new data concerning the use of serotonin reuptake inhibitors in subtypes of alcoholism and the use of combination pharmacotherapy.
Histochemical Characterization of Renal Phosphatases in Rats Treated With Lead
Authors: Jarrar Bashir M. , Mahmoud Zuheir N.
Published: July 2002

Seven groups of 84 male white rats ( Rattus norvegicus) were exposed to lead acetate (0, 0.125, 0.25, 0.5, 1.0, 2.0, and 4% for 1 to 5 months) in drinking water to investigate possible histochemical changes in the activity of renal phosphatases due to lead intoxication. In comparison with respective control rats, chronic exposure to subtoxic doses of lead increased the activities of acid-, neutral-, alkaline-, and membrane-bound Na+-K+ adenosine triphosphatase and glucose-6-phosphatase. In contrast, significant reduction in the activities of 5-nucleotidase and mitochondrial adenosine triphosphatase was observed. Changes in the activity of renal phosphatases were seen mainly in the epithelium of the proximal tubules and, to a lesser extent, in the medullary interstitial cells and the glomeruli as well. These histochemical findings led us to conclude that such changes in the activity of phosphatases were due to long-term exposure to subtoxic doses and could be an adaptation to the metabolic, structural, and functional alterations in the organelles of the renal cells due to lead intoxication.
Ultra‐rapid, antagonist‐precipitated opiate detoxification under general anaesthesia or sedation
Authors: BREWER COLIN
Published: July 2002

Ultra‐rapid opiate withdrawal precipitated by naloxone or naltrexone during general anaesthesia or varying degrees of sedation, usually with the addition of α‐2 adrenergic agonists, is an increasingly popular technique. As with anaesthesia or effective analgesia for dentistry or obstetrics, it can be justified on general humane principles. It also simplifies and speeds the transition to treatment programmes involving naltrexone. This paper reviews its development and the current range of techniques, including the use of octreotide to prevent diarrhoea. The prospect of avoiding or shortening the worst discomforts of withdrawal may encourage more opiate addicts to attempt detoxification and opiate‐free management, including patients who have been successfully rehabilitated on long‐term methadone maintenance treatment (MMT). Despite good motivation and preparation, a significant minority of MMT patients consistently fail to complete conventional withdrawal programmes because of increasing physical discomfort. It is emphasized that there is usually more to the treatment of opiate dependence than withdrawal.
Alcohol, smoking and lung disease
Authors: STRÖM KERSTIN
Published: July 2002

Alcohol and smoking are two well‐known health hazards. Alcohol and tobacco consumption are strongly correlated and heavy drinkers have more trouble quitting smoking than do light drinkers. Death from tobaccorelated causes was more common than alcohol‐related death in a follow‐up study on patients admitted to an addiction programme for treatment of alcoholism and non‐nicotine drug dependence. In British male doctors in the middle and elderly age group, a protective effect of light and moderate alcohol consumption (1–3 British units of alcohol per day) compared with abstinence has been shown in one large survey. This protective effect was shown in overall mortality as well as in mortality from respiratory disease. Higher alcohol intakes were associated with an increase in mortality. This characteristic U‐formed, or J‐formed, dose‐response curve has been found in most studies with an apparent beneficial effect of modest alcohol intake and a harmful effect of high intakes. The anti‐inflammatory effect of alcohol has been considered to be responsible for its limited protective effect on smoking‐related lung function decline. Recently, a hitherto unconfirmed report suggests that the beneficial effect of alcohol on lung function in men is restricted to subjects with Lewis‐negative blood group (10% of the Caucasian population). On the other hand, the protective effect in those individuals is large enough to be clinically relevant. Prospective investigations including both men and women are needed to elucidate which individuals have a protective effect of light and moderate alcohol intake. The major deleterious effect of smoking, including passive smoking, must be kept in mind‐drinking alcohol surrounded by cigarette smoke might not be beneficial for respiratory health.
Mother‐child interactions in Canada and Italy: Linguistic responsiveness to late‐talking toddlers
Authors: Girolametto Luigi , Bonifacio Serena , Visini Cristiana , Weitzman Elaine , Zocconi Elisabetta , Pearce Patsy Steig
Published: July 2002

