bluuebunny/crossref_metadata_2025 huggingface dataset:
Definition and interactions of a positive regulatory element of the Arabidopsis INNER NO OUTER promoter
Authors: Meister Robert J. , Williams Luis A. , Monfared Mona M. , Gallagher Thomas L. , Kraft Edward A. , Nelson Christian G. , Gasser Charles S.
Published: January 2004

INNER NO OUTER (INO) expression is limited to the abaxial cell layer of the incipient and developing outer integument in Arabidopsis ovules. Using deletion analysis of the previously defined INO promoter (P‐INO), at least three distinct regions that contribute to the endogenous INO expression pattern were identified. One such positive element, designated POS9, which comprises at least three distinct subelements, was found to include sufficient information to duplicate the INO expression pattern when four or more copies were used in conjunction with a heterologous minimal promoter. While known regulators of INO, including INO, SUPERMAN, BELL1, and AINTEGUMENTA, did not detectably interact with POS9 in yeast one‐hybrid assays, two groups of proteins that interact specifically with POS9 were identified in one‐hybrid library screens. Members of one group include C2H2 zinc finger motifs. Members of the second group contain a novel, conserved DNA‐binding region and were designated the BASIC PENTACYSTEINE (BPC) proteins on the basis of conserved features of this region. The BPC proteins are nuclear localized and specifically bind in vitro to GA dinucleotide repeats located within POS9. The widespread expression patterns of the BPCs and the large number of GA repeat potential target sequences in the Arabidopsis genome indicate that BPC proteins may affect expression of genes involved in a variety of plant processes.
The Effect of Nitrogen Fertilization on Fusarium Head Blight Development and Deoxynivalenol Contamination in Wheat
Authors: Lemmens M. , Haim K. , Lew H. , Ruckenbauer P.
Published: January 2004

The impact of nitrogen (N) fertilization on the development of Fusarium head blight (FHB) in wheat and the resulting deoxynivalenol (DON) contamination in the kernels was studied. In a first experiment, the disease was assessed on two locations under natural infection pressure. Five different types of nitrogen fertilizer (both organic and mineral) were investigated, each applied at five input rates from 0 to 160 kg N/ha. With all fertilizers, a significant increase of disease intensity was observed with increasing N input, while the type of N fertilizer had poor or no effects on FHB. Depending on the fertilizer used, the percentage of diseased spikelets increased from 2.2% at zero N rate up to 6.6% at 160 kg N input per hectare. In a second series of trials, three spring wheat varieties including one Durum wheat line were artificially inoculated with a Fusarium graminearum and a F. culmorum strain, known producers of DON. A mineral N fertilizer was applied at five input levels from 0 to 160 kg N/ha. A significant increase in FHB intensity and DON contamination in the grain was observed with increasing N from 0 to 80 kg/ha. At higher input rates, relevant in contemporary crop husbandry, disease intensity and toxin contamination remained at constant levels. It is concluded that adaptation of N fertilization represents no relevant tool in managing FHB in practical wheat cultivation.
Vegetative Compatibility Among Isolates of Colletotrichum gloeosporioides from Yam (Dioscorea spp.) in Nigeria
Authors: Abang M. M. , Hoffmann P. , Winter S. , Green K. R. , Wolf G. A.
Published: January 2004

Isolates of Colletotrichum gloeosporioides obtained from yam‐based cropping systems in Nigeria, previously characterized on the basis of morphology, virulence and rDNA internal transcribed spacer (ITS) sequence variation were further compared for vegetative compatibility (VC). Chlorate‐resistant nitrate non‐utilizing (nit) mutants were generated from the isolates and used in complementation (heterokaryon) tests. Tests of VC between complementary mutants from different isolates indicated the presence of several genotypes within a single field, suggesting limited clonal spread. In some cases, isolates obtained from the same lesion were observed to belong to different vegetative compatibility groups (VCGs). No compatibility was observed between isolates of the highly virulent slow‐growing grey (SGG), the moderately virulent fast‐growing salmon (FGS) and the avirulent/weakly virulent fast‐growing grey (FGG) strains. Forty‐one C. gloeosporioides isolates belonged to 28 VCGs, giving a genotype diversity estimate of 0.68. This diversity confirmed the high variability of the pathogen population as revealed by previous characterization studies, however, a correlation between VCGs and isolate groupings based on morphology and virulence was not found. The finding that an isolate from weed was compatible with yam isolates indicated that transfer of important traits, such as virulence, may take place between isolates from yam and non‐yam hosts. The VCG diversity revealed by this study suggests that in addition to asexual reproduction, sexual reproduction may play an important role in the epidemiology of anthracnose on yam.
Occurrence of Antimicrobial Serin‐Proteinases in Sunflower Seeds
Authors: Mendieta J. R. , Giudici A. M. , de la Canal L.
Published: January 2004

