bluuebunny/crossref_metadata_2025 huggingface dataset:
Localisation of phosphorylated MAP kinase during the transition from meiosis I to meiosis II in pig oocytes
Authors: Lee Jibak , Miyano Takashi , Moor Robert M.
Published: July 2002

Mitogen-activated protein kinase (MAPK) has been reported to be involved in oocyte maturation in all animals so far examined. In the present study we investigate the expression and localisation of active phosphorylated MAPKs (p44ERK1/p42ERK2) during maturation of pig oocytes. In immunoblot analysis using anti-p44ERK1 antibody which recognised both active and inactive forms of p44ERK1 and p42ERK2, we confirmed that MAPKs were phosphorylatred around the time of germinal vesicle breakdown (GVBD) and the active phosphorylated MAPKs (pMAKs) were maintained until metaphase II, as has been reported. On immunofluorescent confocal microscopy using anti-pMAPK antibody which recognised only phosphorylated forms of MAPKs, pMAPK was localised at the spindle poles in pig mitotic cells. On the other hand, in pig oocytes, no signal was detected during GV stage. After GVBD, the area around condensed chromosomes was preferentially stained at metaphase I although whole cytoplasm was faintly stained. At early anaphase I, the polar regions of the meiotic spindle were prominently stained. However, during the progression of anaphase I and telophase I pMAPK was detected at the mid-zone of the elongated spindle, gradually becoming concentrated at the centre. Finally, at the time of emission of the first polar body, pMAPK was detected as a ring-like structure between the condensed chromosomes and the first polar body, and the staining was maintained even after the metaphase II spindle was formed. The inhibition of MAPK activity with the MAPK kinase inhibitor U0126 during the meiosis I/meiosis II transition suppressed chromosome separation, first polar body emission and formation of the metaphase II spindle. From these results, we propose that the spindle-associated pMAPKs play an important role in the events occurring during the meiosis I/meiosis II transition, such as chromosome separation, spindle elongation and cleavage furrow formation in pig oocytes.
Sperm binding and penetration of the zona pellucida in vitro but not sperm–egg fusion in an Australian marsupial, the brushtail possum (Trichosurus vulpecula)
Authors: Mate K.E. , Sidhu K.S. , Molinia F.C. , Glazier A.M. , Rodger J.C.
Published: July 2002

Sperm capacitation and in vitro fertilisation (IVF) have been achieved in most eutherian mammals and American marsupials under relatively simple culture conditions. In contrast sperm capacitation in Australian marsupials has not been achieved in vitro and attempts at IVF have previously been characterised by a complete lack of sperm–zona pellucida (ZP) binding. Recently, co-culture of sperm with oviduct epithelial cell monolayers or with oviductal explant conditioned media has been shown to prolong the viability and motility of brushtail possum spermatozoa, as well as to induce capacitation-associated changes such as transformation of sperm to the T-shape orientation. In this study we report that these in vitro produced T-shaped sperm, and in vivo derived T-shaped sperm flushed from the oviduct of artificially inseminated possums as a control, are able to bind to and penetrate the ZP of approximately 25% of eggs recovered from PMSG/LH-superovulated possums in vitro. Development of ZP receptivity and penetrability towards sperm was also identified as a major factor affecting the outcome of IVF. Neither in vivo nor in vitro derived T-shaped sperm were able to bind to or penetrate the ZP if eggs were obtained from animals that were treated with pLH less than 76 h after PMSG. Thus this study provides preliminary evidence for the necessity of sperm–oviduct epithelial cell interactions for capacitation in Australian species and lends further support to the suggestion that the T-shape head orientation is indicative of sperm capacitation. Despite the occurrence of sperm–ZP binding and penetration, sperm–egg membrane fusion and egg activation were not observed. Although the factor(s) responsible for the lack of sperm–egg membrane fusion in the possum have not been identified it is possible that egg capacity for membrane fusion develops independently of zona receptivity and is defective in these eggs, or alternatively that membrane fusion requires strictly defined ionic conditions which are not provided by the IVF media used in this study.
Fertilisability of ovine, bovine or minke whale (Balaenoptera acutorostrata) spermatozoa intracytoplasmically injected into bovine oocytes
Authors: Wei Hong , Fukui Yutaka
Published: July 2002

This study was conducted to investigate the possibility of using bovine oocytes for a heterologous fertility test by intracytoplasmic sperm injection (ICSI) and to compare the pronuclear formation of ram, bull and minke whale spermatozoa after injection into bovine oocytes. Bovine oocytes were cultured in vitro for 24 h and those with a polar body were selected for ICSI. Frozen-thawed semen from the three species were treated with 5 mM dithiothreitol for 1 h and spermatozoa were killed by storing them in a -20 °C refrigerator before use. ICSI was performed using a Piezo system. Three experiments were designed. In experiment 1, a higher (p < 0.05) male pronuclear formation rate was found in the oocytes injected with ram (52.6%) or bull (53.4%) spermatozoa than with minke whale spermatozoa (39.1%). In experiment 2, sperm head decondensation was detected at 2 h after ICSI in the oocytes injected with a spermatozoon of each species. Male pronuclei were first observed at 4 h in the oocytes injected with ram or bull spermatozoa and at 6 h in oocytes injected with minke whale spermatozoa. The mean diameters of male pronuclei derived from both whale and bull spermatozoa were larger than those from ram spermatozoa (30.4 μm and 28.3 μm vs 22.4 μm, p < 0.005). The mean diameter of female pronuclei in the oocytes injected with whale spermatozoa was also larger than with ram spermatozoa (29.3 μm vs 24.7 μm, p < 0.05). The development of male and female pronuclei was synchronous. In experiment 3, ethanol-activated oocytes injected with a spermatozoon from any of the three species achieved significantly higher (p < 0.05-0.001) cleavage rates than control oocytes. Blastocyst formation was only observed when bull spermatozoa were used. The results of this study indicate that dead foreign spermatozoa can participate in fertilisation activities in bovine oocytes after ICSI.
Air and road surface temperature variations during weather change
Authors: Postgård U , Lindqvist S
Published: July 2002