The aim was to examine cross‐cultural variation in linguistic responsiveness to young children in 10 English‐speaking mother‐child dyads and 10 Italian‐speaking mother‐child dyads. All 20 children were late talkers who possessed delays in expressive vocabulary development but age‐appropriate cognitive and receptive language skills. Dyads were filmed in 15‐minute free play contexts, which were transcribed and coded for measures of maternal linguistic input (e.g. rate, MLU, labels, expansions) and child language productivity (e.g. utterances, different words used). The results revealed that the Italian mothers used more utterances, spoke more quickly and used a more diverse vocabulary than the Canadian mothers. The Italian children mirrored their mothers and also used more utterances and a more diverse vocabulary than the Canadian children. Mothers in both groups used similar percentages of responsive labels and expansions. However, Italian mothers responded to fewer of their children's vocalizations, using a smaller percentage of imitations and interpretations than the Canadian mothers. Correlations between maternal input and children's language productivity revealed that contingent language measures (e.g. imitations, interpretations, expansions) were related to high levels of productivity in children in both cultural groups. The results support the use of language interventions based on increasing maternal responsiveness for these children at the one‐word stage of language development. They also point to differences that may be culturally based. For example, Italian mothers use faster rates of interaction and appear to have higher expectations for their children's verbal participation in interaction. This is reflected in higher rates of language production from their children, even though children in both cultural groups have similar vocabulary sizes.
Telethonin and Other New Proteins of the Z‐Disc of Skeletal Muscle
Authors: Faulkner Georgine , Lanfranchi Gerolamo , Valle Giorgio
Published: July 2002

This brief review outlines some of the most relevant proteins of the Z‐disc and the complex network of interactions that link them together in a stable structure. Apart from the well‐known Z‐disc proteins such as actin, cap‐Z, titin, nebulin, and α‐actinin 2, several other Z‐disc proteins have been recently discovered, including telethonin and myotilin that have been linked to limb girdle muscular dystrophies. Some proteins including ALP and ZASP have known interaction domains (PDZ and LIM motifs), whereas others like FATZ have no canonical interaction domains, although they are known to bind several proteins. Another new Z‐disc protein is γ‐filamin that could provide a link between the plasma membrane and myofibrils because it binds directly to γ‐ and δ‐sarcoglycans and indirectly to α‐actinin 2 via FATZ and myotilin. A greater knowledge of Z‐disc proteins and their interactions is essential for understanding their role in the structure and function of muscle.
Interactions between Newly Synthesized Glycoproteins, Calnexin and a Network of Resident Chaperones in the Endoplasmic Reticulum
Authors: Tatu Utpal , Helenius Ari
Published: July 2002

Calnexin is a membrane-bound lectin and a molecular chaperone that binds newly synthesized glycoproteins in the endoplasmic reticulum (ER). To analyze the oligomeric properties of calnexin and calnexin-substrate complexes, sucrose velocity gradient centrifugation and chemical cross-linking were used. After CHAPS solubilization of Chinese Hamster Ovary cells, the unoccupied calnexin behaved as a monomer sedimenting at 3.5 S20,W. For calnexin-substrate complexes the S-values ranged between 3.5–8 S20,W, the size increasing with the molecular weight of the substrate. Influenza hemagglutinin, a well-characterized substrate associated with calnexin in complexes that sedimented at 5–5.5 S20,W. The majority of stable complexes extracted from cells, appeared to contain a single calnexin and a single substrate molecule, with about one third of the calnexin in the cell being unoccupied or present in weak associations. However, when chemical cross-linking was performed in intact cells, the calnexin-substrate complexes and calnexin itself was found to be part of a much larger heterogeneous protein network that included other ER proteins. Pulse-chase analysis of influenza-infected cells combined with chemical cross-linking showed that HA was part of large, heterogeneous, cross-linked entities during the early phases of folding, but no longer after homotrimer assembly. The network of weakly associated resident ER chaperones which included BiP, GRP94, calreticulin, calnexin, and other proteins, may serve as a matrix that binds early folding and assembly intermediates and restricts their exit from the ER.
Distribution and Function of Laminins in the Neuromuscular System of Developing, Adult, and Mutant Mice
Authors: Patton Bruce L. , Miner Jeffrey H. , Chiu Arlene Y. , Sanes Joshua R.
Published: July 2002

