bluuebunny/crossref_metadata_2025 huggingface dataset:
Role of Rab5 in the Recruitment of hVps34/p150 to the Early Endosome
Authors: Murray James T. , Panaretou Christina , Stenmark Harald , Miaczynska Marta , Backer Jonathan M.
Published: March 2003

PI 3‐kinases are important regulators of endocytic trafficking. We have previously proposed a model in which the Rab5 GTPase recruits EEA1 to the early endosome both directly, by binding to EEA1, and indirectly, through the recruitment of the p150/hVps34 PI 3‐kinase and the production of PI[3]P in the endosomal membrane. In this study we have examined this model in vivo. We find that both endogenous hVps34 and p150 are targeted to enlarged endosomal structures in cells expressing constitutively activated Rab5, where they are significantly colocalized with EEA1. Recombinant fragments of p150 disrupt the endosomal localization of EEA1, showing that p150 is required for EEA1 targeting. We further analyzed the mechanism of GTP‐dependent Rab5‐p150 binding, and showed the p150 HEAT and WD40 domains are required for binding, whereas deletion of the protein kinase domain increases binding to Rab5. Overexpression of constitutively active Rab5 caused a redistribution of epitope‐tagged hVps34 and p150 to Rab5‐positive endosomes. However, subcellular fractionation showed that this was not due to a significant recruitment of hVps34 or p150 from the cytosolic to the particulate fraction. These data suggest that the binding of Rab5 to the HEAT/WD40 domains of p150 is important in regulating the localization of hVps34/p150. However, Rab5 does not appear to act by directly recruiting p150/hVps34 complexes from the cytosol to the endosomal membrane.
Human Macrophages Accumulate HIV‐1 Particles in MHC II Compartments
Authors: Raposo Graça , Moore Marilyn , Innes Donald , Leijendekker Richtje , Leigh‐Brown Andrew , Benaroch Philippe , Geuze Hans
Published: March 2003

Macrophages are important targets for HIV‐1 infection and harbor the virions in an as yet unidentified organelle. To determine the location of HIV‐1 in these cells, an extensive analysis of primary human macrophages infected in vitro with HIV‐1 was carried out by immuno‐electron microscopy. Virus particles were found to accumulate in intracellular multivesicular compartments which were enriched in major histocompatibility complex class II molecules and CD63. These features are characteristics of major histocompatibility complex class II compartments where maturing class II molecules acquire their peptide cargo. The membrane‐delimited, electron‐dense virus particles of 100–110 nm diameter labeled strongly for HIV‐1 p24 antigen, major histocompatibility complex class II molecules, CD63 and, to a lesser extent for HIV‐1 gp120 envelope protein and Lamp 1. Our data suggest that virus particles may access the lumen of the major histocompatibility complex class II compartment by budding from the limiting membrane, thus acquiring proteins of this membrane such as class II and CD63. Viral assembly and budding would therefore occur in macrophages by a process similar to the formation of the internal vesicles in multivesicular bodies and at the same location. This could account for the particular content in lipids and proteins previously found in the membrane wrapping HIV particles. Our observations also suggest direct fusion of the virus containing major histocompatibility complex class II compartment with the plasma membrane, leading to massive release of viral particles into the extracellular medium.
Ligand‐Independent Degradation of Epidermal Growth Factor Receptor Involves Receptor Ubiquitylation and Hgs, an Adaptor Whose Ubiquitin‐Interacting Motif Targets Ubiquitylation by Nedd4
Authors: Katz Menachem , Shtiegman Keren , Tal‐Or Pazit , Yakir Liat , Mosesson Yaron , Harari Daniel , Machluf Yossi , Asao Hironobu , Jovin Thomas , Sugamura Kazuo , Yarden Yosef
Published: March 2003

Ligand‐dependent endocytosis of the epidermal growth factor receptor (EGFR) involves recruitment of a ubiquitin ligase, and sorting of ubiquitylated receptors to lysosomal degradation. By studying Hgs, a mammalian homolog of a yeast vacuolar‐sorting adaptor, we provide information on the less understood, ligand‐independent pathway of receptor endocytosis and degradation. Constitutive endocytosis involves receptor ubiquitylation and translocation to Hgs‐containing endosomes. Whereas the lipid‐binding motif of Hgs is necessary for receptor endocytosis, the ubiquitin‐interacting motif negatively regulates receptor degradation. We demonstrate that the ubiquitin‐interacting motif is endowed with two functions: it binds ubiquitylated proteins and it targets self‐ubiquitylation by recruiting Nedd4, an ubiquitin ligase previously implicated in endocytosis. Based upon the dual function of the ubiquitin‐interacting motif and its wide occurrence in endocytic adaptors, we propose a ubiquitin‐interacting motif network that relays ubiquitylated membrane receptors to lysosomal degradation through successive budding events.
Endosome Fusion and Microtubule‐Based Dynamics in the Early Endocytic Pathway of Dictyostelium
Authors: Clarke Margaret , Köhler Jana , Heuser John , Gerisch Günther
Published: March 2003

Dictyostelium amoebae, like mammalian macrophages, take up fluid by macropinocytosis. The present study used fluorescent fluid‐phase markers and GFP‐labeled microtubules to visualize the uptake, dynamics, and fusion of early endosomes in Dictyostelium. Consecutive labeling with two fluorescent fluid‐phase markers demonstrated that within the first few minutes after uptake, new macropinosomes underwent fusion with pre‐existing endosomes. The fusing endosomes, which represent the mixing compartment, displayed extreme shape changes and rapid transport about the cell in association with microtubules. The great plasticity of endosomes at this stage of maturation was also evident by electron microscopy. The constant undulatory motion of microtubules was implemental in establishing contact with endosomes. Treatment of cells with agents that selectively disrupted either actin filaments or microtubules confirmed that endosome dynamics were microtubule based. Further maturation of endosomes led to loss of pleiomorphy in favor of a spherical shape, inability to fuse with new macropinosomes, and diminished motility.
PrPC Is Sorted to the Basolateral Membrane of Epithelial Cells Independently of its Association with Rafts
Authors: Sarnataro Daniela , Paladino Simona , Campana Vincenza , Grassi Jacques , Nitsch Lucio , Zurzolo Chiara
Published: March 2003