Using an experimental approach directed to the isolation of antimicrobial proteins, we have detected the presence of a trypsin inhibitor (TI) with associated antifungal activity in sunflower seeds. Purification of the isolated protein by affinity chromatography on a trypsin‐agarose matrix confirmed that a trypsin inhibitor was responsible for the inhibition of spore germination of the fungal pathogen Sclerotinia sclerotiorum. The protein is a potent antifungal compound as it can completely inhibit the germination of S. sclerotiorum ascospores at a concentration of 14 μg/ml. The putative contribution of this TI to control fungal invasion is discussed.
Hypovirulence of Didymella bryoniae Associated with dsRNA
Authors: Urs R. R. , Mayhew D. E. , Roberts P. D.
Published: January 2004

Didymella bryoniae, isolate 98–18, recovered from watermelon seedlings with symptoms of gummy stem blight, showed abnormal growth, mycelial lysis, sectoring, barrage and limited production of fruiting bodies in culture. A dsRNA (approximately 6.5 kbp) was associated with isolate 98–18 and other isolates showing abnormal mycelial growth. Transfer of dsRNA from 98–18 to virulent isolates Tk659 and TK671 via hyphal anastamosis was achieved only by consecutive exposure to 98–18, but not to virulent isolates RJ1 and RJ2. Transfer of dsRNA via infiltration through growth media to virulent isolates RJ1, RJ2, TK659 and TK671 was unsuccessful. The disease severity index of isolate 98–18 was reduced 53% on watermelon, 32% on cataloupe and 26% on yellow squash compared with isolate RJ2, suggesting that the presence of the dsRNA is associated with reduced virulence of D. bryoniae isolate 98–18. Zucchini and yellow squash were resistant to both isolates when tested.
Bcl-2 overexpression corrects mitochondrial defects and ameliorates inherited desmin null cardiomyopathy
Authors: Weisleder Noah , Taffet George E. , Capetanaki Yassemi
Published: January 2004

One of the hallmarks of cardiomyopathy and heart failure is pronounced and progressive cardiomyocyte death. Understanding the mechanisms involved in cardiomyocyte cell death is a topic of great interest for treatment of cardiac disease. Mice null for desmin, the muscle-specific member of the intermediate filament gene family, develop cardiomyopathy characterized by extensive cardiomyocyte death, fibrosis, calcification, and eventual heart failure. The earliest ultrastructural defects are observed in mitochondria. In the present study, we have demonstrated that these mitochondrial abnormalities are the primary cause of the observed cardiomyopathy and that these defects can be ameliorated by overexpression of bcl-2 in desmin null heart. Overexpression of bcl-2 in the desmin null heart results in correction of mitochondrial defects, reduced occurrence of fibrotic lesions in the myocardium, prevention of cardiac hypertrophy, restoration of cardiomyocyte ultrastructure, and significant improvement of cardiac function. Furthermore, we have found that loss of desmin also diminishes the capacity of mitochondria to resist exposure to calcium, a defect that can be partially restored by bcl-2 overexpression. These results point to a unique function for desmin in protection of mitochondria from calcium exposure that can be partially rescued by overexpression of bcl-2. We show that bcl-2 cardiac overexpression has provided significant improvement of an inherited form of cardiomyopathy, revealing the potential forbcl-2, and perhaps other genes in the family, as therapeutic agents for heart disease of many types, including inherited forms.
Gene expression profiling identifies clinically relevant subtypes of prostate cancer
Authors: Lapointe Jacques , Li Chunde , Higgins John P. , van de Rijn Matt , Bair Eric , Montgomery Kelli , Ferrari Michelle , Egevad Lars , Rayford Walter , Bergerheim Ulf , Ekman Peter , DeMarzo Angelo M. , Tibshirani Robert , Botstein David , Brown Patrick O. , Brooks James D. , Pollack Jonathan R.
Published: January 2004

Prostate cancer, a leading cause of cancer death, displays a broad range of clinical behavior from relatively indolent to aggressive metastatic disease. To explore potential molecular variation underlying this clinical heterogeneity, we profiled gene expression in 62 primary prostate tumors, as well as 41 normal prostate specimens and nine lymph node metastases, using cDNA microarrays containing ≈26,000 genes. Unsupervised hierarchical clustering readily distinguished tumors from normal samples, and further identified three subclasses of prostate tumors based on distinct patterns of gene expression. High-grade and advanced stage tumors, as well as tumors associated with recurrence, were disproportionately represented among two of the three subtypes, one of which also included most lymph node metastases. To further characterize the clinical relevance of tumor subtypes, we evaluated as surrogate markers two genes differentially expressed among tumor subgroups by using immunohistochemistry on tissue microarrays representing an independent set of 225 prostate tumors. Positive staining for MUC1, a gene highly expressed in the subgroups with “aggressive” clinicopathological features, was associated with an elevated risk of recurrence (
P
= 0.003), whereas strong staining for AZGP1, a gene highly expressed in the other subgroup, was associated with a decreased risk of recurrence (
P
= 0.0008). In multivariate analysis, MUC1 and AZGP1 staining were strong predictors of tumor recurrence independent of tumor grade, stage, and preoperative prostate-specific antigen levels. Our results suggest that prostate tumors can be usefully classified according to their gene expression patterns, and these tumor subtypes may provide a basis for improved prognostication and treatment stratification.
Mislocalization to the nuclear envelope: An effect of the dystonia-causing torsinA mutation
Authors: Goodchild Rose E. , Dauer William T.
Published: January 2004