In this study, thermal mapping along the road and recordings from field stations in the Swedish Road Weather Information System (RWIS) are analysed. The purpose is to explore how long it takes for air and road surface temperatures to adjust from clear to overcast conditions after a warm front arrival, and to see if there are any variations in reaction time between the two temperatures. The study also focuses on whether different sitings affect the adjustment time when the warm front arrives and if the time of arrival is important for the resulting temperature variations. The results show that weather preceding the front, temperature difference when the front arrives, arrival time of the front, wind speed and the occurrence of precipitation all have an effect on the adjustment time. Air temperature reacts faster to a weather change than the road surface temperature. During the evening the road surface temperature can still be affected by previous shading for up to four hours after a front arrival. When the total cloud cover increases above 4 oktas before noon there are no or small lags in reaction time between air and road surface temperature, since there is insufficient time to establish shading effects. Copyright © 2001 Royal Meteorological Society
Prediction of hoar‐frost by use of a Road Weather information System
Authors: Karlsson Maria
Published: July 2002

This study measured the amount of hoar‐frost on the road surface with the help of copper plates. The amounts were compared to data collected at a station in the Road Weather information System (RWiS) together with additional measurements of the temperature and humidity profile above the road surface. The data were used to specify the weather parameters when hoar‐frost forms on the road surface and relate the measured amount of hoar‐frost and resulting friction to measured weather parameters. The aim was also to see whether there was an increased possibility of predicting the amount of hoar‐frost with additional measurements of the temperature and humidity profiles. The results of this study confirm that the amount of hoar‐frost deposited on the road surface increases with increasing average wind speed, increasing maximum difference between dew point (Td) and road surface temperature (Tr) and increased duration when Tr<Td, measured at the RWiS station. However, the difference between Td and Tr failed to indicate hoar‐frost on several occasions due to the location and poor accuracy of the measuring equipment. Measurements of temperature and humidity in a profile up to 2.5 m above the road surface showed that the air layer up to 0.4 m reflected conditions at the road surface whereas the surrounding areas influenced the measurements at higher levels. A forward multiple regression performed on the data showed that measurements of temperature and humidity at lower levels improved predictions of deposition of hoar‐frost on the road surface. Measurements of friction on the road surface when hoar‐frost was present showed that friction decreased with increasing difference between Td and Tr and it also increased with the duration of Tr<Td. There was no correlation between the amount of hoar‐frost and friction at the road surface. The friction is probably influenced not only by the amount of hoar‐frost but also by the structure of the hoar‐frost, which has not been determined in this study. Copyright © 2001 Royal Meteorological Society
Synoptic analysis of an extreme heat wave over Argentina in March 1980
Authors: Campetella Claudia , Rusticucci Matilde
Published: July 2002

During the last two weeks of March 1980 an intense heat wave developed in central and northern Argentina, with its greatest intensity over the city of Buenos Aires. The sequence of temperature anomalies showed the importance of this heat wave. During the period of maximum intensity the daily minimum temperatures were over 27° and the apparent temperatures over 30°. It was the most intense heat wave of the last two decades and the only one that has occurred in March. The intensity of the event is shown by the high values of several biometeorological indices that indicate dangerous levels of thermal stress on people. The persistence of an anticyclonic circulation at all tropospheric levels and the extension of the barotropic zone to the south led to the maintenance of high values of temperature and humidity. The advective effect added to the intense solar radiation was the most important cause of the high temperatures. Copyright © 2007 John Wiley & Sons, Ltd.
RATIONALE FOR A NEW MEASURE OF COMPETENCE IN THERAPY
Authors: Milne Derek , Claydon Tony , Blackburn Ivy-Marie , James Ian , Sheikh Alia
Published: July 2002

Therapy process research has made surprisingly little headway during the past 25 years, which has been
attributed to a range of methodological and conceptual problems. As a result, appeals have been made for fresh
approaches to psychotherapy process research. Here we provide an experiential learning model as one promising way
to crystallize conceptualizations of competence in therapy and to advance research. The model provides an integrative
account of the moderators and mediators believed to explain the way that therapy achieves its outcomes. Its breadth
takes it beyond the scope of its closest rival, the assimilation model. By way of illustration, the theoretical components
of the experiential metaphor were operationalized and mapped onto an existing competence scale, the Cognitive
Therapy Scale.
COGNITIVE-BEHAVIORAL SELF-INTERVENTION VERSUS SELF-MONITORING OF ANGER: EFFECTS ON ANGER FREQUENCY, DURATION, AND INTENSITY
Authors: Fernandez Ephrem , Beck Richard
Published: July 2002

