bluuebunny/crossref_metadata_2025 huggingface dataset:
Rapid calcium-current kinetics in synaptic terminals of
goldfish retinal bipolar neurons
Authors: MENNERICK STEVEN , MATTHEWS GARY
Published: July 2002

Onset and offset of calcium current in response
to short, depolarizing voltage pulses were studied in giant
synaptic terminals of goldfish bipolar neurons. Tail-current
decays were extremely rapid (80 μs at −70 mV),
and exponentially slowed at more positive repolarization
potentials. The amplitude of tail current following voltage
pulses varied with the pulse amplitude, according to a
Bolzmann distribution with a V1/2
of −27.6 mV and a slope factor of 13.8 mV. When interpreted
with a Hodgkin-Huxley model, calcium-current onset was best
described by m2 kinetics and a time constant
(τm) of 0.62 ms at −10 mV. The
kinetics of calcium-current onset and offset are at least
two- to threefold faster than those described in other native
cells. Combined with other recent data, the results suggest
that the rapid kinetics may participate in a fast signaling
mode in the goldfish ON-center pathway.
A comparison of the inhibitory actions of glycine and GABA
on acetylcholine release from the rabbit retina
Authors: LINN DAVID M.
Published: July 2002

The inhibition of [3H]acetylcholine
(ACh) release from cholinergic amacrine cells by glycine
and GABA was studied using an in vivo eyecup preparation
in anesthetized rabbits. Glycine (1 mM) had no effect on
basal ACh release, but completely blocked the light-evoked
release of ACh. Glycine also blocked the strong potentiating
effects of picrotoxin (20 μM) normally observed on
basal and light-evoked release. Strychnine (20 μM)
increased basal release, albeit less than picrotoxin, but
partially inhibited and altered the shape of light-evoked
responses. Co-perfusion of picrotoxin and strychnine after
strychnine application resulted in a larger additional
basal increase. However, light-evoked responses were not
restored to a control shape and magnitude, or to potentiated
levels as with picrotoxin alone, but remained altered and
partially inhibited. These results support the concept
of a sustained GABA-mediated inhibition of the cholinergic
pathway in the intact retina. In contrast, glycine-mediated
inhibition of the cholinergic pathway differs, with the
present results indicating a significantly smaller sustained
inhibition of basal release and a temporal inhibition of
light-evoked release. The lack of effect of any of these
compounds on kainate-evoked responses indicates that these
effects are predominately indirect, possibly on the presynaptic
bipolar cell.
A dissection of the electroretinogram from the isolated
rat retina with microelectrodes and drugs
Authors: GREEN DANIEL G. , KAPOUSTA-BRUNEAU NATALIA V.
Published: July 2002

The origins of the a- and b-wave
of the ERG were studied using simultaneous recordings made
across the receptor layer and the full thickness of a piece
of isolated albino rat retina. An inwardly directed current
flowing across the rod outer segments was eliminated from
the recording when postsynaptic activity was blocked with
cobalt or when current source density measurements were
made along the length of the outer segments. Rod photovoltages
were inferred by removing extraneous field potentials from
the recordings made across the photoreceptor layer. The
spatial properties of the photovoltage indicates the responses
came from an area about 100 μm in diameter. The glutamate
analog, APB, which blocks depolarizing bipolar cells, eliminated
the b-wave but left the a-wave unaffected.
The ERG component due to depolarizing bipolar cells was
inferred by subtracting recordings obtained before and
after APB. After treatment with APB a slow component remained.
This component was completely blocked by barium (200 μM),
which blocks potassium channels on Müller cells. Barium
had virtually no effect on low-intensity photovoltages
but did affect the amplitude and shape of the saturated
responses. Barium increased the amplitude of the component
of the ERG which underlies the b-wave. It was
concluded that the depolarizing bipolar cells directly
generate the b-wave of the ERG.
Projections from V1 to lateral suprasylvian cortex: An efferent
pathway in the cat's visual cortex that originates
preferentially from CO blob columns
Authors: BOYD JAMIE D. , MATSUBARA JOANNE A.
Published: July 2002

The patchy pattern of retrograde labeling produced
by injections of anatomical tracers into the lateral suprasylvian
(LS) visual area was compared to the cytochrome oxidase
(CO) blobs in cat visual cortex. Following large injections
of anatomical tracers in LS, retrograde labeling formed
an irregular lattice of patches with a spacing of slightly
less than 1 mm in area 17, and slightly greater than 1
mm in area 18. By comparing labeling in alternate serial
sections, patches of LS-projecting cells in both areas
were found to align with CO blobs. The conclusion of alignment
between CO blob columns and patches of LS-projecting cells
was confirmed by a quantitative analysis which showed a
significant correlation between the local density of LS-projecting
cells in reconstructions of charted cells and the intensity
of CO staining in the CO-reacted sections. As for areas
17 and 18, labeling in other afferent areas of LS was also
patchy with a spacing on the order of 1 mm except for area
19 where we found patches of LS-projecting cells with a
larger spacing, roughly 2 mm. No matching fluctuations
in CO density could be discerned in area 19, however. In
conjunction with recent evidence that CO blob columns in
cats receive strong input from Y-cells of the lateral geniculate
nucleus (Boyd & Matsubara, 1996; Shoham, et al., 1996),
these data support the hypothesis (Shipp & Grant, 1991)
that the patches of LS-projecting cells correspond to Y-cell
input columns. As a relationship between the CO architecture
and certain classes of efferent cells has previously been
shown in primates, these findings show new similarities
between CO blobs in different mammalian species.
A stereological analysis of the lateral geniculate nucleus
in adult Macaca nemestrina monkeys
Authors: BLASCO BEATRIZ , AVENDAÑO CARLOS , CAVADA CARMEN
Published: July 2002