Primary dystonia is a disease characterized by involuntary twisting movements caused by CNS dysfunction without underlying histopathology. DYT1 dystonia is a form of primary dystonia caused by an in-frame GAG deletion (ΔE302/3) in the
TOR1A
gene that encodes the endoplasmic reticulum luminal protein torsinA. We show that torsinA is also present in the nuclear envelope (NE), where it appears to interact with substrate, and that the ΔE302/3 mutation causes a striking redistribution of torsinA from the endoplasmic reticulum to the NE. In addition, ΔE302/3-torsinA recruits WT torsinA to the NE, potentially providing insight into an understanding of the dominant inheritance of the disease. DYT1 dystonia appears to be a previously uncharacterized NE disease and the first, to our knowledge, to selectively affect CNS function.
High-resolution molecular structure of a peptide in an amyloid fibril determined by magic angle spinning NMR spectroscopy
Authors: Jaroniec Christopher P. , MacPhee Cait E. , Bajaj Vikram S. , McMahon Michael T. , Dobson Christopher M. , Griffin Robert G.
Published: January 2004

Amyloid fibrils are self-assembled filamentous structures associated with protein deposition conditions including Alzheimer's disease and the transmissible spongiform encephalopathies. Despite the immense medical importance of amyloid fibrils, no atomic-resolution structures are available for these materials, because the intact fibrils are insoluble and do not form diffraction-quality 3D crystals. Here we report the high-resolution structure of a peptide fragment of the amyloidogenic protein transthyretin, TTR(105–115), in its fibrillar form, determined by magic angle spinning NMR spectroscopy. The structure resolves not only the backbone fold but also the precise conformation of the side chains. Nearly complete
13
C and
15
N resonance assignments for TTR(105–115) formed the basis for the extraction of a set of distance and dihedral angle restraints. A total of 76 self-consistent experimental measurements, including 41 restraints on 19 backbone dihedral angles and 35
13
C–
15
N distances between 3 and 6 Å were obtained from 2D and 3D NMR spectra recorded on three fibril samples uniformly
13
C,
15
N-labeled in consecutive stretches of four amino acids and used to calculate an ensemble of peptide structures. Our results indicate that TTR(105–115) adopts an extended β-strand conformation in the amyloid fibrils such that both the main- and side-chain torsion angles are close to their optimal values. Moreover, the structure of this peptide in the fibrillar form has a degree of long-range order that is generally associated only with crystalline materials. These findings provide an explanation of the unusual stability and characteristic properties of this form of polypeptide assembly.
Multiplier operator algebras and applications
Authors: Blecher David P. , Zarikian Vrej
Published: January 2004

The one-sided multipliers of an operator space
X
are a key to “latent operator algebraic structure” in
X
. We begin with a survey of these multipliers, together with several of the applications that they have had to operator algebras. We then describe several new results on one-sided multipliers, and new applications, mostly to one-sided
M
-ideals.
Ectopic expression of the homeobox gene
Cdx2
is the transforming event in a mouse model of t(12;13)(p13;q12) acute myeloid leukemia
Authors: Rawat Vijay P. S. , Cusan Monica , Deshpande Aniruddha , Hiddemann Wolfgang , Quintanilla-Martinez Leticia , Humphries R. Keith , Bohlander Stefan K. , Feuring-Buske Michaela , Buske Christian
Published: January 2004