Laminins, heterotrimers of α, β, and γ chains, are prominent constituents of basal laminae (BLs) throughout the body. Previous studies have shown that laminins affect both myogenesis and synaptogenesis in skeletal muscle. Here we have studied the distribution of the 10 known laminin chains in muscle and peripheral nerve, and assayed the ability of several heterotrimers to affect the outgrowth of motor axons. We show that cultured muscle cells express four different α chains (α1, α2, α4, and α5), and that developing muscles incorporate all four into BLs. The portion of the muscle's BL that occupies the synaptic cleft contains at least three α chains and two β chains, but each is regulated differently. Initially, the α2, α4, α5, and β1 chains are present both extrasynaptically and synaptically, whereas β2 is restricted to synaptic BL from its first appearance. As development proceeds, α2 remains broadly distributed, whereas α4 and α5 are lost from extrasynaptic BL and β1 from synaptic BL. In adults, α4 is restricted to primary synaptic clefts whereas α5 is present in both primary and secondary clefts. Thus, adult extrasynaptic BL is rich in laminin 2 (α2β1γ1), and synaptic BL contains laminins 4 (α2β2γ1), 9 (α4β2γ1), and 11 (α5β2γ1). Likewise, in cultured muscle cells, α2 and β1 are broadly distributed but α5 and β2 are concentrated at acetylcholine receptor–rich “hot spots,” even in the absence of nerves. The endoneurial and perineurial BLs of peripheral nerve also contain distinct laminin chains: α2, β1, γ1, and α4, α5, β2, γ1, respectively. Mutation of the laminin α2 or β2 genes in mice not only leads to loss of the respective chains in both nerve and muscle, but also to coordinate loss and compensatory upregulation of other chains. Notably, loss of β2 from synaptic BL in β2−/− “knockout” mice is accompanied by loss of α5, and decreased levels of α2 in dystrophic α2dy/dy mice are accompanied by compensatory retention of α4. Finally, we show that motor axons respond in distinct ways to different laminin heterotrimers: they grow freely between laminin 1 (α1β1γ1) and laminin 2, fail to cross from laminin 4 to laminin 1, and stop upon contacting laminin 11. The ability of laminin 11 to serve as a stop signal for growing axons explains, in part, axonal behaviors observed at developing and regenerating synapses in vivo.
Fusion of Lysosomes with Late Endosomes Produces a Hybrid Organelle of Intermediate Density and Is NSF Dependent
Authors: Mullock Barbara M. , Bright Nicholas A. , Fearon Clare W. , Gray Sally R. , Luzio J.
Published: July 2002

Using a cell-free content mixing assay containing rat liver endosomes and lysosomes in the presence of pig brain cytosol, we demonstrated that after incubation at 37°C, late endosome–lysosome hybrid organelles were formed, which could be isolated by density gradient centrifugation. ImmunoEM showed that the hybrids contained both an endocytosed marker and a lysosomal enzyme. Formation of the hybrid organelles appeared not to require vesicular transport between late endosomes and lysosomes but occurred as a result of direct fusion. Hybrid organelles with similar properties were isolated directly from rat liver homogenates and thus were not an artifact of cell-free incubations. Direct fusion between late endosomes and lysosomes was an N-ethylmaleimide–sensitive factor– dependent event and was inhibited by GDP-dissociation inhibitor, indicating a requirement for a rab protein. We suggest that in cells, delivery of endocytosed ligands to an organelle where proteolytic digestion occurs is mediated by direct fusion of late endosomes with lysosomes. The consequences of this fusion to the maintenance and function of lysosomes are discussed.
KSA Antigen Ep-CAM Mediates Cell–Cell Adhesion of Pancreatic Epithelial Cells: Morphoregulatory Roles in Pancreatic Islet Development
Authors: Cirulli V. , Crisa L. , Beattie G.M. , Mally M.I. , Lopez A.D. , Fannon A. , Ptasznik A. , Inverardi L. , Ricordi C. , Deerinck T. , Ellisman M. , Reisfeld R.A. , Hayek A.
Published: July 2002