In the self-regulation of anger, it is unclear whether the effects are due to self-monitoring or actual self-intervention. This is especially important in the light of the reactivity often attributed to self-monitoring. The present
study compared both procedures within a reversal design involving college students with moderate levels of anger.
Intervention consisted of behavioral contracting and relaxation as well as cognitive strategies of reappraisal and
imagery. Measures included anger frequency, duration, and intensity, all of which were obtained in naturalistic settings.
Results revealed significant and dramatic reductions in anger frequency and duration during self-intervention but no
such improvements with self-monitoring apart from a slight reduction in anger intensity. The return to baseline self-monitoring was associated with no relapse in anger but a maintenance of treatment gains. Secondary findings of interest
were a correlation between anger intensity and anger duration and a predominance of anger directed at persons rather
than situations. In general, the outcome of this study adds strength to the inference that cognitive-behavioral self-interventions are effective for anger and that this effectiveness goes beyond mere reactivity. The effects are most
pronounced for anger frequency and duration and this offers clues for future focal points in the practice of anger
management.
Conceptual apraxia in probable Alzheimer's disease
as demonstrated by the Florida Action Recall Test
Authors: SCHWARTZ RONALD L. , ADAIR JOHN C. , RAYMER ANASTASIA M. , WILLIAMSON DAVID J.G. , CROSSON BRUCE , ROTHI LESLIE J.G. , NADEAU STEPHEN E. , HEILMAN KENNETH M.
Published: July 2002

Patients with probable Alzheimer's disease
(AD) often have difficulties associated with semantic knowledge.
Therefore, conceptual apraxia, a defect of action semantics
and mechanical knowledge, may be an early sign of this
disease. The Florida Action Recall Test (FLART), developed
to assess conceptual apraxia, consists of 45 line drawings
of objects or scenes. The subject must imagine the proper
tool to apply to each pictured object or scene and then
pantomime its use. Twelve participants with Alzheimer's
disease (NINCDS–ADRDA criteria) and 21 age- and education-matched
controls were tested. Nine Alzheimer's disease participants
scored below a 2-standarddeviation cutoff on conceptual
accuracy, and the three who scored above the cutoff were
beyond a 2-standard-deviation cutoff on completion time.
The FLART appears to be a sensitive measure of conceptual
apraxia in the early stages of Alzheimer's disease.
(JINS, 2000, 6, 265–270.)
Avoiding health risks from drinking water in Moscow: An empirical analysis
Authors: LARSON BRUCE A. , GNEDENKO EKATERINA D.
Published: July 2002

Casual observation suggests that many households in Moscow boil water, settle water in pans for some periods (e.g., overnight) before consuming, filter water, and buy bottled water. To date, there has been little empirical analysis of such avoidance behavior. Based on a recently completed survey of 615 households in Moscow, this paper investigates the types and amounts of avoidance measures that are used by households in Moscow to adjust drinking water quality. Survey results show that this is clearly the case: over 88 per cent of the sample boil water regularly due to concerns about water quality; 23 per cent filter water regularly; over 30 per cent settle water regularly; and about 13 per cent buy bottled water regularly. On the other hand, residents are generally content with their cold water supply and quality of delivery. Based on a microeconomic model of household avoidance behavior, logit regression results show how avoidance decisions relate to income, opinions of water quality, and location in the city. It is expected that this analysis from Moscow can also be used as a guide for future studies in other cities in Russia to evaluate opinions of quality, avoidance measures, and citizens' willingness to support public infrastructure projects designed to improve water supply.
Pendulum-based instruments, percussive video, sound art, and the permanence of ephemeral public art
Authors: SENN DAN
Published: July 2002

In this article, and within the context of sound art as a basis for artistic
freedom in experimental music and art, the development of pendulum-based
sculptural instruments and an improvisational form of video called
‘percussive video’ are described. Methods used to produce
time-based public art events are also outlined, and their effectiveness in
directing social change at the local level is discussed.
Optimal expressions in Indian English
Authors: BHATT RAKESH M.
Published: July 2002

This paper presents an account of sociolinguistic–syntactic variation in Indian English. It focuses on the differences between Standard Indian English (SIE) and Indian Vernacular English (IVE), restricting the discussion to the syntax of wh-questions, Focus constructions, null subjects and objects (pro drop), and null expletive subjects. The major claim of this paper is that the differences between the observed patterns of generalization in SIE and IVE are best accounted for in a conceptualization of grammar that is based on a general notion of priority of syntactic constraints on well-formedness. This optimality-theoretic (OT) conceptualization (Grimshaw, 1997; Prince & Smolensky, 1993) is able to capture the following generalizations: (a) IVE is just as systematic and logical as SIE; (b) the grammars of IVE and SIE are bound by the same set of grammatical constraints; and (c) the differences between the two varieties is a function of how each grammar prioritizes these constraints. This paper thus also presents a strong argument for using OT as a framework for research on language variation and use.
A major restructuring in the English consonant system: the de-linearization of [h] and the de-consonantization of [w] and [j]
Authors: Anderson John
Published: July 2002

This article deals in outline with two interrelated aspects of the history of English phonology: aspects that are argued here to involve the loss of contrastive linearization for /h/ and of consonantal status for /w/ and /j/. It is suggested that these histories are clarified if proper attention is paid to contrastivity: it is necessary to identify those aspects of phonological representation which are contrastive, both segmentally and sequentially. Contrastive status can change. In this case, it is proposed here that the position of /h/ in a word has come to be noncontrastive, and that [w] and [j] are no longer contrastively consonantal, but sequential variants of their full-vowel congeners. The characterization of these restructurings involves the recognition of changing patterns of contrastivity. And, crucially, contrastivity involves not just paradigmatic distinctions between segments but also syntagmatic relations between them.
Introduction
Authors: Green David W.
Published: July 2002