Creation of fusion genes by balanced chromosomal translocations is one of the hallmarks of acute myeloid leukemia (AML) and is considered one of the key leukemogenic events in this disease. In t(12;13)(p13;q12) AML, ectopic expression of the homeobox gene
CDX2
was detected in addition to expression of the
ETV6-CDX2
fusion gene, generated by the chromosomal translocation. Here we show in a murine model of t(12;13)(p13;q12) AML that myeloid leukemogenesis is induced by the ectopic expression of
CDX2
and not by the
ETV6-CDX2
chimeric gene. Mice transplanted with bone marrow cells retrovirally engineered to express
Cdx2
rapidly succumbed to fatal and transplantable AML. The transforming capacity of
Cdx2
depended on an intact homeodomain and the N-terminal transactivation domain. Transplantation of bone marrow cells expressing
ETV6-CDX2
failed to induce leukemia. Furthermore, coexpression of
ETV6-CDX2
and
Cdx2
in bone marrow cells did not accelerate the course of disease in transplanted mice compared to
Cdx2
alone. These data demonstrate that activation of a protooncogene by a balanced chromosomal translocation can be the pivotal leukemogenic event in AML, characterized by the expression of a leukemia-specific fusion gene. Furthermore, these findings link protooncogene activation to myeloid leukemogenesis, an oncogenic mechanism so far associated mainly with lymphoid leukemias and lymphomas.
DNA nanotubes self-assembled from triple-crossover tiles as templates for conductive nanowires
Authors: Liu Dage , Park Sung Ha , Reif John H. , LaBean Thomas H.
Published: January 2004

DNA-based nanotechnology is currently being developed as a general assembly method for nanopatterned materials that may find use in electronics, sensors, medicine, and many other fields. Here we present results on the construction and characterization of DNA nanotubes, a self-assembling superstructure composed of DNA tiles. Triple-crossover tiles modified with thiol-containing double-stranded DNA stems projected out of the tile plane were used as the basic building blocks. Triple-crossover nanotubes display a constant diameter of ≈25 nm and have been observed with lengths up to 20 μm. We present high-resolution images of the constructs, experimental evidence of their tube-like nature as well as data on metallization of the nanotubes to form nanowires, and electrical conductivity measurements through the nanowires. DNA nanotubes represent a potential breakthrough in the self-assembly of nanometer-scale circuits for electronics layout because they can be targeted to connect at specific locations on larger-scale structures and can subsequently be metallized to form nanometer-scale wires. The dimensions of these nanotubes are also perfectly suited for applications involving interconnection of molecular-scale devices with macroscale components fabricated by conventional photolithographic methods.
A respiratory hemocyanin from an insect
Authors: Hagner-Holler Silke , Schoen Axel , Erker Wolfgang , Marden James H. , Rupprecht Rainer , Decker Heinz , Burmester Thorsten
Published: January 2004

Insects possess an elaborate tracheal system that enables transport of gaseous oxygen from the atmosphere directly to the inner organs. Therefore, the presence of specialized oxygen-transport proteins in the circulatory system of insects has been considered generally unnecessary. Here, we show for the first time, to our knowledge, the presence of an ancestral and functional hemocyanin (Hc) in an insect. In the hemolymph of nymphs and adults of the stonefly
Perla marginata
, a hexameric Hc was identified, which consists of two distinct subunit types of 659 and 655 amino acids.
P. marginata
Hc displays cooperative oxygen binding with a moderately high oxygen affinity [(half-saturation pressure,
P
50
≈8 torr (1 torr = 133 Pa)]. No evidence was found for the presence of Hcs in the more evolutionarily advanced holometabolan insects, suggesting that this type of respiratory protein was lost later in insect evolution. However, our results demonstrate that, in contrast to the accepted paradigm, certain basal insects have retained an ancestral blood-based mechanism of gas exchange.
Phthalate-induced Leydig cell hyperplasia is associated with multiple endocrine disturbances
Authors: Akingbemi Benson T. , Ge Renshan , Klinefelter Gary R. , Zirkin Barry R. , Hardy Matthew P.
Published: January 2004

The possibility that exposures to environmental agents are associated with reproductive disorders in human populations has generated much public interest recently. Phthalate esters are used most commonly as plasticizers in the food and construction industry, and di-(2-ethylhexyl) phthalate (DEHP) is the most abundant phthalate in the environment. Daily human exposure to DEHP in the U.S. is significant, and occupational and clinical exposures from DEHP-plasticized medical devices, e.g., blood bags, hemodialysis tubing, and nasogastric feeding tubes, increase body burden levels. We investigated the effects of chronic exposures to low environmentally relevant DEHP levels on testicular function. Our data show that prolonged exposures to this agent induced high levels of the gonadotropin luteinizing hormone and increased the serum concentrations of sex hormones [testosterone and 17β-estradiol (E2)] by >50%. Increased proliferative activity in Leydig cells was evidenced by enhanced expression of cell cycle proteins, as determined by RT-PCR. The numbers of Leydig cells in the testis of DEHP-treated rats were 40–60% higher than in control rats, indicating induction of Leydig cell hyperplasia. DEHP-induced elevations in serum testosterone and E2 levels suggest the possibility of multiple crosstalks between androgen, estrogen, and steroid hormone receptors, whereas the presence of estrogen receptors in nonreproductive tissues, e.g., cardiovascular system and bones, implies that the increases in serum E2 levels have implications beyond reproduction, including systemic physiology. Analysis of the effects of phthalate exposures on gonadotropin and steroid hormone levels should form part of overall risk assessment in human populations.
Increased primary tumor growth in mice null for β3- or β3/β5-integrins or selectins
Authors: Taverna Daniela , Moher Heather , Crowley Denise , Borsig Lubor , Varki Ajit , Hynes Richard O.
Published: January 2004