The Cavalieri method and the optical fractionator
were employed to estimate the volume and neuron numbers,
respectively, of the dorsal lateral geniculate nucleus
(dLGN) in seven adult male pigtail monkeys (Macaca
nemestrina). Unbiased estimates were selectively obtained
for the parvocellular (P), magnocellular (M), and interlaminar
plus superficial (I + S) layers of the nucleus. The dLGN
had a mean volume of 56.5 mm3, and contained
on average 1.79 million neurons. The P layers contributed
64% of the volume and 83% of the neurons in the dLGN; the
corresponding proportions for the other layers were 13%
and 9% (M), and 23% and 8% (I + S). Interindividual variability
was large for neuron counts, which varied within a two-fold
range, and lower for volume estimates. Since no published
data are available for the pigtail dLGN, the present results
are compared with quantitative studies of the dLGN in other
macaque species, placing special emphasis in the discussion
of the methodologies used.
Spectral sensitivity of macaque monkeys measured with ERG
flicker photometry
Authors: JACOBS GERALD H. , DEEGAN JESS F.
Published: July 2002

The following is a correction for the article “Spectral
sensitivity of macaque monkeys measured with ERG flicker
photometry,” by Gerald H. Jacobs and Jess F. Deegan
II, that appeared in Visual Neuroscience, 1997,
14, 921–928. As a result of a transcription error
the spectral sensitivity values included in Table 1 were
incorrect.
Most calretinin-containing amacrine cells in the rabbit
retina co-localize glycine
Authors: GÁBRIEL ROBERT , VÖLGYI BÉLA , POLLÁK EDIT
Published: July 2002

Calretinin-containing retinal amacrine cells are
heterogeneous with regard to their neurochemical properties.
In the rabbit retina, about 90% of them contain glycine,
as evidenced in the present study by double-label immunocytochemistry.
In a previous report, we showed that a small population
of amacrine cells contains both γ-aminobutyric acid
and calretinin. In this study, we further identified this
cell population by means of known secondary markers. However,
none of the markers we tested (choline acetyltransferase,
serotonin accumulation, NADPH-diaphorase, vasoactive intestinal
polypeptide) co-localized with calretinin. A small population
(1%) of the cells in the ganglion cell layer contains both
calretinin and glycine. Since calretinin-positive cells
in the ganglion cell layer have been identified as ganglion
cells based on soma size and presence of calretinin-positive
axons in the optic nerve fiber layer, this population may
represent a class of ganglion cell which contains glycine.
Our results, together with those of other studies, suggest
that calretinin is not a general marker of any of the well-known
amacrine cell types in the mammalian retina. Rather, calretinin,
just as other calcium-binding proteins, is distributed
in a species-specific manner. At the same time it appears
that, as shown for horizontal cells, one or more of the
major buffer-type calcium-binding proteins of the EF-hand
family is present in most of the retinal amacrine cells.
Unusual coupling patterns of a cone bipolar cell in the
rabbit retina
Authors: MILLS STEPHEN L.
Published: July 2002

In mammals, gap junctions between retinal bipolar
cells are generally small and tracer coupling has not been
previously demonstrated. In this study, Neurobiotin was
injected into the Ba3-type cone bipolar cell, a medium-field
cone bipolar cell that ramifies in sublamina a
of the rabbit retina. Tracer spread to many other Ba3 bipolar
cells, presumably through gap junctions. It also spread
to a smaller field bipolar cell called the Ba1 that ramifies
at the same depth of the inner plexiform layer. Injection
of Neurobiotin into Ba1 bipolar cells did not produce staining
beyond the injected cell. Tracer coupling from the Ba3
was therefore both heterologous, in that different cell
types were stained, and asymmetric. The unusual properties
of this bipolar cell suggest that its function may differ
from that of most cone bipolar cells, which are narrow-field,
do not overlap, and are poorly coupled to one another.
Morphological differentiation of bipolar cells in the ferret retina
Authors: MILLER E.D. , TRAN M.N. , WONG G.-K. , OAKLEY D.M. , WONG R.O.L.
Published: July 2002

Bipolar cells are not only important for visual
processing but input from these cells may underlie the
reorganization of ganglion cell dendrites in the inner
plexiform layer (IPL) during development. Because little
is known about the development of bipolar cells, here we
have used immunocytochemical markers and dye labeling to
identify and follow their differentiation in the neonatal
ferret retina. Putative cone bipolar cells were immunoreacted
for calbindin and recoverin, and rod bipolar cells were
immunostained for protein kinase C (PKC). Our results show
that calbindin-immunoreactive cone bipolar cells appear
at postnatal day 15 (P15), at which time their axonal terminals
are already localized to the inner half of the IPL. By
contrast, recoverin-immunoreactive cells with terminals
in the IPL are present at birth, but many of these cells
may be immature photoreceptors. By the second postnatal
week, recoverin-positive cells resembling cone bipolar
cells were clearly present, and with increasing age, two
distinct strata of immunolabeled processes occupied the
IPL. PKC-containing rod bipolar cells emerged by the fourth
postnatal week and at this age have stratified arbors in
the inner IPL. The early bias of bipolar axonal arbors
in terminating in the inner or outer half of the IPL is
confirmed by dye labeling of cells with somata in the inner
nuclear layer. At P10, several days before ribbon synapses
have been previously observed in the ferret IPL, the axon
terminals of all dye-labeled bipolar cells were clearly
stratified. The results suggest that bipolar cells could
provide spatially localized interactions that are suitable
for guiding dendritic lamination in the inner retina.
Light-induced changes of extracellular ions and volume in
the isolated chick retina–pigment epithelium preparation
Authors: DMITRIEV A.V. , GOVARDOVSKII V.I. , SCHWAHN H.N. , STEINBERG R.H.
Published: July 2002