Cell adhesion molecules (CAMs) are important mediators of cell–cell interactions and regulate cell fate determination by influencing growth, differentiation, and organization within tissues. The human pancarcinoma antigen KSA is a glycoprotein of 40 kD originally identified as a marker of rapidly proliferating tumors of epithelial origin. Interestingly, most normal epithelia also express this antigen, although at lower levels, suggesting that a dynamic regulation of KSA may occur during cell growth and differentiation. Recently, evidence has been provided that this glycoprotein may function as an epithelial cell adhesion molecule (Ep-CAM). Here, we report that Ep-CAM exhibits the features of a morphoregulatory molecule involved in the development of human pancreatic islets. We demonstrate that Ep-CAM expression is targeted to the lateral domain of epithelial cells of the human fetal pancreas, and that it mediates calcium-independent cell–cell adhesion. Quantitative confocal immunofluorescence in fetal pancreata identified the highest levels of Ep-CAM expression in developing islet-like cell clusters budding from the ductal epithelium, a cell compartment thought to comprise endocrine progenitors. A surprisingly reversed pattern was observed in the human adult pancreas, displaying low levels of Ep-CAM in islet cells and high levels in ducts. We further demonstrate that culture conditions promoting epithelial cell growth induce upregulation of Ep-CAM, whereas endocrine differentiation of fetal pancreatic epithelial cells, transplanted in nude mice, is associated with a downregulation of Ep-CAM expression. In addition, a blockade of Ep-CAM function by KS1/4 mAb induced insulin and glucagon gene transcription and translation in fetal pancreatic cell clusters. These results indicate that developmentally regulated expression and function of Ep-CAM play a morphoregulatory role in pancreatic islet ontogeny.
Apical Vesicles Bearing Inositol 1,4,5-trisphosphate Receptors in the Ca2+Initiation Site of Ductal Epithelium of Submandibular Gland
Authors: Yamamoto-Hino Miki , Miyawaki Atsushi , Segawa Akihisa , Adachi Eijiro , Yamashina Shohei , Fujimoto Toyoshi , Sugiyama Tomoyasu , Furuichi Teiichi , Hasegawa Mamoru , Mikoshiba Katsuhiko
Published: July 2002

In polarized epithelial cells, agonists trigger Ca2+ waves and oscillations. These patterns may be caused by the compartmentalization of inositol 1,4,5-trisphosphate (IP3)-sensitive Ca2+ pools into specific regions. We have investigated the relationship between the distribution of IP3 receptors (IP3Rs) and the spatiotemporal pattern of Ca2+ signaling in the duct cells of the rat submandibular gland (SMG). Using immunofluorescence, although labeling was somewhat heterogeneous, the IP3Rs were colocalized to the apical pole of the duct cells. Immunoelectron microscopy identified small apical vesicles bearing IP3R2 in some types of duct cells. Real-time confocal imaging of intact ducts demonstrated that, after carbachol stimulation, an initial Ca2+ spike occurred in the apical region. Subsequently, repetitive Ca2+ spikes spread from the apical to the middle cytoplasm. These apical Ca2+ initiation sites were found only in some “pioneer cells,” rather than in all duct cells. We performed both Ca2+ imaging and immunofluorescence on the same ducts and detected the strongest immunosignals of IP3R2 in the Ca2+ initiation sites of the pioneer cells. The subcellular localization and expression level of IP3Rs correlated strongly with the spatiotemporal nature of the intracellular Ca2+ signal and distinct Ca2+ responses among the rat SMG duct cells.
Directed Expression of Keratin 16 to the Progenitor Basal Cells of Transgenic Mouse Skin Delays Skin Maturation
Authors: Paladini Rudolph D. , Coulombe Pierre A.
Published: July 2002

We previously hypothesized that the type I keratin 16 (K16) plays a role in the process of keratinocyte activation that occurs in response to skin injury (Paladini, R.D., K. Takahashi, N.S. Bravo, and P.A. Coulombe. 1996. J. Cell Biol. 132:381–397). To further examine its properties in vivo, the human K16 cDNA was constitutively expressed in the progenitor basal layer of transgenic mouse skin using the K14 gene promoter. Mice that express approximately as much K16 protein as endogenous K14 display a dramatic postnatal phenotype that consists of skin that is hyperkeratotic, scaly, and essentially devoid of fur. Histologically, the epidermis is thickened because of hyperproliferation of transgenic basal cells, whereas the hair follicles are decreased in number, poorly developed, and hypoproliferative. Microscopically, the transgenic keratinocytes are hypertrophic and feature an altered keratin filament network and decreased cell–cell adhesion. The phenotype normalizes at ∼5 wk after birth. In contrast, control mice expressing a K16-K14 chimeric protein to comparable levels are normal. The character and temporal evolution of the phenotype in the K16 transgenic mice are reminiscent of the activated EGF receptor– mediated signaling pathway in skin. In fact, tyrosine phosphorylation of the EGF receptor is increased in the newborn skin of K16 transgenic mice. We conclude that expression of K16 can significantly alter the response of skin keratinocytes to signaling cues, a distinctive property likely resulting from its unique COOH-terminal tail domain.
The RET–Glial Cell-derived Neurotrophic Factor (GDNF) Pathway Stimulates Migration and Chemoattraction of Epithelial Cells
Authors: Tang Ming-Jer , Worley Dane , Sanicola Michele , Dressler Gregory R.
Published: July 2002