Expression of αvβ3- or αvβ5-integrins and selectins is widespread on blood cells and endothelial cells. Here we report that human tumor cells injected s.c. into mice lacking β3- or β3/β5-integrins or various selectins show enhanced tumor growth compared with growth in control mice. There was increased angiogenesis in mice lacking β3-integrins, but no difference in structure of the vessels was observed by histology or by staining for NG2 and smooth muscle actin in pericytes. Bone marrow transplants suggest that the absence of β3-integrins on bone marrow-derived host cells contributes to the enhanced tumor growth in β3-null mice, although few, if any, bone marrow-derived endothelial cells were found in the tumor vasculature. Tumor growth also was affected by bone marrow-derived cells in mice lacking any one or all three selectins, implicating both leukocyte and endothelial selectins in tumor suppression. Reduced infiltration of macrophages was observed in tumors grown in mice lacking either β3-integrins or selectins. These results implicate cells of the innate immune system, macrophages or perhaps natural killer cells, in each case dependent on integrins and selectins, in tumor suppression.
Global gene repression by KaiC as a master process of prokaryotic circadian system
Authors: Nakahira Yoichi , Katayama Mitsunori , Miyashita Hiroshi , Kutsuna Shinsuke , Iwasaki Hideo , Oyama Tokitaka , Kondo Takao
Published: January 2004

A
kaiABC
clock gene cluster was previously identified from cyanobacterium
Synechococcus elongatus
PCC 7942, and the feedback regulation of
kai
genes was proposed as the core mechanism generating circadian oscillation. In this study, we confirmed that the Kai-based oscillator is the dominant circadian oscillator functioning in cyanobacteria. We probed the nature of this regulation and found that excess KaiC represses not only
kaiBC
but also the rhythmic components of all genes in the genome. This result strongly suggests that the KaiC protein primarily coordinates genomewide gene expression, including its own expression. We also found that a promoter derived from
E. coli
is feedback controlled by KaiC and restores the complete circadian rhythm in
kaiBC
-inactivated arrhythmic mutants, provided it can express
kaiB
and
kaiC
genes at an appropriate level. Unlike eukaryotic models, specific regulation of the
kaiBC
promoter is not essential for cyanobacterial circadian oscillations.
Hydrogen storage in molecular compounds
Authors: Mao Wendy L. , Mao Ho-kwang
Published: January 2004

At low temperature (
T
) and high pressure (
P
), gas molecules can be held in ice cages to form crystalline molecular compounds that may have application for energy storage. We synthesized a hydrogen clathrate hydrate, H
2
(H
2
O)
2
, that holds 50 g/liter hydrogen by volume or 5.3 wt %. The clathrate, synthesized at 200–300 MPa and 240–249 K, can be preserved to ambient
P
at 77 K. The stored hydrogen is released when the clathrate is warmed to 140 K at ambient
P
. Low
T
also stabilizes other molecular compounds containing large amounts of molecular hydrogen, although not to ambient
P
, e.g., the stability field for H
2
(H
2
O) filled ice (11.2 wt % molecular hydrogen) is extended from 2,300 MPa at 300 K to 600 MPa at 190 K, and that for (H
2
)
4
CH
4
(33.4 wt % molecular hydrogen) is extended from 5,000 MPa at 300 K to 200 MPa at 77 K. These unique characteristics show the potential of developing low-
T
molecular crystalline compounds as a new means for hydrogen storage.
Chemogenomic profiling: Identifying the functional interactions of small molecules in yeast
Authors: Giaever Guri , Flaherty Patrick , Kumm Jochen , Proctor Michael , Nislow Corey , Jaramillo Daniel F. , Chu Angela M. , Jordan Michael I. , Arkin Adam P. , Davis Ronald W.
Published: January 2004

We demonstrate the efficacy of a genome-wide protocol in yeast that allows the identification of those gene products that functionally interact with small molecules and result in the inhibition of cellular proliferation. Here we present results from screening 10 diverse compounds in 80 genome-wide experiments against the complete collection of heterozygous yeast deletion strains. These compounds include anticancer and antifungal agents, statins, alverine citrate, and dyclonine. In several cases, we identified previously known interactions; furthermore, in each case, our analysis revealed novel cellular interactions, even when the relationship between a compound and its cellular target had been well established. In addition, we identified a chemical core structure shared among three therapeutically distinct compounds that inhibit theERG24heterozygous deletion strain, demonstrating that cells may respond similarly to compounds of related structure. The ability to identify on-and-off target effectsin vivois fundamental to understanding the cellular response to small-molecule perturbants.
Persistence of a small subpopulation of cancer stem-like cells in the C6 glioma cell line
Authors: Kondo Toru , Setoguchi Takao , Taga Tetsuya
Published: January 2004