The papers in this Special Issue focus on the use of neuroimaging techniques to answer questions about the neural representation, processing and control of two languages. Neuropsychological data from bilingual aphasics remain vital if we are to establish the neural basis of language (see Paradis, 1995) but lesion-deficit studies alone cannot tell us how neural activity relates to ongoing language processing. Modern neuroimaging methods provide a means to do so. There are two broad classes of such methods: electrophysiological methods allow us to answer questions about when a particular process occurs whereas haemodynamic methods allow us to answer the complementary question of where in the brain such a process is carried out. Before giving a thumb-nail sketch of the papers in this Special Issue, I briefly discuss each class of method.
“Where is my brush-teeth?” Acquisition of compound nouns in a French–English bilingual child
Authors: Nicoladis Elena
Published: July 2002

This study had two purposes: (i) to see if bilingual children can differentiate their languages with respect to the ability to form compound nouns and (ii) to test the validity of previous explanations of the acquisition of compounds. Compound nouns are right-headed in English and left-headed in French. If the French–English bilingual child in this study could differentiate between the two compounding rules, his compounds should show differential order based on the language of the semantic head. The analysis was based on the child's spontaneous compound productions from 2;9 to 3;3. The results showed that the language choice of the semantic head noun predicted the order of his compounds, suggesting that he had two distinct compounding rules. The pattern of errors made by the child cannot be accounted for by any previous explanation alone. It is suggested that children use various cues to learn compound structure.
Astrobiology – the final frontier in chemical reaction dynamics
Authors: Kaiser Ralf I. , Balucani Nadia
Published: July 2002

Crossed-beam experiments on the reactions of cyano CN(X2Σ+) and ethinyl C2H(X2Σ+) radicals with the unsaturated hydrocarbons acetylene, ethylene, methylacetylene allene and benzene have been carried out under single-collision conditions to investigate synthetic routes to form nitriles, polyynes and substituted allenes in hydrocarbon-rich atmospheres of planets and their moons. All reactions were found to proceed without an entrance barrier, to have exit barriers well below the energy of the reactant molecules and to be strongly exothermic. The predominant identification of the radical versus atomic hydrogen exchange channel makes these reactions compelling candidates for the formation of complex organic chemicals – precursors to biologically important amino acids – in Solar system environments and in the interstellar medium.
A murine tumor progression model for pancreatic cancer recapitulating the genetic alterations of the human disease
Authors: Wagner Martin , Greten Florian R. , Weber Christoph K. , Koschnick Stefan , Mattfeldt Torsten , Deppert Wolfgang , Kern Horst , Adler Guido , Schmid Roland M.
Published: July 2002

This study describes a tumor progression model for ductal pancreatic cancer in mice overexpressing TGF-α. Activation of Ras and Erk causes induction of cyclin D1-Cdk4 without increase of cyclin E or PCNA in ductal lesions. Thus, TGF-α is able to promote progression throughout G1, but not S phase. Crossbreeding with p53 null mice accelerates tumor development in TGF-α transgenic mice dramatically. In tumors developing in these mice, biallelic deletion ofInk4a/Arf or LOH of the Smad4 locus is found suggesting that loci in addition to p53 are involved in antitumor activities. We conclude that these genetic events are critical for pancreatic tumor formation in mice. This model recapitulates pathomorphological features and genetic alterations of the human disease.
The transcriptional repressor CDP (Cutl1) is essential for epithelial cell differentiation of the lung and the hair follicle
Authors: Ellis Tammy , Gambardella Laure , Horcher Markus , Tschanz Stefan , Capol Janine , Bertram Paula , Jochum Wolfram , Barrandon Yann , Busslinger Meinrad
Published: July 2002

The mammalian Cutl1 gene codes for the CCAAT displacement protein (CDP), which has been implicated as a transcriptional repressor in diverse processes such as terminal differentiation, cell cycle progression, and the control of nuclear matrix attachment regions. To investigate the in vivo function of Cutl1, we have replaced the C-terminal Cut repeat 3 and homeodomain exons with an in-framelacZ gene by targeted mutagenesis in the mouse. The CDP–lacZ fusion protein is retained in the cytoplasm and fails to repress gene transcription, indicating that the Cutl1lacZ allele corresponds to a null mutation. Cutl1 mutant mice on inbred genetic backgrounds are born at Mendelian frequency, but die shortly after birth because of retarded differentiation of the lung epithelia, which indicates an essential role of CDP in lung maturation. A less pronounced delay in lung development allows Cutl1 mutant mice on an outbred background to survive beyond birth. These mice are growth-retarded and develop an abnormal pelage because of disrupted hair follicle morphogenesis. The inner root sheath (IRS) is reduced, and the transcription of Sonic hedgehog and IRS-specific genes is deregulated in Cutl1 mutant hair follicles, consistent with the specific expression of Cutl1 in the progenitors and cell lineages of the IRS. These data implicate CDP in cell-lineage specification during hair follicle morphogenesis, which resembles the role of the related Cut protein in specifying cell fates duringDrosophila development.
Fine structure of E. coli RNA polymerase-promoter interactions: α subunit binding to the UP element minor groove
Authors: Ross Wilma , Ernst Alexander , Gourse Richard L.
Published: July 2002