To better understand the mechanisms of extracellular
space volume regulation and their possible effects on retinal
function, light-induced changes in the concentrations of
the principal extracellular ions (Na+, K+,
Ca2+, and Cl−) were measured
with ion-sensitive microelectrodes in the chick retina–pigment
epithelium–choroid preparation. Changes of extracellular
space volume were assessed by measuring the concentration
of an impermeant marker, tetramethylammonium. In the inner
retina, transient ON/OFF Na+ decrease was about
twice as large as K+ increase, and the charge
difference was compensated by a decrease in Cl−
concentration. The ion changes were accompanied by extracellular
space-volume decreases here. In the subretinal space,
[Na+]o increase was about
twice as large as K+ decrease, yet
[Cl−]o also decreased;
this was accompanied by a sustained extracellular space-volume
increase. The ionic changes in the inner retina are consistent
with a model of extracellular space-volume regulation which
assumes that neuronal depolarization causes net uptake
of NaCl, cell swelling, and extracellular space shrinkage.
However, to prevent the apparent violation of electroneutrality
in the subretinal space, our simple model should be expanded
to include the involvement of unidentified anion(s). Substantial
changes in the subretinal space volume may influence interaction
between the neural retina and pigment epithelium. Among ionic changes,
only the light-induced [K+]o decrease
around the photoreceptors and the [Ca2+]o
increase near the photoreceptor bodies and synaptic terminals
are large enough (−25% and 7.5%, respectively) to
be likely candidates for integrated intercellular signaling.
Emergent properties of neural systems: How focal molecular neurobiological alterations can

affect behavior
Authors: POST ROBERT M. , WEISS SUSAN R. B.
Published: July 2002

Whereas the basic wiring diagram of the mammalian central nervous system (CNS) is

genetically preprogramed, its fine tuning throughout different phases of infancy, childhood, and

adulthood are highly experience dependent. The potential neurobiological mechanisms and

behavioral effects of such experience-dependent neuroplasticity as a function of stage of

development are outlined. A basic thesis of this paper is that mechanisms involved in neuronal

learning and memory, such as long-term potentiation (LTP) and long-term depression (LTD), are

used and reused not only in the sculpting of the CNS in the initial establishment of connections,

but also again in the molding of personality and behavior based on experience. It is postulated

that for higher order processes such as emotional memory, such neuroplasticity is occurring at

increasingly larger numbers of synapses and cell assemblies with increasing mechanistic

complexity and self-organization. Just as overexcitation or deprivation can profoundly affect the

development of the visual system, it is postulated that similar phenomena exist in the neural

substrates of emotional and cognitive development. In addition, a secondary and potentially more

widespread series of ramifications are likely to occur in the higher order and integrative systems,

such as secondary and tertiary cortical association areas and prefrontal cortex that become the

ultimate integrators of emotion and experience, leading to subsequent actions and plans for the

future. This process is, by definition, plastic, and such remodeling is likely to take place not only

at the level of the single synapse, but also at higher levels of network integration of which we

currently have only the barest glimpse. Nonetheless, beginning to discuss the neurobiology of

such self-organizing plastic systems may begin to change our conceptual approaches to

psychopathology and open new avenues of therapeutics for the major psychiatric illnesses that

are critically dependent on such higher order learning and memory mechanisms.
Prenatal stress alters brain biogenic amine levels in primates
Authors: SCHNEIDER MARY L. , CLARKE A. SUSAN , KRAEMER GARY W. , ROUGHTON ELIZABETH C. , LUBACH GABRIELE R. , RIMM–KAUFMAN SARA , SCHMIDT DENNIS , EBERT MICHAEL
Published: July 2002

In this study, we assessed behavioral responses to social separation at 8 months of age and
cerebrospinal fluid (CSF) concentrations of biogenic amines and metabolites at 8 and 18 months
of age in 12 rhesus monkeys derived from either stressed or undisturbed pregnancies. Compared
to controls from undisturbed pregnancies, prenatal stress-derived monkeys had higher
concentrations of 3-methoxy-4-hydroxyphenylglycol (MHPG), and 3,4-dihydroxyphenylacetic
acid in CSF than controls. Norepinephrine and MHPG response to stress were both correlated
between 8 and 18 months of age. There were few group differences in behavior during social
separation; however, several behavioral differences between groups were found when monkeys
were reunited with cage mates. Prenatally stressed monkeys spent more time clinging to their
surrogates and exploring (including eating and drinking), while controls showed more
locomotion and social play with their cage mates. Collectively, our findings suggest that chronic
unpredictable psychological stress during pregnancy has long-lasting effects on noradrenergic
and dopaminergic activity and behavior in the offspring of gestationally stressed primate
mothers.
Measuring developmental changes in
exposure to adversity: A Life Chart and rating scale approach
Authors: GEST SCOTT D. , REED MARIE–GABRIELLE J. , MASTEN ANN S.
Published: July 2002