Embryonic development requires cell migration in response to positional cues. Yet, how groups of cells recognize and translate positional information into morphogenetic movement remains poorly understood. In the developing kidney, the ureteric bud epithelium grows from the nephric duct towards a group of posterior intermediate mesodermal cells, the metanephric mesenchyme, and induces the formation of the adult kidney. The secreted protein GDNF and its receptor RET are required for ureteric bud outgrowth and subsequent branching. However, it is unclear whether the GDNF–RET pathway regulates cell migration, proliferation, survival, or chemotaxis. In this report, we have used the MDCK renal epithelial cell line to show that activation of the RET pathway results in increased cell motility, dissociation of cell adhesion, and the migration towards a localized source of GDNF. Cellular responses to RET activation include the formation of lamellipodia, filopodia, and reorganization of the actin cytoskeleton. These data demonstrate that GDNF is a chemoattractant for RET-expressing epithelial cells and thus account for the developmental defects observed in RET and GDNF mutant mice. Furthermore, the RET-transfected MDCK cells described in this report are a promising model for delineating RET signaling pathways in the renal epithelial cell lineage.
Chromosomal Proteins HMG-14 and HMG-17 Are Released from Mitotic Chromosomes and Imported into the Nucleus by Active Transport
Authors: Hock Robert , Scheer Ulrich , Bustin Michael
Published: July 2002

The high mobility group 14/17 (HMG-14/ -17) proteins form specific complexes with nucleosome core particles and produce distinct footprints on nucleosomal DNA. Therefore, they could be an integral part of the chromatin fiber. Here we show that during the cell cycle these proteins are transiently dissociated from chromatin. They colocalize with the nuclear DNA in interphase and prophase but not in metaphase and anaphase. They relocate into the nucleus and colocalize again with the DNA in late telophase, concomitantly with the appearance of the nuclear envelope. Thus, these nucleosomal binding proteins are not always associated with chromatin. Using reconstituted nuclei and permeabilized cells, we demonstrate that these two small proteins, with a molecular mass <10 kD, are actively imported into the nucleus. We identify the major elements involved in the nuclear import of these chromosomal proteins: HMG-14/-17 proteins contain an intrinsic bipartite nuclear localization signal, and their entry into the nucleus through nuclear pores requires energy and the participation of importin α. These findings suggest that the cell cycle–related association of HMG-14/-17 with chromatin is dependent on, and perhaps regulated by, nuclear import processes.
Regulation of Bone Morphogenetic Protein Activity by Pro Domains and Proprotein Convertases
Authors: Constam Daniel B. , Robertson Elizabeth J.
Published: July 2002

Bone morphogenetic proteins (BMPs) are derived from inactive precursor proteins by endoproteolytic cleavage. Here we show that processing of Nodal and Myc-tagged BMP4 is significantly enhanced by SPC1/Furin or SPC4/PACE4, providing direct evidence that regulation of BMP signaling is likely to be controlled by subtilisin-like proprotein convertase (SPC) activities. Nodal processing is dramatically enhanced if two residues adjacent to the precursor cleavage site are substituted with amino acids found at the equivalent positions of Activin, demonstrating that structural constraints at the precursor cleavage site limit the processing efficiency. However, in transfection assays, mature Nodal is undetectable either in culture supernatants or in cell lysates, despite efficient cleavage of the precursor protein, suggesting that mature Nodal is highly unstable. Domain swap experiments support this conclusion since mature BMP4 or Dorsalin are also destabilized when expressed in conjunction with the Nodal pro domain. By contrast, mature Nodal is stabilized by the Dorsalin pro domain, which mediates the formation of stable complexes. Collectively, these data show that the half-life of mature BMPs is greatly influenced by the identity of their pro regions.
The Spinal Muscular Atrophy Disease Gene Product, Smn
Authors: Carvalho Teresa , Almeida Fátima , Calapez Alexandre , Lafarga Miguel , Berciano Maria T. , Carmo-Fonseca Maria
Published: July 2002