The α subunit of E. coli RNAP plays an important role in the recognition of many promoters by binding to the A+T-rich UP element, a DNA sequence located upstream of the recognition elements for the ς subunit, the −35 and −10 hexamers. We examined DNA–RNAP interactions using high resolution interference and protection footprinting methods and using the minor groove-binding drug distamycin. Our results suggest that α interacts with bases in the DNA minor groove and with the DNA backbone along the minor groove, but that UP element major groove surfaces do not make a significant contribution to α binding. On the basis of these and previous results, we propose a model in which α contacts UP element DNA through amino acid residues located in a pair of helix–hairpin–helix motifs. Furthermore, our experiments extend existing information about recognition of the core promoter by ς70 by identifying functional groups in the major grooves of the −35 and −10 hexamers in which modifications interfere with RNAP binding. These studies greatly improve the resolution of our picture of the promoter–RNAP interaction.
GT Repeats Are Associated with Recombination on Human Chromosome 22
Authors: Majewski Jacek , Ott Jurg
Published: July 2002

The rate of meiotic recombination is not a constant function of physical distance across chromosomes. This variation is manifested by recombination hot spots and cold spots, observed in all organisms ranging from bacteria to humans. It is generally believed that factors such as primary and secondary DNA sequence, as well as chromatin structure and associated proteins, influence the frequency of recombination within a specific region. Several such factors, for example repetitive sequences, gene promoters, or regions with the ability to adopt Z-DNA conformation, have been hypothesized to enhance recombination. However, apart from specific examples, no general trends of association between recombination rates and particular DNA sequence motifs have been reported. In this paper, we analyze the complete sequence data from human chromosome 22 and compare microsatellite repeat distributions with mitotic recombination patterns available from earlier genetic studies. We show significant correlation between long tandem GT repeats, which are known to form Z-DNA and interact with several components of the recombination machinery, and recombination hot spots on human chromosome 22.
Eukaryote-specific Domains in Translation Initiation Factors: Implications for Translation Regulation and Evolution of the Translation System
Authors: Aravind L. , Koonin Eugene V.
Published: July 2002

Computational analysis of sequences of proteins involved in translation initiation in eukaryotes reveals a number of specific domains that are not represented in bacteria or archaea. Most of these eukaryote-specific domains are known or predicted to possess an α-helical structure, which suggests that such domains are easier to invent in the course of evolution than are domains of other structural classes. A previously undetected, conserved region predicted to form an α-helical domain is delineated in the initiation factor eIF4G, inNonsense-mediated mRNA decay2 protein (NMD2/UPF2), in the nuclear cap-binding CBP80, and in other, poorly characterized proteins, which is named the NIC (NMD2, eIF4G, CBP80) domain. Biochemical and mutagenesis data on NIC-containing proteins indicate that this predicted domain is one of the central adapters in the regulation of mRNA processing, translation, and degradation. It is demonstrated that, in the course of eukaryotic evolution, initiation factor eIF4G, of which NIC is the core, conserved portion, has accreted several additional, distinct predicted domains such as MI (MA-3 and eIF4G ) and W2, which probably was accompanied by acquisition of new regulatory interactions.
Primacy Versus Recency in a Quantitative Model: Activity Is the Critical Distinction
Authors: Greene Anthony J. , Prepscius Colin , Levy William B.
Published: July 2002

Behavioral and neurobiological evidence shows that primacy and recency are subserved by memory systems for intermediate- and short-term memory, respectively. A widely accepted explanation of recency is that in short-term memory, new learning overwrites old learning. Primacy is not as well understood, but many hypotheses contend that initial items are better encoded into long-term memory because they have had more opportunity to be rehearsed. A simple, biologically motivated neural network model supports an alternative hypothesis of the distinct processing requirements for primacy and recency given single-trial learning without rehearsal. Simulations of the model exhibit either primacy or recency, but not both simultaneously. The incompatibility of primacy and recency clarifies possible reasons for two neurologically distinct systems. Inhibition, and its control of activity, determines those list items that are acquired and retained. Activity levels that are too low do not provide sufficient connections for learning to occur, while higher activity diminishes capacity. High recurrent inhibition, and progressively diminishing activity, allows acquisition and retention of early items, while later items are never acquired. Conversely, low recurrent inhibition, and the resulting high activity, allows continuous acquisition such that acquisition of later items eventually interferes with the retention of early items.
Inhibition of Nitric Oxide Synthesis and Gene Knockout of Neuronal Nitric Oxide Synthase Impaired Adaptation of Mouse Optokinetic Response Eye Movements
Authors: Katoh Akira , Kitazawa Hiromasa , Itohara Shigeyoshi , Nagao Soichi
Published: July 2002