The reliability and validity of a
comprehensive, integrative strategy for assessing adversity exposure in childhood and
adolescence was examined in a community-based longitudinal study of 205 children. Extensive
data from multiple measures completed by adolescents and their parents regarding chronic and
acute life experiences were aggregated into a Life Chart record of lifetime adversity exposure.
Severity of adversity in three nonoverlapping time spans was rated by trained judges with
excellent interrater reliability. As hypothesized, Family adversity predominated over all other
types of adversity in childhood, remained salient into midadolescence, and showed considerable
stability over time. Adversity related to an individual's own behavior and psychological
functioning rose to equal prominence by late adolescence. Adversity arising from physical
ailments that were independent of the individual's psychological functioning declined over
time, while adversity arising from the community rose; physical and community events were
uncommon for most individuals but substantial for some. Results suggest that coherent
developmental trends in adversity exposure may be identified effectively through judges'
ratings of severity of adversity over multiple-year time spans.
Exceptional syntax
Authors: BENTON NICK , KENNEDY ANDREW
Published: July 2002

From the points of view of programming pragmatics, rewriting and operational semantics,
the syntactic construct used for exception handling in ML-like programming languages, and
in much theoretical work on exceptions, has subtly undesirable features. We propose and
discuss a more well-behaved construct.
Scope, goals and methods in CALL research: questions of coherence and autonomy
Authors: LEVY MIKE
Published: July 2002

This paper considers the problem of coherence and direction in CALL research. Rather than suggesting
a top-down approach to setting goals for research, it argues for a much closer examination and a much
stronger emphasis on existing CALL research work as a platform for directing and informing future CALL work.
Based on a corpus of 47 CALL research articles published in books and journals in 1999, it sets out a
framework for the description and analysis of CALL research as it is represented in the literature. Two major
directions and three important, though less frequent, directions are described in detail, using examples from
the corpus, and the implications for research in the future are considered. Particular emphasis will be
placed on identifying the goals of CALL researchers and on clarifying the unique attributes of research in
this field.
The imminent extinction of the Kites Milvus milvus fasciicauda and Milvus m. migrans on the Cape Verde Islands
Authors: Hille Sabine , Thiollay Jean-marc
Published: July 2002

We investigated the decline and current status of the Black Kite Milvus m. migrans Boddaert 1783 and the endemic and distinct Red Kite (Cape Verde Kite or Cape Verde Red Kite) Milvus milvus fasciicauda Hartert 1914 on the Cape Verde archipelago. In 1996 and 1997, the first census of both kites on the entire archipelago (Hille 1998) revealed that fewer than 10 individuals of each species still existed and that extinction of all kites on Cape Verde was imminent. As expected from this first survey, the tiny island populations of kites further declined and only two individual Red Kites and one Black Kite were found in 1999. Both populations are expected to become extinct very soon. The presumed causes of extinction such as indirect poisoning, lack of food in a changing landscape, and human persecution are discussed.
Precedent Autonomy: Life-Sustaining Intervention and the
Demented Patient
Authors: NEWTON MICHAEL J.
Published: July 2002

How aggressively should we pursue life-sustaining
treatment of the demented patient? This question becomes
increasingly important as our population ages and medical
technology offers ever more life-prolongation. In Life's
Dominion, Ronald Dworkin addresses the issue in the
context of an Alzheimer patient who had previously declared
the desire to avoid life-sustaining intervention. Dworkin
argues for the primacy of what he calls precedent autonomy:
“A competent person's right to autonomy requires
that his past decisions about how he is to be treated if
he becomes demented be respected even if they contradict
the desires he has at a later point.” In 1995, the
Hastings Center Report carried thoughtful
rebuttals by Daniel Callahan and Rebecca Dresser. Much
of Callahan's article is devoted to patients who never
executed an advance directive, but he states unequivocally
that such directives can be overridden, basing much of
his argument on the writings of Sanford Kadish. Together,
Dresser, Callahan, and Kadish define a clear position opposed
to precedent autonomy.
Proxy, Health, and Personal Care Preferences: Implications
for End-of-Life Care
Authors: AIKMAN PETER J. , THIEL ELAINE C. , MARTIN DOUGLAS K. , SINGER PETER A.
Published: July 2002

The Institute of Medicine's report, “Approaching
Death: Improving Care at the End of Life,”
the American Medical Association's “Education
for Physicians on End-of-Life Care” project, the
Open Society Institute's “Project on Death in
America,” and the “Last Acts” initiative
sponsored by the Robert Wood Johnson Foundation have focused
attention on improving the care of dying patients. These
efforts include advance care planning and the use of written
advance directives (ADs). Although previous studies have
provided quantitative descriptions of patient preferences
for life-sustaining treatment, including those documented
in written ADs, to our knowledge open-ended written
preferences have not been studied. Studies of these
open-ended preferences could highlight issues with respect
to quality end-of-life care. The purpose of this study
was to explore the open-ended proxy, health, and personal
care preferences of people with HIV as expressed in a written
AD form.
An Upper Bound on the Number of Edges of a 2-Connected Graph
Authors: WU POU-LIN
Published: July 2002

Let G be a loopless 2-connected graph whose largest

cycle has c edges, and whose largest bond has

c* edges. We show that

|E(G)| [les ]

½cc*.
Asymptotic Enumeration of Eulerian Circuits
in the Complete Graph
Authors: McKAY BRENDAN D. , ROBINSON ROBERT W.
Published: July 2002