The spliceosomal snRNAs U1, U2, U4, and U5 are synthesized in the nucleus, exported to the cytoplasm to assemble with Sm proteins, and reimported to the nucleus as ribonucleoprotein particles. Recently, two novel proteins involved in biogenesis of small nuclear ribonucleoproteins (snRNPs) were identified, the Spinal muscular atrophy disease gene product (SMN) and its associated protein SIP1. It was previously reported that in HeLa cells, SMN and SIP1 form discrete foci located next to Cajal (coiled) bodies, the so-called “gemini of coiled bodies” or “gems.” An intriguing feature of gems is that they do not appear to contain snRNPs. Here we show that gems are present in a variable but small proportion of rapidly proliferating cells in culture. In the vast majority of cultured cells and in all primary neurons analyzed, SMN and SIP1 colocalize precisely with snRNPs in the Cajal body. The presence of SMN and SIP1 in Cajal bodies is confirmed by immunoelectron microscopy and by microinjection of antibodies that interfere with the integrity of the structure. The association of SMN with snRNPs and coilin persists during cell division, but at the end of mitosis there is a lag period between assembly of new Cajal bodies in the nucleus and detection of SMN in these structures, suggesting that SMN is targeted to preformed Cajal bodies. Finally, treatment of cells with leptomycin B (a drug that blocks export of U snRNAs to the cytoplasm and consequently import of new snRNPs into the nucleus) is shown to deplete snRNPs (but not SMN or SIP1) from the Cajal body. This suggests that snRNPs flow through the Cajal body during their biogenesis pathway.
P0 Glycoprotein Overexpression Causes Congenital Hypomyelination of Peripheral Nerves
Authors: Wrabetz Lawrence , Feltri Maria Laura , Quattrini Angelo , Imperiale Daniele , Previtali Stefano , D'Antonio Maurizio , Martini Rudolf , Yin Xinghua , Trapp Bruce D. , Zhou Lei , Chiu Shing-Yan , Messing Albee
Published: July 2002

We show that normal peripheral nerve myelination depends on strict dosage of the most abundantly expressed myelin gene, myelin protein zero (Mpz). Transgenic mice containing extra copies of Mpz manifested a dose-dependent, dysmyelinating neuropathy, ranging from transient perinatal hypomyelination to arrested myelination and impaired sorting of axons by Schwann cells. Myelination was restored by breeding the transgene into the Mpz-null background, demonstrating that dysmyelination does not result from a structural alteration or Schwann cell-extrinsic effect of the transgenic P0 glycoprotein. Mpz mRNA overexpression ranged from 30–700%, whereas an increased level of P0 protein was detected only in nerves of low copy-number animals. Breeding experiments placed the threshold for dysmyelination between 30 and 80% Mpz overexpression. These data reveal new points in nerve development at which Schwann cells are susceptible to increased gene dosage, and suggest a novel basis for hereditary neuropathy.
Leydig Cell Loss and Spermatogenic Arrest in Platelet-Derived Growth Factor (Pdgf)-a–Deficient Mice
Authors: Gnessi Lucio , Basciani Sabrina , Mariani Stefania , Arizzi Mario , Spera Giovanni , Wang Chiayeng , Bondjers Cecilia , Karlsson Linda , Betsholtz Christer
Published: July 2002

Platelet-derived growth factor (PDGF)- A–deficient male mice were found to develop progressive reduction of testicular size, Leydig cells loss, and spermatogenic arrest. In normal mice, the PDGF-A and PDGF-Rα expression pattern showed positive cells in the seminiferous epithelium and in interstitial mesenchymal cells, respectively. The testicular defects seen in PDGF-A−/− mice, combined with the normal developmental expression of PDGF-A and PDGF-Rα, indicate that through an epithelial-mesenchymal signaling, the PDGF-A gene is essential for the development of the Leydig cell lineage. These findings suggest that PDGF-A may play a role in the cascade of genes involved in male gonad differentiation. The Leydig cell loss and the spermatogenic impairment in the mutant mice are reminiscent of cases of testicular failure in man.
Introducing a Null Mutation in the Mouse K6α and K6β Genes Reveals Their Essential Structural Role in the Oral Mucosa
Authors: Wong Pauline , Colucci-Guyon Emma , Takahashi Kenzo , Gu Changhong , Babinet Charles , Coulombe Pierre A.
Published: July 2002