PrPC is a glycosylphosphatidylinositol‐anchored protein expressed in neurons as well as in the cells of several peripheral tissues. Although the normal function of PrPC remains unknown, a conformational isoform called PrPSc (scrapie) has been proposed to be the infectious agent of transmissible spongiform encephalopathies in animals and humans. Where and how the PrPC to PrPSc conversion occurs in the cells is not yet known. Therefore, dissecting the intracellular trafficking of the wild‐type prion protein, as well as of the scrapie isoform, can be of major relevance to the pathogenesis of the diseases. In this report we have analyzed the exocytic pathway of transfected mouse PrPC in thyroid and kidney polarized epithelial cells. In contrast to the majority of glycosylphosphatidylinositol‐anchored proteins, we found that PrPC is localized mainly on the basolateral domain of the plasma membrane of both cell lines. This is reminiscent of the predominant somatodendritic localization found in neurons. However, similarly to apical glycosylphosphatidylinositol‐proteins, PrPC associates with detergent‐resistant microdomains, which have been suggested to have a role in apical sorting of glycosylphosphatidylinositol‐proteins, as well as in the conversion process of PrPC to PrPSc. In order to discriminate whether detergent‐resistant microdomains have a direct role in PrPSc conversion, or whether they are involved in the transport of the protein to the site of its conversion, we have examined the effect of disruption of detergent‐resistant microdomain association on PrPC intracellular traffic. Consistent with the unusual basolateral localization of this glycosylphosphatidylinositol‐linked protein, our data exclude a classical role for detergent‐resistant microdomains in the post‐trans‐Golgi network sorting and transport of PrPC to the plasma membrane.
Arf Regulates Interaction of GGA with Mannose‐6‐Phosphate Receptor
Authors: Hirsch Dianne Snow , Stanley Katherine T. , Chen Ling‐Xin , Jacques Kerry M. , Puertollano Rosa , Randazzo Paul A.
Published: March 2003

The role of ADP‐ribosylation factor (Arf) in Golgi associated, γ‐adaptin homologous, Arf‐interacting protein (GGA)‐mediated membrane traffic was examined. GGA is a clathrin adaptor protein that binds Arf through its GAT domain and the mannose‐6‐phosphate receptor through its VHS domain. The GAT and VHS domains interacted such that Arf and mannose‐6‐phosphate receptor binding to GGA were mutually exclusive.In vivo, GGA bound membranes through either Arf or mannose‐6‐phosphate receptor. However, mannose‐6‐phosphate receptor excluded Arf from GGA‐containing structures outside of the Golgi. These data are inconsistent with predictions based on the model for Arf's role in COPI veside coat function. We propose that Arf recruits GGA to a membrane and then, different from the current model, ‘hands‐off’ GGA to mannose‐6‐phosphate receptor. GGA and mannose‐6‐phosphate receptor are then incorporated into a transport intermediate that excludes Arf.
Changes in the prevalence of HIV‐associated mucosal disease at a dedicated clinic over 7 years
Authors: Greenwood I , Zakrzewska JM , Robinson PG
Published: March 2003

OBJECTIVE: To investigate changes in the prevalence of mucosal disease in patients with HIV/AIDS.

DESIGN: Retrospective analysis of clinic database.

SETTING: An open‐access dental clinic dedicated to the dental treatment of patients infected with HIV in London, UK.

SUBJECTS: Dental patients attending for routine assessment (check‐up).

MAIN OUTCOME MEASURES: Prevalence of HIV associated oral lesions, diagnosed in accordance with EC classification criteria, in patients taking and not taking antiretroviral therapies (ARTs).

RESULTS: Data were collected at 2789 dental examinations for 1590 patients between 1992 and 1998. More patients taking ART had one or more oral manifestations in the years up to 1996, after which the prevalence fell to a similar level to those not taking medications. The prevalence of mucosal disease in patients not taking ART was relatively constant over time. These trends were observed for oral hairy leukoplakia, pseudomembranous and erythematous candidiasis and Kaposi's sarcoma but not for oral papillomata.

CONCLUSIONS: These data provide evidence that decreases in the prevalence of oral disease in people with HIV infection can be attributed to the effectiveness of ART.
A collaborative approach for improving information literacy skills of dental hygiene students
Authors: Cobban Sandra J. , Seale Linda N.
Published: March 2003

Abstract 

Background: Evidence‐based practitioners need to develop proficiency in locating and evaluating necessary information from a variety of sources, a concept known as information literacy. One potential source of information is the World Wide Web; however, the wide range in the quality of information available through the Internet indicates a need for strong critical appraisal skills.

Objectives:To facilitate development of these skills in dental hygiene students, dental hygiene faculty collaborated with a librarian to implement a project to improve information literacy skills and integrate the use of the Internet into dental hygiene course work.

Methods:Students were required to develop a scenario about an aspect of dental hygiene practice, such as client health concern, practice management, or selection of an appropriate dental hygiene therapy. Students were to search the Internet, locate and evaluate a website that could address their scenario, and post this information and additional comments to a text‐based computer conferencing section of a class website. WebCT software, particularly the computer conferencing function, was used to support asynchronous online activities. Student evaluation of the assignment was conducted using a 5‐point Likert scale.