We determine the asymptotic behaviour of the number of Eulerian
circuits in a complete
graph of odd order. One corollary of our result is the following. If a
maximum random
walk, constrained to use each edge at most once, is taken on
Kn, then the probability
that all the edges are eventually used is asymptotic to
e3/4n−½. Some
similar results are
obtained about Eulerian circuits and spanning trees in random regular tournaments.
We
also give exact values for up to 21 nodes.
Use of METEOSAT water‐vapour images for the diagnosis of a vigorous stratospheric intrusion over the central Mediterranean
Authors: Lagouvardos K , Kotroni V
Published: July 2002

The diagnosis of a vigorous dry intrusion over the central Mediterranean is performed using water‐vapour images from METEOSAT. This dry intrusion was located on the rear side of a cold front (propagating from Italy to Greece) and played an important role in the onset of thunderstorms over the western Greek coasts. A combination of satellite imagery and potential vorticity analyses showed that the dry air originated in the lower‐stratospheric and higher‐tropospheric layers. The interaction of the dry air with the moist air masses within the warm conveyor belt ahead of the cold front (overrun of warm air by low equivalent potential temperature air) produced a potentially unstable region over the area of reported thunderstorms. Copyright © 2000 Royal Meteorological Society
The Verbmobil prototype system – a software
engineering perspective
Authors: BUB THOMAS , SCHWINN JOHANNES
Published: July 2002

Verbmobil represents a new generation of speech-to-speech translation systems in which
spontaneously spoken language, speaker independence and adaptability as well as the combination
of deep and shallow approaches to the analysis and transfer problems are the main features.
The project brought together researchers from the fields of signal processing, computational
linguistics and artificial intelligence. Verbmobil goes beyond the state-of-the-art in each of
these areas, but its main achievement is the seamless integration of them. The first project
phase (1993–1996) has been followed up by the second project phase (1997–2000), which
aims at applying the results to further languages and at integrating innovative telecooperation
techniques. Quite apart from the speech and language processing issues, the size and
complexity of the project represent an extreme challenge on the areas of project management
and software engineering:

[bull ] 50 researchers from 29 organizations at different sites in different countries are involved
in the software development process,

[bull ] to reuse existing software, hardware, knowledge and experience, only a few technical
restrictions could be given to the partners.

In this article we describe the Verbmobil prototype system from a software-engineering
perspective. We discuss:

[bull ] the modularized functional architecture,

[bull ] the flexible and extensible software architecture which reflects that functional architecture,

[bull ] the evolutionary process of system integration,

[bull ] the communication-based organizational structure of the project,

[bull ] the evaluation of the system operational by the end of the first project phase.
CONVERGENCE IN MONETARY INFLATION MODELS WITH HETEROGENEOUS LEARNING RULES
Authors: Evans George W. , Honkapohja Seppo , Marimon Ramon
Published: July 2002

Inflation and the monetary financing of deficits are analyzed in a model in which the deficit is constrained to be less than a given fraction of a measure of aggregate market activity. Depending on parameter values, the model can have multiple steady states. Under adaptive learning with heterogeneous learning rules, there is convergence to a subset of these steady states. In some cases, a high-inflation constrained steady state will emerge. However, with a sufficiently tight fiscal constraint, the low-inflation steady state is globally stable. We provide experimental evidence in support of our theoretical results.
Osa-containing Brahma chromatin remodeling complexes are required for the repression of Wingless target genes
Authors: Collins Russell T. , Treisman Jessica E.
Published: July 2002

The Wingless signaling pathway directs many developmental processes in Drosophila by regulating the expression of specific downstream target genes. We report here that the product of thetrithorax group gene osa is required to repress such genes in the absence of the Wingless signal. The Wingless-regulated genes nubbin, Distal-less, and decapentaplegicand a minimal enhancer from the Ultrabithorax gene are misexpressed in osa mutants and repressed by ectopic Osa. Osa-mediated repression occurs downstream of the up-regulation of Armadillo but is sensitive both to the relative levels of activating Armadillo/Pangolin and repressing Groucho/Pangolin complexes present and to the responsiveness of the promoter to Wingless. Osa functions as a component of the Brahma chromatin-remodeling complex; other components of this complex are likewise required to repress Wingless target genes. These results suggest that altering the conformation of chromatin is an important mechanism by which Wingless signaling activates gene expression.
Using viral species specificity to define a critical protein/RNA interaction surface
Authors: Coburn Glen A. , Wiegand Heather L. , Kang Yibin , Ho Dona N. , Georgiadis Millie M. , Cullen Bryan R.
Published: July 2002

The Tap protein mediates the sequence-specific nuclear export of mRNAs bearing the retroviral constitutive transport element (CTE) and also plays a critical role in the sequence nonspecific export of cellular mRNAs. Previously, we have demonstrated that CTE function displays species specificity, that is, the CTE functions in human but not quail cells. Here, we demonstrate that quail Tap fails to support CTE function because it cannot bind the CTE. However, changing a single residue in quail Tap, glutamine 246, to arginine, the residue found in human Tap, rescues both CTE function and CTE binding. This residue, which is located on the exterior of a recently reported molecular structure of Tap, defines a surface on Tap that is critical for CTE binding. These data emphasize the potential importance of cross-species genetic complementation in the identification and characterization of cellular factors that are critical for different aspects of viral replication.
Pyruvate,Orthophosphate Dikinase in Leaves and Chloroplasts of C3 Plants Undergoes Light-/Dark-Induced Reversible Phosphorylation
Authors: Chastain Chris J. , Fries Jason P. , Vogel Julie A. , Randklev Christa L. , Vossen Adam P. , Dittmer Sharon K. , Watkins Erin E. , Fiedler Lucas J. , Wacker Sarah A. , Meinhover Katherine C. , Sarath Gautam , Chollet Raymond
Published: July 2002