Nitric oxide (NO) plays a key role in synaptic transmission efficiency in the central nervous system. To gain an insight on the role of NO in cerebellar functions, we, here, measured the dynamics of the horizontal optokinetic response (HOKR) and vestibulo-ocular reflex (HVOR), and the adaptation of HOKR in mice locally injected withNG-monomethyl-l-arginine (L-NMMA) that inhibits NO synthesis and in mice devoid of neuronal nitric oxide synthase (nNOS). Local application of L-NMMA into the cerebellar flocculi induced no change in the dynamics of the HOKR but markedly depressed the adaptation of the HOKR induced by 1 hr of sustained screen oscillation. A slight difference was seen in the HOKR but not in the HVOR dynamics between nNOS−/− mutant and wild-type mice. One hour of sustained screen oscillation induced adaptation of the HOKR gains in wild-type mice but not in mutants. These observations suggest that NO is essential for the adaptation of the HOKR and that nNOS is the major enzyme for NO synthesis in the process.
Hormone and Seed-Specific Regulation of Pea Fruit Growth
Authors: Ozga Jocelyn A. , van Huizen Rika , Reinecke Dennis M.
Published: July 2002

Growth of young pea (Pisum sativum) fruit (pericarp) requires developing seeds or, in the absence of seeds, treatment with gibberellin (GA) or auxin (4-chloroindole-3-acetic acid). This study examined the role of seeds and hormones in the regulation of cell division and elongation in early pea fruit development. Profiling histone H2A and γ-tonoplast intrinsic protein (TIP) gene expression during early fruit development identified the relative contributions of cell division and elongation to fruit growth, whereas histological studies identified specific zones of cell division and elongation in exocarp, mesocarp, and endocarp tissues. Molecular and histological studies showed that maximal cell division was from −2 to 2 d after anthesis (DAA) and elongation from 2 to 5 DAA in pea pericarp. Maximal increase in pericarp γ-TIP message level preceded the maximal rate of fruit growth and, in general, γ-TIP mRNA level was useful as a qualitative marker for expanding tissue, but not as a quantitative marker for cell expansion. Seed removal resulted in rapid decreases in pericarp growth and in γ-TIP and histone H2A message levels. In general, GA and 4-chloroindole-3-acetic acid maintained these processes in deseeded pericarp similarly to pericarps with seeds, and both hormones were required to obtain mesocarp cell sizes equivalent to intact fruit. However, GA treatment to deseeded pericarps resulted in elevated levels of γ-TIP mRNA (6 and 7 DAA) when pericarp growth and cell enlargement were minimal. Our data support the theory that cell division and elongation are developmentally regulated during early pea fruit growth and are maintained by the hormonal interaction of GA and auxin.
Two Wound-Inducible Soybean Cysteine Proteinase Inhibitors Have Greater Insect Digestive Proteinase Inhibitory Activities than a Constitutive Homolog
Authors: Zhao Y. , Botella M. A. , Subramanian L. , Niu X. , Nielsen S. S. , Bressan R. A. , Hasegawa P. M.
Published: July 2002

Diverse functions for three soybean (Glycine max L. Merr.) cysteine proteinase inhibitors (CysPIs) are inferred from unique characteristics of differential regulation of gene expression and inhibitory activities against specific Cys proteinases. Based on northern blot analyses, we found that the expression in leaves of one soybean CysPI gene (L1) was constitutive and the other two (N2 and R1) were induced by wounding or methyl jasmonate treatment. Induction of N2 and R1 transcript levels in leaves occurred coincidentally with increased papain inhibitory activity. Analyses of kinetic data from bacterial recombinant CysPI proteins indicated that soybean CysPIs are noncompetitive inhibitors of papain. The inhibition constants against papain of the CysPIs encoded by the wound and methyl jasmonate-inducible genes (57 and 21 nM for N2 and R1, respectively) were 500 to 1000 times lower than the inhibition constant of L1 (19,000 nM). N2 and R1 had substantially greater inhibitory activities than L1 against gut cysteine proteinases of the third-instar larvae of western corn rootworm and Colorado potato beetle. Cysteine proteinases were the predominant digestive proteolytic enzymes in the guts of these insects at this developmental stage. N2 and R1 were more inhibitory than the epoxide trans-epoxysuccinyl-L-leucylamide-(4-guanidino)butane (E-64) against western corn rootworm gut proteinases (50% inhibition concentration = 50, 200, and 7000 nM for N2, R1, and E-64, respectively). However, N2 and R1 were less effective than E-64 against the gut proteinases of Colorado potato beetle. These results indicate that the wound-inducible soybean CysPIs, N2 and R1, function in host plant defense against insect predation, and that substantial variation in CysPI activity against insect digestive proteinases exists among plant CysPI proteins.
Genetic Engineering of Plant Secondary Metabolism (Accumulation of 4-Hydroxybenzoate Glucosides as a Result of the Expression of the Bacterial ubiC Gene in Tobacco)
Authors: Siebert M. , Sommer S. , Li Sm. , Wang Zx. , Severin K. , Heide L.
Published: July 2002

The ubiC gene of Escherichia coli encodes chorismate pyruvatelyase, an enzyme that converts chorismate into 4-hydroxybenzoate (4HB) and is not normally present in plants. The ubiC gene was expressed in Nicotiana tabacum L. plants under control of a constitutive plant promoter. The gene product was targeted into the plastid by fusing it to the sequence for the chloroplast transit peptide of the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Transgenic plants showed high chorismate pyruvate-lyase activity and accumulated 4HB as β-glucosides, with the glucose attached to either the hydroxy or the carboxyl function of 4HB. The total content of 4HB glucosides was approximately 0.52% of dry weight, which exceeded the content of untransformed plants by at least a factor of 1000. Feeding experiments with [1,7-13C2]shikimic acid unequivocally proved that the 4HB that was formed in the transgenic plants was not derived from the conventional phenylpropanoid pathway but from the newly introduced chorismate pyruvate-lyase reaction.
Cytokinin, Acting through Ethylene, Restores Gravitropism to Arabidopsis Seedlings Grown under Red Light
Authors: Golan A. , Tepper M. , Soudry E. , Horwitz B. A. , Gepstein S.
Published: July 2002