Results:Mean ratings for structure variables ranged from 3.94 to 3.81. Slightly lower mean ratings for process variables, ranging from 3.74 to 3.21, suggest room for improvement to activities. Mean ratings for outcome variables, ranging from 4.42 to 3.29, suggest achievement of the desired learning outcomes.

Conclusion:Evaluation results warrant the continued inclusion of asynchronous online activities in conjunction with face‐to‐face classroom instruction.
Value of Exercise Capacity and Heart Rate Recovery in Older People
Authors: Messinger‐Rapport Barbara , Snader Claire E. Pothier , Blackstone Eugene H. , Yu David , Lauer Michael S.
Published: March 2003

OBJECTIVES:To evaluate the prognostic value in older adults of two predictors of mortality: impaired functional capacity and an attenuated heart rate recovery.

SETTING:Academic medical center.

DESIGN:Prospective study with mean 3.7 years follow‐up.

PARTICIPANTS:Seven thousand three hundred fifty‐four adults aged 65 and older consecutively referred for exercise testing between 1990 and 1999. Patients with heart failure, valvular disease, atrial fibrillation, and pacemakers were excluded.

MEASUREMENTS:The primary endpoint was all‐cause mortality. Impaired functional capacity was defined as the peak exercise workload in the lowest quintile of metabolic equivalents achieved according to prespecified strata of age and sex. Heart rate recovery was defined as the fall in heart rate during the first minute after exercise and was abnormal if 12 or fewer beats per minute, except for patients undergoing stress echocardiography, in which case 18 or fewer beats per minute was abnormal.

RESULTS:There were 842 deaths. Patients with impaired functional capacity were at increased risk for death (23% vs 9%, hazard ratio (HR) = 2.7, 95% confidence interval (CI) = 2.2–3.1,P< .0001) as were patients with an abnormal heart rate recovery (17% vs 9%, HR = 2.0, 95% CI = 1.8–2.3,P< .0001). After adjusting for age, sex, coronary history, and other confounders, impaired functional capacity (adjusted HR = 2.1, 95% CI = 1.8–2.4) and an abnormal heart rate recovery (adjusted HR = 1.5, 95% CI = 1.3–1.7) independently predicted death. No interactions between these two variables with age were noted.

CONCLUSIONS:In older patients, impaired functional capacity and heart rate recovery were independent predictors of death.
The MDS‐CHESS Scale: A New Measure to Predict Mortality in Institutionalized Older People
Authors: Hirdes John P. , Frijters Dinnus H. , Teare Gary F.
Published: March 2003

OBJECTIVES: To develop a scale predicting mortality and other adverse outcomes associated with frailty.

DESIGN: Observational study based on Minimum Data Set (MDS) 2.0 and mortality data.

SETTING: Ontario chronic hospitals.

PARTICIPANTS: All chronic hospital patients (N = 28,495) assessed with the MDS 2.0 after mandatory implementation in July 1996 followed until May 1999.

MEASUREMENTS: MDS 2.0 assessments done as part of normal practice mainly by registered nurses or multidisciplinary teams in a chronic hospital. Mortality data are available from the accompanying discharge tracking form.

RESULTS: The MDS‐Changes in Health, End‐stage disease and Symptoms and Signs (CHESS) score is a composite measure addressing changes in health, end‐stage disease, and symptoms and signs of medical problems. It is a strong predictor of mortality (P < .0001) independent of the effects of age, sex, activities of daily living impairment, cognition, and do‐not‐resuscitate orders. It is also strongly associated with physician activity, complex medical procedures, and pain (P < .001 for each dependent variable).

CONCLUSIONS: The CHESS score provides a useful new MDS‐based test to predict mortality and to measure instability in health as a clinical outcome.
Testosterone Supplementation Therapy for Older Men: Potential Benefits and Risks
Authors: Gruenewald David A. , Matsumoto Alvin M.
Published: March 2003

Serum testosterone levels decline gradually and progressively with aging in men. Many manifestations associated with aging in men, including muscle atrophy and weakness, osteoporosis, reduced sexual functioning, and increased fat mass, are similar to changes associated with testosterone deficiency in young men. These similarities suggest that testosterone supplementation may prevent or reverse the effects of aging. A MEDLINE search was performed to identify studies of testosterone supplementation therapy in older men. A structured, qualitative review was performed of placebo‐controlled trials that included men aged 60 and older and evaluated one or more physical, cognitive, affective, functional, or quality‐of‐life outcomes. Studies focusing on patients with severe systemic diseases and hormone deficiencies related to specific diseases were excluded.

In healthy older men with low‐normal to mildly decreased testosterone levels, testosterone supplementation increased lean body mass and decreased fat mass. Upper and lower body strength, functional performance, sexual functioning, and mood were improved or unchanged with testosterone replacement. Variable effects on cognitive function were reported, with improvements in some cognitive domains (e.g., spatial, working, and verbal memory). Testosterone supplementation improved exercise‐induced coronary ischemia in men with coronary heart disease, whereas angina pectoris was improved or unchanged. In a few studies, men with low testosterone levels were more likely to experience improvements in lumbar bone mineral density, self‐perceived functional status, libido, erectile function, and exercise‐induced coronary ischemia with testosterone replacement than men with less marked testosterone deficiency. No major unfavorable effects on lipids were reported, but hematocrit and prostate specific antigen levels often increased.