Pyruvate,orthophosphate (Pi) dikinase (PPDK) is best recognized as a chloroplastic C4 cycle enzyme. As one of the key regulatory foci for controlling flux through this photosynthetic pathway, it is strictly and reversibly regulated by light. This light/dark modulation is mediated by reversible phosphorylation of a conserved threonine residue in the active-site domain by the PPDK regulatory protein (RP), a bifunctional protein kinase/phosphatase. PPDK is also present in C3 plants, although it has no known photosynthetic function. Nevertheless, in this report we show that C3 PPDK in leaves of several angiosperms and in isolated intact spinach (Spinacia oleracea) chloroplasts undergoes light-/dark-induced changes in phosphorylation state in a manner similar to C4 dikinase. In addition, the kinetics of this process closely resemble the reversible C4 process, with light-induced dephosphorylation occurring rapidly (≤15 min) and dark-induced phosphorylation occurring much more slowly (≥30–60 min). In intact spinach chloroplasts, light-induced dephosphorylation of C3 PPDK was shown to be dependent on exogenous Pi and photosystem II activity but independent of electron transfer from photosystem I. These in organello results implicate a role for stromal pools of Pi and adenylates in regulating the reversible phosphorylation of C3-PPDK. Last, we used an in vitro RP assay to directly demonstrate ADP-dependent PPDK phosphorylation in desalted leaf extracts of the C3 plantsVicia faba and rice (Oryza sativa). We conclude that an RP-like activity mediates the light/dark modulation of PPDK phosphorylation state in C3 leaves and chloroplasts and likely represents the ancestral isoform of this unusual and key C4 pathway regulatory “converter” enzyme.
Inhibitors of Protein Kinases and Phosphatases Alter Root Morphology and Disorganize Cortical Microtubules
Authors: Baskin T. I. , Wilson J. E.
Published: July 2002

To investigate molecular mechanisms controlling plant morphogenesis, we examined the morphology of primary roots of Arabidopsis thaliana and the organization of cortical microtubules in response to inhibitors of serine/threonine protein phosphatases and kinases. We found that cantharidin, an inhibitor of types 1 and 2A protein phosphatases, as previously reported for okadaic acid and calyculin A (R.D. Smith, J.E. Wilson, J.C. Walker, T.I. Baskin [1994] Planta 194: 516–524), inhibited elongation and stimulated radial expansion. Of the protein kinase inhibitors tested, chelerythrine, 6-dimethylaminopurine, H-89, K252a, ML-9, and staurosporine all inhibited elongation, but only staurosporine appreciably stimulated radial expansion. To determine the basis for the root swelling, we examined cortical microtubules in semithin sections of material embedded in butyl-methyl-methacrylate. Chelerythrine and 100 nM okadaic acid, which inhibited elongation without causing swelling, did not change the appearance of cortical arrays, but calyculin A, cantharidin, and staurosporine, which caused swelling, disorganized cortical microtubules. The stability of the microtubules in the aberrant arrays was not detectably different from those in control arrays, as judged by similar sensitivity to depolymerization by cold or oryzalin. These results identify protein phosphorylation and dephosphorylation
Gibberellin Deficiency and Response Mutations Suppress the Stem Elongation Phenotype of Phytochrome-Deficient Mutants of Arabidopsis
Authors: Peng J. , Harberd N. P.
Published: July 2002

Plant growth and development are regulated by numerous internal and external factors. Among these, gibberellin (GA) (an endogenous plant growth regulator) and phytochrome (a photoreceptor) often influence the same processes. For example, in plants grown in the light Arabidopsis thaliana hypocotyl elongation is reduced by GA deficiency and increased by phytochrome deficiency. Here we describe experiments in which the phenotypes of Arabidopsis plants doubly homozygous for GA-related and phytochrome-related mutations were examined. The double mutants were studied at various stages in the plant life cycle, including the seed germination, young seedling, adult, and reproductive phases of development. The results of these experiments are complex, but indicate that a fully functional GA system is necessary for full expression of the elongated phenotypes conferred by phytochrome deficiency.
Requirement for the H Phosphoprotein in Photosystem II of Chlamydomonas reinhardtii
Authors: Summer E. J. , Schmid VHR. , Bruns B. U. , Schmidt G. W.
Published: July 2002