Mammalian genomes feature multiple genes encoding highly related keratin 6 (K6) isoforms. These type II keratins show a complex regulation with constitutive and inducible components in several stratified epithelia, including the oral mucosa and skin. Two functional genes, K6α and K6β, exist in a head-to-tail tandem array in mouse genomes. We inactivated these two genes simultaneously via targeting and homologous recombination. K6 null mice are viable and initially indistinguishable from their littermates. Starting at two to three days after birth, they show a growth delay associated with reduced milk intake and the presence of white plaques in the posterior region of dorsal tongue and upper palate. These regions are subjected to greater mechanical stress during suckling. Morphological analyses implicate the filiform papillae as being particularly sensitive to trauma in K6α/K6β null mice, and establish the complete absence of keratin filaments in their anterior compartment. All null mice die about a week after birth. These studies demonstrate an essential structural role for K6 isoforms in the oral mucosa, and implicate filiform papillae as being the major stress bearing structures in dorsal tongue epithelium.
Sorting of Yeast Membrane Proteins into an Endosome-to-Golgi Pathway Involves Direct Interaction of Their Cytosolic Domains with Vps35p
Authors: Nothwehr Steven F. , Ha Seon-Ah , Bruinsma Paul
Published: July 2002

Resident late-Golgi membrane proteins in Saccharomyces cerevisiae are selectively retrieved from a prevacuolar–endosomal compartment, a process dependent on aromatic amino acid–based sorting determinants on their cytosolic domains. The formation of retrograde vesicles from the prevacuolar compartment and the selective recruitment of vesicular cargo are thought to be mediated by a peripheral membrane retromer protein complex. We previously described mutations in one of the retromer subunit proteins, Vps35p, which caused cargo-specific defects in retrieval. By genetic and biochemical means we now show that Vps35p directly associates with the cytosolic domains of cargo proteins. Chemical cross-linking, followed by coimmunoprecipitation, demonstrated that Vps35p interacts with the cytosolic domain of A-ALP, a model late-Golgi membrane protein, in a retrieval signal–dependent manner. Furthermore, mutations in the cytosolic domains of A-ALP and another cargo protein, Vps10p, were identified that suppressed cargo-specific mutations in Vps35p but did not suppress the retrieval defects of a vps35 null mutation. Suppression was shown to be due to an improvement in protein sorting at the prevacuolar compartment. These data strongly support a model in which Vps35p acts as a “receptor” protein for recognition of the retrieval signal domains of cargo proteins during their recruitment into retrograde vesicles.
Dnm1p Gtpase-Mediated Mitochondrial Fission Is a Multi-Step Process Requiring the Novel Integral Membrane Component Fis1p
Authors: Mozdy A.D. , McCaffery J.M. , Shaw J.M.
Published: July 2002

Yeast Dnm1p is a soluble, dynamin-related GTPase that assembles on the outer mitochondrial membrane at sites where organelle division occurs. Although these Dnm1p-containing complexes are thought to trigger constriction and fission, little is known about their composition and assembly, and molecules required for their membrane recruitment have not been isolated. Using a genetic approach, we identified two new genes in the fission pathway, FIS1 and FIS2. FIS1 encodes a novel, outer mitochondrial membrane protein with its amino terminus exposed to the cytoplasm. Fis1p is the first integral membrane protein shown to participate in a eukaryotic membrane fission event. In a related study (Tieu, Q., and J. Nunnari. 2000. J. Cell Biol. 151:353–365), it was shown that the FIS2 gene product (called Mdv1p) colocalizes with Dnm1p on mitochondria. Genetic and morphological evidence indicate that Fis1p, but not Mdv1p, function is required for the proper assembly and distribution of Dnm1p-containing fission complexes on mitochondrial tubules. We propose that mitochondrial fission in yeast is a multi-step process, and that membrane-bound Fis1p is required for the proper assembly, membrane distribution, and function of Dnm1p-containing complexes during fission.
Caspases Disrupt the Nuclear-Cytoplasmic Barrier
Authors: Faleiro Lavina , Lazebnik Yuri
Published: July 2002

During apoptosis, caspases, a family of proteases, disassemble a cell by cleaving a set of proteins. Caspase-3 plays a major role in the disassembly of the nucleus by processing several nuclear substrates. The question is how caspase-3, which is usually cytoplasmic, gains access to its nuclear targets. It was suggested that caspase-3 is actively transported to the nucleus through the nuclear pores. We found that caspase-9, which is activated earlier than caspase-3, directly or indirectly inactivates nuclear transport and increases the diffusion limit of the nuclear pores. This increase allows caspase-3 and other molecules that could not pass through the nuclear pores in living cells to enter or leave the nucleus during apoptosis by diffusion. Hence, caspase-9 contributes to cell disassembly by disrupting the nuclear-cytoplasmic barrier.
Skeletal Malformations Caused by Overexpression of Cbfa1 or Its Dominant Negative Form in Chondrocytes
Authors: Ueta Chisato , Iwamoto Masahiro , Kanatani Naoko , Yoshida Carolina , Liu Yang , Enomoto-Iwamoto Motomi , Ohmori Tomoharu , Enomoto Hirayuki , Nakata Ken , Takada Kenji , Kurisu Kojiro , Komori Toshihisa
Published: July 2002