Based on these results, testosterone supplementation cannot be recommended at this time for older men with normal or low‐normal testosterone levels and no clinical manifestations of hypogonadism. However, testosterone replacement may be warranted in older men with markedly decreased testosterone levels, regardless of symptoms, and in men with mildly decreased testosterone levels and symptoms or signs suggesting hypogonadism. The long‐term safety and efficacy of testosterone supplementation remain uncertain. Establishment of evidence‐based indications will depend on further demonstrations of favorable clinical outcomes and symptomatic, functional, and quality‐of‐life benefits in carefully performed, long‐term, randomized, placebo‐controlled clinical trials. J Am Geriatr Soc 51:101–115, 2003.
Bidirectional DNA unwinding by a ternary complex of T antigen, nucleolin and topoisomerase I
Authors: Seinsoth Stephanie , Uhlmann‐Schiffler Heike , Stahl Hans
Published: March 2003

The simian virus 40 large tumour‐antigen (T antigen) DNA helicase is a hexameric structure; it has been proposed that, in viral DNA replication, two of these hexamers are combined to form a bipartite holoenzyme that acts concurrently at both forks of a replication bubble. In a search for structural components of this helicase complex, we have identified nucleolin as a specific binding protein for the T‐antigen hexamer. We show that nucleolin, in co‐operation with human topoisomerase I, mediates the cohesion of the T‐antigen helicase holoenzyme during plasmid unwinding. Our results provide biochemical evidence for a direct role of nucleolin in DNA replication, in addition to its known function in ribosome biogenesis. The data presented here suggest that nucleolin enables the formation of a functional ‘helicase‐swivelase’ complex at the replication fork.
The adapter protein APS associates with the multifunctional docking sites Tyr-568 and Tyr-936 in c-Kit
Authors: WOLLBERG Patrik , LENNARTSSON Johan , GOTTFRIDSSON Eva , YOSHIMURA Akihiko , RÖNNSTRAND Lars
Published: March 2003

The adapter protein APS has previously been shown to be involved in recruiting the ubiquitin E3 ligase c-Cbl to the insulin receptor, the platelet-derived growth factor β-receptor and the erythropoietin receptor, leading to increased degradation of the receptors and inhibition of mitogenesis. Here we demonstrate, by use of immobilized synthetic phosphopeptides corresponding to various autophosphorylated tyrosine residues in the receptor for stem-cell factor (c-Kit), that APS preferentially associates with phosphorylated Tyr-568 and Tyr-936. Tyr-568 has previously been identified as the binding site of the Src family of tyrosine kinases, the Csk-homologous kinase CHK, and the protein tyrosine phosphatase SHP-2. We have recently demonstrated that Tyr-936 is an autophosphorylation site involved in binding the adapter proteins Grb2 and Grb7. We could further demonstrate that the critical determinant for binding of APS is the presence of either a leucine or an isoleucine residue in the position +3 to the phosphorylated tyrosine. This allowed us to design mutants that selectively failed to associate with APS, while still associating with Src family members, SHP-2 and Grb2, respectively.
Novel splice variants of the receptor for advanced glycation end-products expressed in human vascular endothelial cells and pericytes, and their putative roles in diabetes-induced vascular injury
Authors: YONEKURA Hideto , YAMAMOTO Yasuhiko , SAKURAI Shigeru , PETROVA Ralica G. , ABEDIN Md. Joynal , LI Hui , YASUI Kiyoshi , TAKEUCHI Masayoshi , MAKITA Zenji , TAKASAWA Shin , OKAMOTO Hiroshi , WATANABE Takuo , YAMAMOTO Hiroshi
Published: March 2003

The binding of advanced glycation end-products (AGE) to the receptor for AGE (RAGE) is known to deteriorate various cell functions and is implicated in the pathogenesis of diabetic vascular complications. In the present study, we show that the cellular constituents of small vessels, endothelial cells (EC) and pericytes express novel splice variants of RAGE mRNA coding for the isoforms that lack the N-terminal V-type immunoglobulin-like domain (N-truncated) or the C-terminal transmembrane domain (C-truncated), as well as the known full-length mRNA. The ratio of the expression of the three variants was different between EC and pericytes; the content of the C-truncated form was highest in EC, whereas the full-length form was the most abundant in pericytes. Transfection experiments with COS-7 cells demonstrated that those variant mRNAs were translated into proteins as deduced; C-truncated RAGE was efficiently secreted into the culture media, and N-truncated RAGE was located mainly on the plasma membrane. The three isoforms were also detected in primary cultured human EC and pericytes. Further, full-length and C-truncated forms of RAGE bound to an AGE-conjugated column, whereas N-truncated RAGE did not. The AGE induction of extracellular-signal-related kinase phosphorylation and vascular endothelial growth factor in EC and of the growth and cord-like structure formation of EC was abolished completely by C-truncated RAGE, indicating that this endogenous secretory receptor (endogenous secretory RAGE) is cytoprotective against AGE. The results may contribute to our understanding of the molecular basis for the diversity of cellular responses to AGE and for individual variations in the susceptibility to diabetic vascular complications.
Early stimulation and late inhibition of peroxisome proliferator-activated receptor gamma (PPARgamma) gene expression by transforming growth factor beta in human aortic smooth muscle cells: role of early growth-response factor-1 (Egr-1), activator protein 1 (AP1) and Smads
Authors: FU Mingui , ZHANG Jifeng , LIN Yimin , ZHU Xiaojun , ZHAO Luning , AHMAD Mushtaq , EHRENGRUBER Markus U. , CHEN Yuqing E.
Published: March 2003