To dissect the expression of the psbB gene cluster of the Chlamydomonas reinhardtii chloroplast genome and to assess the role of the photosystem II H-phosphoprotein (PSII-H) in the biogenesis and/or stabilization of PSII, an aadA gene cassette conferring spectinomycin resistance was employed for mutagenesis. Disruption of the gene cluster has no effect on the abundance of transcripts of the upstream psbB/T locus. Likewise, interruption of psbB/T and psbH with a strong transcriptional terminator from the rbcL gene does not influence transcript accumulation. Thus, psbB/T and psbH may be independently transcribed, and the latter gene seems to have its own promoter in C. reinhardtii. In the absence of PSII-H, translation and thylakoid insertion of chloroplast PSII core proteins is unaffected, but PSII proteins do not accumulate. Because the deletion mutant also exhibits PSII deficiency when dark-grown, the effect is unrelated to photoinhibition. Turnover of proteins B and C of PSII and the polypeptides PSII protein A and PSII protein D is faster than in wild-type cells but is much slower than that observed in other PSII-deficient mutants of C. reinhardtii, suggesting a peripheral location of PSII-H in PSII. The role of PSII-H on PSII assembly was examined by sucrose gradient fractionation of pulse-labeled thylakoids; the accumulation of high-molecular-weight forms of PSII is severely impaired in the psbH deletion mutant. Thus, a primary role of PSII-H may be to facilitate PSII assembly/stability through dimerization. PSII-H phosphorylation, which possibly occurs at two sites, may also be germane to its role in regulating PSII structure, stabilization, or activity.
Intracellular Carbonic Anhydrase Is Essential to Photosynthesis in Chlamydomonas reinhardtii at Atmospheric Levels of CO2 (Demonstration via Genomic Complementation of the High-CO2-Requiring Mutant ca-1)
Authors: Funke R. P. , Kovar J. L. , Weeks D. P.
Published: July 2002

Genomic complementation of the high-CO2-requiring mutant ca-1–12-1C of Chlamydomonas reinhardtii was achieved by transformation with DNA pools from an indexed cosmid library of wild-type genomic DNA. Transformation of mutant cells with cosmid DNA from two microtiter plates in the library produced colonies that grew phototrophically at atmospheric CO2 levels. Transformations with cosmid DNA from each of the rows and files of the two plates pinpointed one well in each plate with a cosmid bearing the targeted gene. Sequencing of cosmid subclones revealed a gene encoding a recently identified C. reinhardtii chloroplast carbonic anhydrase (CAH3). Transformations with chimeric constructs combining different portions of the wild-type and mutant genes indicated the presence of a mutation in the 5[prime]-half of the gene. Comparison of mutant and wild-type gene sequences in this region revealed a G-to-A substitution in the mutant gene, which produced a nonsense codon. The data presented demonstrate that the carbonic anhydrase produced from the CAH3 gene is essential to the inorganic carbon-concentrating mechanism in C. reinhardtii and that genomic complementation can be a facile and efficient means for isolating genes associated with defects affecting photosynthesis and other physiological processes in this eukaryotic green alga.
Characterization of New Gibberellin-Responsive Semidwarf Mutants of Arabidopsis
Authors: Sponsel V. M. , Schmidt F. W. , Porter S. G. , Nakayama M. , Kohlstruk S. , Estelle M.
Published: July 2002

Chemical mutagenesis of Arabidopsis thaliana (L.) Heynh. yielded four semidwarf mutants, all of which appeared to be gibberellin (GA)-biosynthesis mutants. All four had atypical response profiles to C20-GAs, suggesting that each had impaired 20-oxidation. One mutant, 11.2, was shown to be allelic to ga5 and has been named ga5–2. It had altered metabolism of [14C]GA15 relative to that in wild-type plants and undetectable levels of C19-GAs in young stems, consistent with the known function of GA5 as a stem-expressed GA 20-oxidase. Two mutants (2.1 and 10.3), which had very short inflorescences and siliques, were allelic to each other but not to the known GA-responding mutants, ga1 to ga5. The locus defined by these two mutations is provisionally named GA6 and is purported to encode an inflorescence- and silique-expressed GA 20-oxidase. A double mutant, ga5–2 ga6–2, had an extreme dwarf phenotype with very short siliques. The fourth mutation, 1.1, gave a phenotype like ga5, but was not allelic to any of the known ga mutations. It has not yet been given a gene symbol pending further studies.
Three Drought-Responsive Members of the Nonspecific Lipid-Transfer Protein Gene Family inLycopersicon pennelliiShow Different Developmental Patterns of Expression1
Authors: Treviño Marcela B. , Connell Mary A. O'
Published: July 2002

Genomic clones of two nonspecific lipid-transfer protein genes from a drought-tolerant wild species of tomato (Lycopersicon pennellii Corr.) were isolated using as a probe a drought- and abscisic acid (ABA)-induced cDNA clone (pLE16) from cultivated tomato (Lycopersicon esculentumMill.). Both genes (LpLtp1 and LpLtp2) were sequenced and their corresponding mRNAs were characterized; they are both interrupted by a single intron at identical positions and predict basic proteins of 114 amino acid residues. Genomic Southern data indicated that these genes are members of a small gene family inLycopersicon spp. The 3′-untranslated regions fromLpLtp1 and LpLtp2, as well as a polymerase chain reaction-amplified 3′-untranslated region from pLE16 (cross-hybridizing to a third gene in L. pennellii, namely LpLtp3), were used as gene-specific probes to describe expression in L. pennellii through northern-blot analyses. All LpLtp genes were exclusively expressed in the aerial tissues of the plant and all were drought and ABA inducible. Each gene had a different pattern of expression in fruit, and LpLtp1 and LpLtp2, unlikeLpLtp3, were both primarily developmentally regulated in leaf tissue. Putative ABA-responsive elements were found in the proximal promoter regions of LpLtp1 andLpLtp2.
Regulation of Oleoresinosis in Grand Fir (Abies grandis)1
Authors: Steele Christopher L. , Katoh Sadanobu , Bohlmann Jörg , Croteau Rodney
Published: July 2002