Cytokinin replaces light in several aspects of the photomorphogenesis of dicot seedlings. Arabidopsis thaliana seedlings grown under red light have been shown to become disoriented, losing the negative hypocotyl gravitropism that has been observed in seedlings grown in darkness or white light. We report here that cytokinin at micromolar concentrations restores gravitropism to seedlings grown under red light. Cytokinin cancels the effect of red light on the gravity-sensing system and at the same time replaces light in the inhibition of hypocotyl elongation. Furthermore, application of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid acts similarly to cytokinin. Cytokinin cannot restore gravitropism under red light to an ethylene-insensitive mutant that is defective at the EIN2 locus. Stimulation of ethylene production, therefore, can explain the action of cytokinin in restoring negative gravitropism to the hypocotyls of Arabidopsis seedlings grown under continuous red light.
Isolation and Characterization of Glutamine Synthetase Genes in Chlamydomonas reinhardtii
Authors: Chen Q. , Silflow C. D.
Published: July 2002

To elucidate the role of glutamine synthetase (GS) in nitrogen assimilation in the green alga Chlamydomonas reinhardtii we used maize GS1 (the cytosolic form) and GS2 (the chloroplastic form) cDNAs as hybridization probes to isolate C. reinhardtii cDNA clones. The amino acid sequences derived from the C. reinhardtii clones have extensive homology with GS enzymes from higher plants. A putative amino-terminal transit peptide encoded by the GS2 cDNA suggests that the protein localizes to the chloroplast. Genomic DNA blot analysis indicated that GS1 is encoded by a single gene, whereas two genomic fragments hybridized to the GS2 cDNA probe. All GS2 cDNA clones corresponded to only one of the two GS2 genomic sequences. We provide evidence that ammonium, nitrate, and light regulate GS transcript accumulation in green algae. Our results indicate that the level of GS1 transcripts is repressed by ammonium but induced by nitrate. The level of GS2 transcripts is not affected by ammonium or nitrate. Expression of both GS1 and GS2 genes is regulated by light, but perhaps through different mechanisms. Unlike in higher plants, no decreased level of GS2 transcripts was detected when cells were grown under conditions that repress photorespiration. Analysis of GS transcript levels in mutants with defects in the nitrate assimilation pathway show that nitrate assimilation and ammonium assimilation are regulated independently.
The Ferredoxin-Binding Site of Ferredoxin:Nitrite Oxidoreductase (Differential Chemical Modification of the Free Enzyme and Its Complex with Ferredoxin)
Authors: Dose M. M. , Hirasawa M. , Kleis-SanFrancisco S. , Lew E. L. , Knaff D. B.
Published: July 2002

Spinach (Spinacea oleracea) leaf ferredoxin (Fd)-dependent nitrite reductase was treated with either the arginine-modifying reagent phenyl-glyoxal or the lysine-modifying reagent pyridoxal-5[prime]-phosphate under conditions where only the Fd-binding affinity of the enzyme was affected and where complex formation between Fd and the enzyme prevented the inhibition by either reagent. Modification with [14C]phenylglyoxal allowed the identification of two nitrite reductase arginines, R375 and R556, that are protected by Fd against labeling. Modification of nitrite reductase with pyridoxal-5[prime]-phosphate, followed by reduction with NaBH4, allowed the identification of a lysine, K436, that is protected by Fd against labeling. Positive charges are present at these positions in all of the Fd-dependent nitrite reductases for which sequences are available, suggesting that these amino acids are directly involved in electrostatic binding of Fd to the enzyme.
Differential Expression of Chitinases in Vitis vinifera L. Responding to Systemic Acquired Resistance Activators or Fungal Challenge
Authors: Busam G. , Kassemeyer H. H. , Matern U.
Published: July 2002

The concept of systemic acquired resistance (SAR) enables a novel approach to crop protection, and particular pathogenesis-related proteins, i.e. an acidic chitinase, have been classified as markers of the SAR response. Basic class I (VCHIT1b) and a class III (VCH3) chitinase cDNAs were cloned from cultured Vitis vinifera L. cv Pinot Noir cells and used to probe the induction response of grapevine cells to salicylic acid or yeast elicitor. Furthermore, the cells were treated with the commercial SAR activators 2,6-dichloroiso-nicotinic acid or benzo(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester. Elicitor or salicylic acid induced both VCHIT1b and VCH3 transcript abundances, whereas 2,6-dichloroiso-nicotinic acid or benzo(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester enhanced exclusively the expression of VCH3. To assess the systemic sensation of chitinase expression, single leaves of Vitis vinifera L. cv Pinot Noir or Vitis rupestris plants were inoculated with Plasmopara viticola spore suspensions, and the VCH3 and VCHIT1b mRNA amounts in the infected versus the adjacent, healthy leaf were monitored. Two VCH3 mRNA maxima were observed 2 and 6 d postinoculation in the infected, susceptible V. vinifera tissue, whereas in the healthy leaf the transcript increased from low levels d 2 postinoculation to prominent levels d 6 to 8 postinoculation. The level of VCH3 mRNA increased also over 4 d in the inoculated, resistant V. rupestris tissue. However, necrotic spots rapidly limited the infection, and the VCH3 transcript was undetectable in the upper-stage, healthy leaf. The expression of VCHIT1b remained negligible under either experimental condition. Overall, the results suggest that the selective expression of VCH3 might be a reliable indicator of the SAR response in V. vinifera L.
Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Activase Deficiency Delays Senescence of Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase but Progressively Impairs Its Catalysis during Tobacco Leaf Development
Authors: He Z. , von Caemmerer S. , Hudson G. S. , Price G. D. , Badger M. R. , Andrews T. J.
Published: July 2002