Transforming growth factor β (TGFβ) and peroxisome proliferator-activated receptor γ (PPARγ) play major roles in the development of vascular diseases. It has been documented that PPARγ activation inhibits the TGFβ signal pathway in vascular smooth muscle cells (VSMC). Here we examined whether TGFβ can regulate PPARγ expression. Northern blot analyses revealed that both TGFβ1 and 2 exert a biphasic effect (early stimulation and late repression) on PPARγ gene expression in VSMC. TGFβ rapidly and transiently induced early growth-response factor-1 (Egr-1) expression through the mitogen-activated protein kinase/extracellular signal-regulated kinase kinase 1 (MEK1)/ERK-mediated pathway. Inhibition of MEK1/ERK by PD98059 not only abrogated the induction of Egr-1 but also abolished the rapid and transient induction of PPARγ by TGFβ. Furthermore, overexpression of NAB2, a repressor of Egr-1 activation, also blocked the induction of PPARγ by TGFβ in VSMC, suggesting that Egr-1 mediates the rapid and transient induction of PPARγ by TGFβ. With regard to the TGFβ repression of PPARγ expression, activator protein 1 (AP1) and Smad3/4 dramatically inhibited the PPARγ promoter activity in transient-transfection studies. In contrast, adenovirus-mediated overexpression of a dominant-negative form of c-Jun partially rescued the TGFβ-induced PPARγ repression in VSMC. Taken together, our data demonstrate that Egr-1, AP1 and Smad are part components of the TGFβ signal transduction pathway that regulates PPARγ expression.
Noradrenaline and Dopamine Regulation of Prolactin Secretion in Sheep: Role in Prolactin Homeostasis but not Photoperiodism
Authors: Lincoln G. A. , Clarke I. J.
Published: March 2003

The role of noradrenaline (NA) and dopamine (DA) in the hypothalamic control of prolactin (PRL) secretion was investigated in hypothalamic intact (control) and hypothalamo‐pituitary disconnected (HPD) Soay rams. The animals were exposed to alternating 16‐weekly periods of short (8 L : 16D) and long days (16 L : 8D) to induce marked cyclical changes in PRL secretion in both groups (as demonstrated previously). Selective NA and DA receptor antagonists (dose: 1.2 µmol/kg) were administered under short days (low endogenous PRL secretion), and agonists (dose: 0.0012–0.12 µmol/kg) were administered under long days (high endogenous PRL secretion). The acute changes in blood PRL concentrations were measured over 4 h as the index of responsiveness. Under short days, treatment with WB4101 (α‐1 adenoceptor antagonist), and rauwolscine (α‐2 antagonist), consistently increased PRL secretion in control, but not in HPD rams. The treatments produced similar acute, drug‐specific behavioural effects in both groups. Propranolol (β antagonist) had no effect on PRL secretion, while sulpiride (DA D‐2 antagonist) induced a marked increase in blood PRL concentrations in control rams (> 4 h), and a transient effect in HPD rams (15 min). Under long days, when endogenous PRL secretion was increased, phenylephrine (α‐1 agonist) produced no effects, while bromocriptine (DA D‐2 agonist) robustly decreased PRL concentrations in both control and HPD rams, even at the lowest treatment dose. Overall, the positive responses to the antagonists in the control rams, support the view that DA (acting via D‐2 receptors), and to a lesser extent NA (acting via α‐1/α‐2 receptors), negatively regulate PRL secretion. In contrast, the lack of responses to the antagonists in the HPD rams, support the view that neither DA, nor NA, mediate the photoperiodic control of PRL secretion.
Decreased Corticotropin‐Releasing Factor Receptor Expression and Adrenocorticotropic Hormone Responsiveness in Anterior Pituitary Cells of Wistar‐Kyoto Rats
Authors: Hauger R. L. , Shelat S. G. , Redei E. E.
Published: March 2003

The Wistar‐Kyoto (WKY) rat shows signs of persistent activation of the hypothalamic‐pituitary‐adrenal axis, but the cause and site of this activation is not yet known. Chronically activated corticotrophs generally show blunted adrenocorticotropic hormone (ACTH) response to corticotropin releasing factor (CRF); therefore, the anterior pituitary responsiveness to ACTH secretagogues, CRF and vasopressin, was compared in male WKY and Wistar rats. Anterior pituitary CRF binding and CRF receptor mRNA expression was significantly decreased in WKY rats. ACTH response to CRF or vasopressin was markedly impaired, and vasopressin failed to potentiate the CRF‐stimulated ACTH release in cultured WKY anterior pituitary cells. In contrast, CRF and vasopressin alone and in combination stimulated large, concentration‐dependent increases in ACTH release in Wistar anterior pituitary cells. By contrast to the decreased ACTH secretory responses, steady‐state anterior pituitary pro‐opiomelanocortin mRNA levels were approximately 12‐fold greater in WKY rats compared to Wistar rats, and they further increased in response to CRF stimulation. These findings suggest that, although the WKY rat corticotroph is under a chronic state of activation or disinhibition, the in vitro secretory responses to classic ACTH secretagogues are impaired.
Waterlogging tolerance in the tribe Triticeae: the adventitious roots ofCritesion marinumhave a relatively high porosity and a barrier to radial oxygen loss
Authors: Mcdonald M. P. , Galwey N. W. , Colmer T. D.
Published: March 2003