Both stem cells and cancer cells are thought to be capable of unlimited proliferation. Paradoxically, however, some cancers seem to contain stem-like cells (cancer stem cells). To help resolve this paradox, we investigated whether established malignant cell lines, which have been maintained for years in culture, contain a subpopulation of stem cells. In this article, we show that many cancer cell lines contain a small side population (SP), which, in many normal tissues, is thought to contain the stem cells of the tissue. We demonstrate that in the absence of serum the combination of basic fibroblast growth factor and platelet-derived growth factor maintains SP cells in the C6 glioma cell line. Moreover, we show that C6 SP cells, but not non-SP cells, can generate both SP and non-SP cells in culture and are largely responsible for the
in vivo
malignancy of this cell line. Finally, we provide evidence that C6 SP cells can produce both neurons and glial cells
in vitro
and
in vivo
. We propose that many cancer cell lines contain a minor subpopulation of stem cells that is enriched in an SP, can be maintained indefinitely in culture, and is crucial for their malignancy.
Prominent roles of the NorR and Fur regulators in the
Escherichia coli
transcriptional response to reactive nitrogen species
Authors: Mukhopadhyay Partha , Zheng Ming , Bedzyk Laura A. , LaRossa Robert A. , Storz Gisela
Published: January 2004

We examined the genomewide transcriptional responses of
Escherichia coli
treated with nitrosylated glutathione or the nitric oxide (NO)-generator acidified sodium nitrite (NaNO
2
) during aerobic growth. These assays showed that NorR, a homolog of NO-responsive transcription factors in
Ralstonia eutrophus
, and Fur, the global repressor of ferric ion uptake, are major regulators of the response to reactive nitrogen species. In contrast, SoxR and OxyR, regulators of the
E. coli
defenses against superoxide-generating compounds and hydrogen peroxide, respectively, have minor roles. Moreover, additional regulators of the
E. coli
response to reactive nitrogen species remain to be identified because several of the induced genes were regulated normally in
norR
,
fur
,
soxRS
, and
oxyR
mutant strains. We propose that the
E. coli
transcriptional response to reactive nitrogen species is a composite response mediated by the modification of multiple transcription factors containing iron or redox-active cysteines, some specifically designed to sense NO and its derivatives and others that are collaterally activated by the reactive nitrogen species.
Antiatherogenic activity of fungal beauveriolides, inhibitors of lipid droplet accumulation in macrophages
Authors: Namatame Ichiji , Tomoda Hiroshi , Ishibashi Shun , Ōmura Satoshi
Published: January 2004

Beauveriolides I and III, isolated from the culture broth of fungal
Beauveria
sp. FO-6979, showed potent inhibitory activity of lipid droplet accumulation in primary mouse peritoneal macrophages. The cellular molecular target of this inhibitory activity was studied in macrophages. Beauveriolides I and III strongly inhibited the cholesteryl ester (CE) synthesis with IC
50
values of 0.78 and 0.41 μM, respectively, without showing significant effects on the triacylglycerol and phospholipid synthesis. Furthermore, lysosomal cholesterol metabolism to CE in macrophages was inhibited by the compounds, indicating that the inhibition site lies within steps between cholesterol departure from the lysosome and CE synthesis in the endoplasmic reticulum. Therefore, acyl-CoA:cholesterol acyltransferase (ACAT) activity in the membrane fractions prepared from mouse macrophages was studied, resulting in a dose-dependent inhibition by beauveriolides I and III with IC
50
values of 6.0 and 5.5 μM, respectively. Thus, we showed that the beauveriolides inhibit macrophage ACAT activity specifically, resulting in blockage of the CE synthesis, leading to a reduction of lipid droplets in macrophages. ACAT activity in the membrane fractions prepared from mouse liver and Caco-2 cells was also inhibited, indicating that the beauveriolides block both ACAT-1 and -2. Moreover, beauveriolides I and III exert antiatherogenic activity in both low-density lipoprotein receptor- and apolipoprotein E-knockout mice without any side effects such as diarrhea or cytotoxicity to adrenal tissues as observed for many synthetic ACAT inhibitors. Beauveriolides I and III are the first microbial cyclodepsipeptides having an
in vivo
antiatherosclerotic effect and show promise as potential lead compounds for antiatherosclerotic agents.
A form of long-lasting, learning-related synaptic plasticity in the hippocampus induced by heterosynaptic low-frequency pairing
Authors: Huang Yan-You , Pittenger Christopher , Kandel Eric R.
Published: January 2004