During skeletogenesis, cartilage develops to either permanent cartilage that persists through life or transient cartilage that is eventually replaced by bone. However, the mechanism by which cartilage phenotype is specified remains unclarified. Core binding factor α1 (Cbfa1) is an essential transcription factor for osteoblast differentiation and bone formation and has the ability to stimulate chondrocyte maturation in vitro. To understand the roles of Cbfa1 in chondrocytes during skeletal development, we generated transgenic mice that overexpress Cbfa1 or a dominant negative (DN)-Cbfa1 in chondrocytes under the control of a type II collagen promoter/enhancer. Both types of transgenic mice displayed dwarfism and skeletal malformations, which, however, resulted from opposite cellular phenotypes. Cbfa1 overexpression caused acceleration of endochondral ossification due to precocious chondrocyte maturation, whereas overexpression of DN-Cbfa1 suppressed maturation and delayed endochondral ossification. In addition, Cbfa1 transgenic mice failed to form most of their joints and permanent cartilage entered the endochondral pathway, whereas most chondrocytes in DN-Cbfa1 transgenic mice retained a marker for permanent cartilage. These data show that temporally and spatially regulated expression of Cbfa1 in chondrocytes is required for skeletogenesis, including formation of joints, permanent cartilages, and endochondral bones.
Intramitochondrial Location and Dynamics of Crithidia fasciculata Kinetoplast Minicircle Replication Intermediates
Authors: Drew Mark E. , Englund Paul T.
Published: July 2002

Kinetoplast DNA, the mitochondrial DNA of Crithidia fasciculata, is organized into a network containing 5,000 topologically interlocked minicircles. This network, situated within the mitochondrial matrix, is condensed into a disk-shaped structure located near the basal body of the flagellum. Fluorescence in situ hybridization revealed that before their replication, minicircles are released vectorially from the network face nearest the flagellum. Replication initiates in the zone between the flagellar face of the disk and the mitochondrial membrane (we term this region the kinetoflagellar zone [KFZ]). The replicating minicircles then move to two antipodal sites that flank the disk-shaped network. In later stages of replication, the number of free minicircles increases, accumulating transiently in the KFZ. The final replication events, including primer removal, repair of many of the gaps, and reattachment of the progeny minicircles to the network periphery, are thought to take place within the antipodal sites.
PKCα regulates the hypertrophic growth of cardiomyocytes through extracellular signal–regulated kinase1/2 (ERK1/2)
Authors: Braz Julian C. , Bueno Orlando F. , De Windt Leon J. , Molkentin Jeffery D.
Published: July 2002

Members of the protein kinase C (PKC) isozyme family are important signal transducers in virtually every mammalian cell type. Within the heart, PKC isozymes are thought to participate in a signaling network that programs developmental and pathological cardiomyocyte hypertrophic growth. To investigate the function of PKC signaling in regulating cardiomyocyte growth, adenoviral-mediated gene transfer of wild-type and dominant negative mutants of PKCα, βII, δ, and ε (only wild-type ζ) was performed in cultured neonatal rat cardiomyocytes. Overexpression of wild-type PKCα, βII, δ, and ε revealed distinct subcellular localizations upon activation suggesting unique functions of each isozyme in cardiomyocytes. Indeed, overexpression of wild-type PKCα, but not βII, δ, ε, or ζ induced hypertrophic growth of cardiomyocytes characterized by increased cell surface area, increased [3H]-leucine incorporation, and increased expression of the hypertrophic marker gene atrial natriuretic factor. In contrast, expression of dominant negative PKCα, βII, δ, and ε revealed a necessary role for PKCα as a mediator of agonist-induced cardiomyocyte hypertrophy, whereas dominant negative PKCε reduced cellular viability. A mechanism whereby PKCα might regulate hypertrophy was suggested by the observations that wild-type PKCα induced extracellular signal–regulated kinase1/2 (ERK1/2), that dominant negative PKCα inhibited PMA-induced ERK1/2 activation, and that dominant negative MEK1 (up-stream of ERK1/2) inhibited wild-type PKCα–induced hypertrophic growth. These results implicate PKCα as a necessary mediator of cardiomyocyte hypertrophic growth, in part, through a ERK1/2-dependent signaling pathway.