Nine species from the tribe Triticeae – three crop, three pasture and three ‘wild’ wetland species – were evaluated for tolerance to growth in stagnant deoxygenated nutrient solution and also for traits that enhance longitudinal O2movement within the roots.Critesion marinum(syn.Hordeum marinum) was the only species evaluated that had a strong barrier to radial O2loss (ROL) in the basal regions of its adventitious roots. Barriers to ROL have previously been documented in roots of several wetland species, although not in any close relatives of dryland crop species. Moreover, the porosity in adventitious roots ofC. marinumwas relatively high: 14% and 25% in plants grown in aerated and stagnant solutions, respectively. The porosity ofC. marinumroots in the aerated solution was 1·8–5·4‐fold greater, and in the stagnant solution 1·2–2·8‐fold greater, than in the eight other species when grown under the same conditions. These traits presumably contributed toC. marinumhaving a 1·4–3 times greater adventitious root length than the other species when grown in deoxygenated stagnant nutrient solution or in waterlogged soil. The length of the adventitious roots and ROL profiles ofC. marinumgrown in waterlogged soil were comparable to those of the extremely waterlogging‐tolerant speciesEchinochloa crus‐galliL. (P. Beauv.). The superior tolerance ofC. marinum, as compared toHordeum vulgare(the closest cultivated relative), was confirmed in pots of soil waterlogged for 21 d;H. vulgaresuffered severe reductions in shoot and adventitious root dry mass (81% and 67%, respectively), whereasC. marinumshoot mass was only reduced by 38% and adventitious root mass was not affected.
Photosynthesis, light and nitrogen relationships in a young deciduous forest canopy under open‐air CO2 enrichment
Authors: Takeuchi Y. , Kubiske M. E. , Isebrands J. G. , Pregtizer K.S. , Hendrey G. , Karnosky D. F.
Published: March 2003

Leaf photosynthesis (Ps), nitrogen (N) and light environment were measured on Populus tremuloides trees in a developing canopy under free‐air CO2 enrichment in Wisconsin, USA. After 2 years of growth, the trees averaged 1·5 and 1·6 m tall under ambient and elevated CO2, respectively, at the beginning of the study period in 1999. They grew to 2·6 and 2·9 m, respectively, by the end of the 1999 growing season. Daily integrated photon flux from cloud‐free days (PPFDday,sat) around the lowermost branches was 16·8 ± 0·8 and 8·7 ± 0·2% of values at the top for the ambient and elevated CO2 canopies, respectively. Elevated CO2 significantly decreased leaf N on a mass, but not on an area, basis. N per unit leaf area was related linearly to PPFDday,sat throughout the canopies, and elevated CO2 did not affect that relationship. Leaf Ps light‐response curves responded differently to elevated CO2, depending upon canopy position. Elevated CO2 increased Pssat only in the upper (unshaded) canopy, whereas characteristics that would favour photosynthesis in shade were unaffected by elevated CO2. Consequently, estimated daily integrated Ps on cloud‐free days (Psday,sat) was stimulated by elevated CO2 only in the upper canopy. Psday,sat of the lowermost branches was actually lower with elevated CO2 because of the darker light environment. The lack of CO2 stimulation at the mid‐ and lower canopy was probably related to significant down‐regulation of photosynthetic capacity; there was no down‐regulation of Ps in the upper canopy. The relationship between Psday,sat and leaf N indicated that N was not optimally allocated within the canopy in a manner that would maximize whole‐canopy Ps or photosynthetic N use efficiency. Elevated CO2 had no effect on the optimization of canopy N allocation.
Acclimation of photosynthetic capacity to irradiance in tree canopies in relation to leaf nitrogen concentration and leaf mass per unit area
Authors: Meir P. , Kruijt B. , Broadmeadow M. , Barbosa E. , Kull O. , Carswell F. , Nobre A. , Jarvis P. G.
Published: March 2003

The observation of acclimation in leaf photosynthetic capacity to differences in growth irradiance has been widely used as support for a hypothesis that enables a simplification of some soil‐vegetation‐atmosphere transfer (SVAT) photosynthesis models. The acclimation hypothesis requires that relative leaf nitrogen concentration declines with relative irradiance from the top of a canopy to the bottom, in 1 : 1 proportion. In combination with a light transmission model it enables a simple estimate of the vertical profile in leaf nitrogen concentration (which is assumed to determine maximum carboxylation capacity), and in combination with estimates of the fraction of absorbed radiation it also leads to simple ‘big‐leaf’ analytical solutions for canopy photosynthesis. We tested how forests deviate from this condition in five tree canopies, including four broadleaf stands, and one needle‐leaf stand: a mixed‐species tropical rain forest, oak (Quercus petraea(Matt.) Liebl), birch (Betula pendulaRoth), beech (Fagus sylvaticaL.) and Sitka spruce (Picea sitchensis(Bong.) Carr). Each canopy was studied when fully developed (mid‐to‐late summer for temperate stands). Irradiance (Q,µmol m−2 s−1) was measured for 20 d using quantum sensors placed throughout the vertical canopy profile. Measurements were made to obtain parameters from leaves adjacent to the radiation sensors: maximum carboxylation and electron transfer capacity (Va,Ja,µmol m−2 s−1), day respiration (Rda,µmol m−2 s−1), leaf nitrogen concentration (Nm, mg g−1) and leaf mass per unit area (La, g m−2).