The late, transcription- and translation-dependent phase of long-term synaptic potentiation (L-LTP) at the Schaffer collateral synapse of the hippocampus is an experimental model of the synaptic plasticity underlying long-lasting memory formation. L-LTP is typically induced by homosynaptic tetanic stimulation; but associative forms of learning are likely to require the heterosynaptic pairing of stimuli. Here we describe L-LTP elicited by such heterosynaptic pairing at the Schaffer collateral synapse in mice. We find that repeated stimulation of one pathway at low frequency (0.2 Hz), which does not by itself induce synaptic potentiation, will produce long-lasting synaptic plasticity when paired with a brief conditioning burst applied to an independent afferent pathway. The induction of heterosynaptic L-LTP is associative and critically depends on the precise time interval of pairing: simultaneous, conjunctional pairing induces L-LTP; in contrast, delayed pairing induces short-lasting early-phase LTP. Heterosynaptically induced early-phase LTP could be depotentiated by repeatedly presenting unpaired test stimuli, whereas L-LTP could not. This heterosynaptically induced L-LTP requires PKA and protein synthesis. In addition, heterosynaptically induced L-LTP is impaired in transgenic mice that express KCREB (a dominant negative inhibitor of adenosine 3′5′-cyclic monophosphate response element-binding protein-mediated transcription) in the hippocampus. These mice have previously been shown to be impaired in spatial memory but have normal L-LTP as induced by a conventional homosynaptic tetanic protocol. These data suggest that at least in some instances this L-LTP-inducing protocol may better model behaviorally relevant information storage and the
in vivo
mechanisms underlying long-lasting memories.
PLASTOCHRON1
, a timekeeper of leaf initiation in rice, encodes cytochrome P450
Authors: Miyoshi Kazumaru , Ahn Byung-Ohg , Kawakatsu Taiji , Ito Yukihiro , Itoh Jun-Ichi , Nagato Yasuo , Kurata Nori
Published: January 2004

During postembryonic development of higher plants, the shoot apical meristem produces lateral organs in a regular spacing (phyllotaxy) and a regular timing (plastochron). Molecular analysis of mutants associated with phyllotaxy and plastochron would greatly increase understanding of the developmental mechanism of plant architecture because phyllotaxy and plastochron are fundamental regulators of plant architecture.
pla1
of rice is not only a plastochron mutant showing rapid leaf initiation without affecting phyllotaxy, but also a heterochronic mutant showing ectopic shoot formation in the reproductive phase. Thus,
pla1
provides a tool for analyzing the molecular basis of temporal regulation in leaf development. In this work, we isolated the
PLA1
gene by map-based cloning. The identified
PLA1
gene encodes a cytochrome P450, CYP78A11, which potentially catalyzes substances controlling plant development.
PLA1
is expressed in developing leaf primordia, bracts of the panicle, and elongating internodes, but not in the shoot apical meristem. The expression pattern and mutant phenotype suggest that the
PLA1
gene acting in developing leaf primordia affects the timing of successive leaf initiation and the termination of vegetative growth.
Atypical protein kinase C (PKCζ/λ) is a convergent downstream target of the insulin-stimulated phosphatidylinositol 3-kinase and TC10 signaling pathways
Authors: Kanzaki Makoto , Mora Silvia , Hwang Joseph B. , Saltiel Alan R. , Pessin Jeffrey E.
Published: January 2004

Insulin stimulation of adipocytes resulted in the recruitment of atypical PKC (PKCζ/λ) to plasma membrane lipid raft microdomains. This redistribution of PKCζ/λ was prevented by Clostridium difficile toxin B and by cholesterol depletion, but was unaffected by inhibition of phosphatidylinositol (PI) 3-kinase activity. Expression of the constitutively active GTP-bound form of TC10 (TC10Q/75L), but not the inactive GDP-bound mutant (TC10/T31N), targeted PKCζ/λ to the plasma membrane through an indirect association with the Par6–Par3 protein complex. In parallel, insulin stimulation as well as TC10/Q75L resulted in the activation loop phosphorylation of PKCζ. Although PI 3-kinase activation also resulted in PKCζ/λ phosphorylation, it was not recruited to the plasma membrane. Furthermore, insulin-induced GSK-3β phosphorylation was mediated by both PI 3-kinase–PKB and the TC10–Par6–atypical PKC signaling pathways. Together, these data demonstrate that PKCζ/λ can serve as a convergent downstream target for both the PI 3-kinase and TC10 signaling pathways, but only the TC10 pathway induces a spatially restricted targeting to the plasma membrane.
FoxO3a regulates erythroid differentiation and induces BTG1, an activator of protein arginine methyl transferase 1
Authors: Bakker Walbert J. , Blázquez-Domingo Montserrat , Kolbus Andrea , Besooyen Janey , Steinlein Peter , Beug Hartmut , Coffer Paul J. , Löwenberg Bob , von Lindern Marieke , van Dijk Thamar B.
Published: January 2004