Transgenic tobacco (Nicotiana tabacum L. cv W38) plants with an antisense gene directed against the mRNA of ribulose-1,5-biphosphate carboxylase/oxygenase (Rubisco) activase grew more slowly than wild-type plants in a CO2-enriched atmosphere, but eventually attained the same height and number of leaves. Compared with the wild type, the anti-activase plants had reduced CO2 assimilation rates, normal contents of chlorophyll and soluble leaf protein, and much higher Rubisco contents, particularly in older leaves. Activase deficiency greatly delayed the usual developmental decline in Rubisco content seen in wild-type leaves. This effect was much less obvious in another transgenic tobacco with an antisense gene directed against chloroplast-located glyceraldehyde-3-phosphate dehydrogenase, which also had reduced photosynthetic rates and delayed development. Although Rubisco carbamylation was reduced in the anti-activase plants, the reduction was not sufficient to explain the reduced photosynthetic rate of older anti-activase leaves. Instead, up to a 10-fold reduction in the catalytic turnover rate of carbamylated Rubisco in vivo appeared to be the main cause. Slower catalytic turnover by carbamylated Rubisco was particularly obvious in high-CO2-grown leaves but was also detectable in air-grown leaves. Rubisco activity measured immediately after rapid extraction of anti-activase leaves was not much less than that predicted from its degree of carbamylation, ruling out slow release of an inhibitor from carbamylated sites as a major cause of the phenomenon. Nor could substrate scarcity or product inhibition account for the impairment. We conclude that activase must have a role in vivo, direct or indirect, in promoting the activity of carbamylated Rubisco in addition to its role in promoting carbamylation.
The PsaD Subunit of Photosystem I (Mutations in the Basic Domain Reduce the Level of PsaD in the Membranes)
Authors: Chitnis V. P. , Ke A. , Chitnis P. R.
Published: July 2002

The PsaD subunit of photosystem I (PSI) is a peripheral protein that provides a docking site for ferredoxin and interacts with the PsaB, PsaC, and PsaL subunits of PSI. We used site-directed mutagenesis to determine the function of a basic region in PsaD of the cyanobacterium Synechocystis sp. PCC 6803. We generated five mutant strains in which one or more charged residues were altered. Western blotting showed that replacement of lysine (Lys)-74 with glutamine or glutamic acid led to a substantial decrease in the level of PsaD in the membranes. The mutant PSI complexes showed reduced NADP+ photoreduction activity mediated by ferredoxin; the decrease in activity correlated with the reduced level of PsaD. Using protein synthesis inhibitors we showed that the degradation rates of the mutant and wild-type PsaD were similar, indicating a defect in the assembly of the mutant protein. Treatment of the mutant PSI complexes with a different concentration of Nal showed that the mutations decreased affinity between PsaD and the transmembrane components of PSI. With glutaraldehyde, the mutant and wild-type PsaD proteins could be cross-linked with PsaC, but the PsaD-PsaL cross-linked product was reduced drastically when arginine-72, Lys-74, and Lys-76 were mutated simultaneously. These studies demonstrate that the basic residues in the central region of PsaD, especially Lys-74, are crucial in the assembly of PsaD into the PSI complex.
Uptake of HCO3− and CO2 in Cells and Chloroplasts from the Microalgae Chlamydomonas reinhardtii and Dunaliella tertiolecta1
Authors: Amoroso Gabi , Sültemeyer Dieter , Thyssen Christoph , Fock Heinrich P.
Published: July 2002

Mass-spectrometric disequilibrium analysis was applied to investigate CO2 uptake and HCO3− transport in cells and chloroplasts of the microalgae Dunaliella tertiolecta andChlamydomonas reinhardtii, which were grown in air enriched with 5% (v/v) CO2 (high-Ci cells) or in ambient air (low-Ci cells). High- and low-Ci cells of both species had the capacity to transport CO2 and HCO3−, with maximum rates being largely unaffected by the growth conditions. In high- and low-Ci cells ofD. tertiolecta, HCO3− was the dominant inorganic C species taken up, whereas HCO3− and CO2 were used at similar rates by C. reinhardtii. The apparent affinities of HCO3− transport and CO2 uptake increased 3- to 9-fold in both species upon acclimation to air. Photosynthetically active chloroplasts isolated from both species were able to transport CO2 and HCO3−. For chloroplasts from C. reinhardtii, the concentrations of HCO3− and CO2 required for half-maximal activity declined from 446 to 33 μm and 6.8 to 0.6 μm, respectively, after acclimation of the parent cells to air; the corresponding values for chloroplasts from D. tertiolecta decreased from 203 to 58 μm and 5.8 to 0.5 μm, respectively. These results indicate the presence of inducible high-affinity HCO3− and CO2 transporters at the chloroplast envelope membrane.