Relative to upper‐canopy values,Vadeclined linearly in 1 : 1 proportion withNa. RelativeVaalso declined linearly with relativeQ, but with a significant intercept at zero irradiance (P < 0·01). This intercept was strongly related toLaof the lowest leaves in each canopy (P < 0·01,r2 = 0·98,n= 5). For each canopy, daily lnQwas also linearly related with lnVa(P < 0·05), and the intercept was correlated with the value for photosynthetic capacity per unit nitrogen (PUN:Va/Na,µmol g−1s−1) of the lowest leaves in each canopy (P < 0·05).Vawas linearly related withLaandNa(P < 0·01), but the slope of theVa : Narelationship varied widely among sites. Hence, whilst there was a uniqueVa : Naratio in each stand, acclimation inVatoQvaried predictably withLaof the lowest leaves in each canopy. The specific leaf area,Lm(cm2g−1), of the canopy‐bottom foliage was also found to predict carboxylation capacity (expressed on a mass basis;Vm,µmol g−1 s−1) at all sites (P < 0·01). These results invalidate the hypothesis of full acclimation to irradiance, but suggest thatLaandLmof the most light‐limited leaves in a canopy are widely applicable indicators of the distribution of photosynthetic capacity with height in forests.
Photosynthetic carbon assimilation and associated metabolism in relation to water deficits in higher plants
Authors: Lawlor D. W. , Cornic G.
Published: March 2003

Experimental studies on CO2 assimilation of mesophytic C3 plants in relation to relative water content (RWC) are discussed. Decreasing RWC slows the actual rate of photosynthetic CO2 assimilation (A) and decreases the potential rate (Apot). Generally, as RWC falls from c. 100 to c. 75%, the stomatal conductance (gs) decreases, and with it A. However, there are two general types of relation of Apot to RWC, which are called Type 1 and Type 2. Type 1 has two main phases. As RWC decreases from 100 to c. 75%, Apot is unaffected, but decreasing stomatal conductance (gs) results in smaller A, and lower CO2 concentration inside the leaf (Ci) and in the chloroplast (Cc), the latter falling possibly to the compensation point. Down‐regulation of electron transport occurs by energy quenching mechanisms, and changes in carbohydrate and nitrogen metabolism are considered acclimatory, caused by low Ci and reversible by elevated CO2. Below 75% RWC, there is metabolic inhibition of Apot, inhibition of A then being partly (but progressively less) reversible by elevated CO2; gs regulates A progressively less, and Ci and CO2 compensation point, Γ rise. It is suggested that this is the true stress phase, where the decrease in Apot is caused by decreased ATP synthesis and a consequent decreased synthesis of RuBP. In the Type 2 response, Apot decreases progressively at RWC 100 to 75%, with A being progressively less restored to the unstressed value by elevated CO2. Decreased gs leads to a lower Ci and Cc but they probably do not reach compensation point: gs becomes progressively less important and metabolic limitations more important as RWC falls. The primary effect of low RWC on Apot is most probably caused by limited RuBP synthesis, as a result of decreased ATP synthesis, either through inhibition of Coupling Factor activity or amount due to increased ion concentration. Carbohydrate synthesis and accumulation decrease. Type 2 response is considered equivalent to Type 1 at RWC below c. 75%, with Apot inhibited by limited ATP and RuBP synthesis, respiratory metabolism dominates and Ci and Γ rise. The importance of inhibited ATP synthesis as a primary cause of decreasing Apot is discussed. Factors determining the Type 1 and Type 2 responses are unknown. Electron transport is maintained (but down‐regulated) in Types 1 and 2 over a wide range of RWC, and a large reduced/oxidized adenylate ratio results. Metabolic imbalance results in amino acid accumulation and decreased and altered protein synthesis. These conditions profoundly affect cell functions and ultimately cause cell death. Type 1 and 2 responses may reflect differences in gs and in sensitivity of metabolism to decreasing RWC.
Identification of canine helper T‐cell epitopes from the fusion protein of canine distemper virus
Authors: Ghosh Souravi , Walker John , Jackson David C.
Published: March 2003

The fusion protein of canine distemper virus (CDV‐F), a 662 amino‐acid envelope protein, was used as the target molecule for identification of canine T helper (Th) epitopes. A library of 94 peptides, each 17 residues in length overlapping by 10 residues and covering the entire sequence of CDV‐F, was screened using a lymphocyte proliferation assay with peripheral blood mononuclear cells (PBMC) obtained from dogs inoculated with canine distemper virus (CDV) vaccine. Initially we observed low and inconsistent proliferation of PBMC in response to these peptides, even when using cells obtained from dogs that had received multiple doses of CDV. Subsequently, the use of expanded cell populations derived by in vitro stimulation of canine PBMC with pools of peptides allowed the identification of a number of putative canine Th‐epitopes within the protein sequence of CDV‐F. There were two major clusters of Th‐epitopes identified close to the cleavage site of the F0 fusion protein, while some others were scattered in both the F1 and F2 fragments of the protein. Some of these peptides, in particular peptide 35 (p35), were stimulatory in dogs of different breeds and ages. The identification of such promiscuous canine Th‐epitopes encouraged us to assemble p35 in tandem with luteinising hormone releasing hormone (LHRH) a 10 amino‐acid residue synthetic peptide representing a B‐cell epitope which alone induces no antibody in dogs. The totally synthetic immunogen was able to induce the production of very high titres of antibodies against LHRH in all dogs tested. These results indicate that p35 could be an ideal candidate for use as a Th‐epitope for use in outbred dogs.
Signalling pathways induced by protease‐activated receptors and integrins in T cells
Authors: Bar‐Shavit Rachel , Maoz Miriam , Yongjun Yin , Groysman Maya , Dekel Idit , Katzav Shulamit
Published: March 2003