Grand fir (Abies grandis Lindl.) has been developed as a model system for the study of wound-induced oleoresinosis in conifers as a response to insect attack. Oleoresin is a roughly equal mixture of turpentine (85% monoterpenes [C10] and 15% sesquiterpenes [C15]) and rosin (diterpene [C20] resin acids) that acts to seal wounds and is toxic to both invading insects and their pathogenic fungal symbionts. The dynamic regulation of wound-induced oleoresin formation was studied over 29 d at the enzyme level by in vitro assay of the three classes of synthases directly responsible for the formation of monoterpenes, sesquiterpenes, and diterpenes from the corresponding C10, C15, and C20 prenyl diphosphate precursors, and at the gene level by RNA-blot hybridization using terpene synthase class-directed DNA probes. In overall appearance, the shapes of the time-course curves for all classes of synthase activities are similar, suggesting coordinate formation of all of the terpenoid types. However, closer inspection indicates that the monoterpene synthases arise earlier, as shown by an abbreviated time course over 6 to 48 h. RNA-blot analyses indicated that the genes for all three classes of enzymes are transcriptionally activated in response to wounding, with the monoterpene synthases up-regulated first (transcripts detectable 2 h after wounding), in agreement with the results of cell-free assays of monoterpene synthase activity, followed by the coordinately regulated sesquiterpene synthases and diterpene synthases (transcription beginning on d 3–4). The differential timing in the production of oleoresin components of this defense response is consistent with the immediate formation of monoterpenes to act as insect toxins and their later generation at solvent levels for the mobilization of resin acids responsible for wound sealing.
Systemin Potentiates the Oxidative Burst in Cultured Tomato Cells1
Authors: Stennis Michael J. , Chandra Sreeganga , Ryan Clarence A. , Low Philip S.
Published: July 2002

Plants that have been wounded by insects or other herbivores may be more susceptible to infection by adventitious microbes. Wound-induced signal molecules, which serve to induce responses in the plant that retard further feeding, might also act to prepare a plant for possible pathogen attack. We have examined the effect of a wound-generated systemic messenger (systemin) on a pathogen-stimulated defense-response marker, the oxidative burst. We observed that neither systemin nor its inactive analog (A-17) was able to directly induce H2O2 biosynthesis in suspension-cultured tomato (Lycopersicon esculentum L.) cells, regardless of the duration of exposure of the cells to the two peptides. Similarly, neither systemin nor A-17 was capable of modifying an oligogalacturonide-elicited oxidative burst, as long as elicitor addition occurred within minutes of treatment with systemin or A-17. In contrast, preexposure of the cell cultures to systemin (but not to A-17) led to a time-dependent enhancement of the oligogalacturonide-elicited oxidative burst. By 12 h of exposure, the H2O2 biosynthetic capacity of systemin-treated cells exceeded that of the control cells by a factor of 16 ± 2. A similar up-regulation by systemin of a mechanically stimulated oxidative burst was also observed. Because the systemin-induced augmentation in oxidant synthesis is quantitatively prevented by coincubation with 2 μm cycloheximide, and because the oxidative burst of oligogalacturonic acid-elicited control cells (no systemin exposure) is unaffected by preincubation with cycloheximide, we conclude that systemin enhancement of the tomato-cell oxidative burst requires protein synthesis.
The Site of Oxygen Limitation in Soybean Nodules1
Authors: Kuzma Monika M. , Winter Heike , Storer Paul , Oresnik Ivan , Atkins Craig A. , Layzell David B.
Published: July 2002

In legume nodules the [O2] in the infected cells limits respiration and nitrogenase activity, becoming more severe if nodules are exposed to subambient O2 levels. To identify the site of O2 limitation, adenylate pools were measured in soybean (Glycine max) nodules that were frozen in liquid N2 before being ground, lyophilized, sonicated, and separated on density gradients of nonaqueous solvents (heptane/tetrachloroethylene) to yield fractions enriched in bacteroid or plant components. In nodules maintained in air, the adenylate energy charge (AEC = [ATP + 0.5 ADP]/[ATP + ADP + AMP]) was lower in the plant compartment (0.65 ± 0.04) than in the bacteroids (0.76 ± 0.095), but did not change when the nodulated root system was exposed to 10% O2. In contrast, 10% O2decreased the bacteroid AEC to 0.56 ± 0.06, leading to the conclusion that they are the primary site of O2 limitation in nodules. To account for the low but unchanged AEC in the plant compartment and for the evidence that mitochondria are localized in O2-enriched microenvironments adjacent to intercellular spaces, we propose that steep adenylate gradients may exist between the site of ATP synthesis (and ADP use) in the mitochondria and the extra-mitochondrial sites of ATP use (and ADP production) throughout the large, infected cells.
Mutations Affecting Induction of Glycolytic and Fermentative Genes during Germination and Environmental Stresses in Arabidopsis1
Authors: Conley Terry R. , Peng Hsiao-Ping , Shih Ming-Che
Published: July 2002

Expression of the alcohol dehydrogenase gene (ADH) of Arabidopsis is known to be induced by environmental stresses and regulated developmentally. We used a negative-selection approach to isolate mutants that were defective in regulating the expression of theADH gene during seed germination; we then characterized three recessive mutants, aar1–1,aar1–2, and aar2–1, which belong to two complementation groups. In addition to their defects during seed germination, mutations in the AAR1 andAAR2 genes also affected anoxic and hypoxic induction ofADH and other glycolytic genes in mature plants. Theaar1 and aar2 mutants were also defective in responding to cold and osmotic stress. The two allelic mutantsaar1–1and aar1–2 exhibited different phenotypes under cold and osmotic stresses. Based on our results we propose that these mutants are defective in a late step of the signaling pathways that lead to increased expression of theADH gene and glycolytic genes.