Recent characterization of the thrombin receptor indicates that it plays a role in T‐cell signalling pathways. However, little is known regarding the signalling events following stimulation of additional members of the protease‐activated receptor (PAR) family, i.e. PAR2 and PAR3. Most of the postligand cascades are largely unknown. Here, we illustrate that in Jurkat T‐leukaemic cells, activation of PAR1, PAR2 and PAR3 induce tyrosine phosphorylation of Vav1. This response was impaired in Jurkat T cells deficient in p56lck (JCaM1.6). Activation of PARs also led to an increase in tyrosine phosphorylation of ZAP‐70 and SLP‐76, two key proteins in T‐cell receptor (TCR) signalling. We also demonstrated that p56lck is meaningful for integrin signalling. Thus, JCaM1.6 cells exhibited a marked reduction in their adherence to fibronectin‐coated plates, as compared to the level of adherence of Jurkat T cells. While the phosphorylation of Vav1 in T cells is augmented following adhesion, no additional increase was noted following treatment of the adhered cells with PARs. Altogether, we have identified key components in the postligand‐signalling cascade of PARs and integrins. Furthermore, we have identified Lck as a critical and possibly upstream component of PAR‐induced Vav1 phosphorylation, as well as integrin activation, in Jurkat T cells.
On the development of the shoulder girdle in Crocidura russula (Soricidae) and other placental mammals: evolutionary and functional aspects
Authors: Großmann Martin , Sánchez‐Villagra Marcelo R. , Maier Wolfgang
Published: March 2003

The development of the shoulder girdle was studied in embryonic stages and a neonate of Crocidura russula using histological sections and 3‐D reconstructions. Neonatal stages of Suncus etruscus and Mesocricetus auratus, both altricial placentals, were also studied. The earliest stage of C. russula, in which the scapula is still partially blastematous, has already a supraspinous fossa. The dorsal portion of the scapular spine does not develop from the anterior margin of the scapula. Its mode of development varies among the placentals studied to date. In some it is completely appositional bone, in others it consists of bone formed mostly by endochondral ossification of a dorsal cartilaginous process stemming from the acromium. During development the supraspinatus muscle increases in size in proportion to the infraspinatus muscle and the humeral head increases in size in relation to the glenoid fossa. Placentals have secondary cartilage in the sternal and acromial ends of the clavicle, a derived feature absent in Marsupialia. Even the most altricial placentals have a more developed shoulder girdle at birth than any newborn marsupial studied to date.
Climate and population density induce long‐term cohort variation in a northern ungulate
Authors: Forchhammer Mads C. , Clutton‐Brock Tim H. , Lindström Jan , Albon Steve D.
Published: March 2003

Density‐dependent and climatic conditions experienced by individuals before and after birth differ considerably between cohorts. Such early environmental variability has the potential to create persistent fitness differences among cohorts. Here we test the hypothesis that conditions experienced by individuals in their early development will have long‐term effects on their life history traits.

 We approached this by analysing and contrasting the effects of climate (the North Atlantic Oscillation, NAO) and population density at year of birth on cohort birth weight, birth date, litter size, age of maturity, survival and fecundity of Soay sheep, Ovies aries L., ewes in the population on the island of Hirta, St Kilda, Scotland.

 Significant intercohort variations were found in life history traits. Cohorts born after warm, wet and windy (high NAO) winters were lighter at birth, born earlier, less likely to have a twin and matured later than cohorts born following cold and dry (low NAO) winters. High population densities in the winter preceding birth also had a negative effect on birth weight, birth date and litter size, whereas high postnatal densities delayed age of first reproduction.

 High NAO winters preceding birth depressed juvenile survival but increased adult survival and fecundity. The negative influence of high NAO winters on juvenile survival is likely to be related to mothers’ compromised physical condition while the cohort is in utero, whereas the positive influence on adult survival and fecundity may relate to the improved postnatal forage conditions following high NAO winters. High pre‐ and postnatal population densities decreased juvenile (neonatal, yearling) and adult (2–4 years) survivorship but had no significant effect fecundity.
Density dependence and colony growth in the ant speciesFormica neorufibarbis
Authors: Billick Ian
Published: March 2003

Production of cocoons and numbers of workers in 25–40 colonies of the ant speciesFormica neorufibarbiswere tracked from 1993 to 1998 in order to test for density dependence in either production or survivorship of workers.

The percentage of workers lost by colonies over winter increased with increasing worker numbers.

There was no evidence that worker production was density‐dependent.

Food supplementation did not affect worker‐cocoon production.

Colonies forced to live under small nest rocks produced fewer cocoons, but this effect was never seen under natural conditions.

In the field, colonies preferentially utilized large rocks for nests and worker number was positively correlated with nest‐rock area.
Root proliferation in decaying roots and old root channels: a nutrient conservation mechanism in oligotrophic mangrove forests?
Authors: McKee Karen L.
Published: March 2003

In oligotrophic habitats, proliferation of roots in nutrient‐rich microsites may contribute to overall nutrient conservation by plants. Peat‐based soils on mangrove islands in Belize are characterized by the presence of decaying roots and numerous old root channels (0.1–3.5 cm diameter) that become filled with living and highly branched roots ofRhizophora mangleandAvicennia germinans. The objectives of this study were to quantify the proliferation of roots in these microsites and to determine what causes this response.

Channels formed by the refractory remains of mangrove roots accounted for only 1–2% of total soil volume, but the proportion of roots found within channels varied from 9 to 24% of total live mass. Successive generations of roots growing inside increasingly smaller root channels were also found.

When artificial channels constructed of PVC pipe were buried in the peat for 2 years, those filled with nutrient‐rich organic matter had six times more roots than empty or sand‐filled channels, indicating a response to greater nutrient availability rather than to greater space or less impedance to root growth.

Root proliferation inside decaying roots may improve recovery of nutrients released from decomposing tissues before they can be leached or immobilized in this intertidal environment. Greatest root proliferation in channels occurred in interior forest zones characterized by greater soil waterlogging, which suggests that this may be a strategy for nutrient capture that minimizes oxygen losses from the whole root system.

Improved efficiency of nutrient acquisition at the individual plant level has implications for nutrient economy at the ecosystem level and may explain, in part, how mangroves persist and grow in nutrient‐poor environments.