Erythropoiesis requires tight control of expansion, maturation, and survival of erythroid progenitors. Because activation of phosphatidylinositol-3-kinase (PI3K) is required for erythropoietin/stem cell factor–induced expansion of erythroid progenitors, we examined the role of the PI3K-controlled Forkhead box, class O (FoxO) subfamily of Forkhead transcription factors. FoxO3a expression and nuclear accumulation increased during erythroid differentiation, whereas untimely induction of FoxO3a activity accelerated differentiation of erythroid progenitors to erythrocytes. We identified B cell translocation gene 1 (BTG1)/antiproliferative protein 2 as a FoxO3a target gene in erythroid progenitors. Promoter studies indicated BTG1 as a direct target of FoxO3a. Expression of BTG1 in primary mouse bone marrow cells blocked the outgrowth of erythroid colonies, which required a domain of BTG1 that binds protein arginine methyl transferase 1. During erythroid differentiation, increased arginine methylation coincided with BTG1 expression. Concordantly, inhibition of methyl transferase activity blocked erythroid maturation without affecting expansion of progenitor cells. We propose FoxO3a-controlled expression of BTG1 and subsequent regulation of protein arginine methyl transferase activity as a novel mechanism controlling erythroid expansion and differentiation.
Regulation of polarized growth initiation and termination cycles by the polarisome and Cdc42 regulators
Authors: Bidlingmaier Scott , Snyder Michael
Published: January 2004

The dynamic regulation of polarized cell growth allows cells to form structures of defined size and shape. We have studied the regulation of polarized growth using mating yeast as a model. Haploid yeast cells treated with high concentration of pheromone form successive mating projections that initiate and terminate growth with regular periodicity. The mechanisms that control the frequency of growth initiation and termination under these conditions are not well understood. We found that the polarisome components Spa2, Pea2, and Bni1 and the Cdc42 regulators Cdc24 and Bem3 control the timing and frequency of projection formation. Loss of polarisome components and mutation of Cdc24 decrease the frequency of projection formation, while loss of Bem3 increases the frequency of projection formation. We found that polarisome components and the cell fusion proteins Fus1 and Fus2 are important for the termination of projection growth. Our results define the first molecular regulators that control the timing of growth initiation and termination during eukaryotic cell differentiation.
Physical and functional interaction of the active zone proteins, CAST, RIM1, and Bassoon, in neurotransmitter release
Authors: Takao-Rikitsu Etsuko , Mochida Sumiko , Inoue Eiji , Deguchi-Tawarada Maki , Inoue Marie , Ohtsuka Toshihisa , Takai Yoshimi
Published: January 2004

We have recently isolated a novel cytomatrix at the active zone (CAZ)–associated protein, CAST, and found it directly binds another CAZ protein RIM1 and indirectly binds Munc13-1 through RIM1; RIM1 and Munc13-1 directly bind to each other and are implicated in priming of synaptic vesicles. Here, we show that all the CAZ proteins thus far known form a large molecular complex in the brain, including CAST, RIM1, Munc13-1, Bassoon, and Piccolo. RIM1 and Bassoon directly bind to the COOH terminus and central region of CAST, respectively, forming a ternary complex. Piccolo, which is structurally related to Bassoon, also binds to the Bassoon-binding region of CAST. Moreover, the microinjected RIM1- or Bassoon-binding region of CAST impairs synaptic transmission in cultured superior cervical ganglion neurons. Furthermore, the CAST-binding domain of RIM1 or Bassoon also impairs synaptic transmission in the cultured neurons. These results indicate that CAST serves as a key component of the CAZ structure and is involved in neurotransmitter release by binding these CAZ proteins.
Novel regulation of mitotic exit by the Cdc42 effectors Gic1 and Gic2
Authors: Höfken Thomas , Schiebel Elmar
Published: January 2004

The guanine nucleotide exchange factor Cdc24, the GTPase Cdc42, and the Cdc42 effectors Cla4 and Ste20, two p21-activated kinases, form a signal transduction cascade that promotes mitotic exit in yeast. We performed a genetic screen to identify components of this pathway. Two related bud cortex–associated Cdc42 effectors, Gic1 and Gic2, were obtained as factors that promoted mitotic exit independently of Ste20. The mitotic exit function of Gic1 was dependent on its activation by Cdc42 and on the release of Gic1 from the bud cortex. Gic proteins became essential for mitotic exit when activation of the mitotic exit network through Cdc5 polo kinase and the bud cortex protein Lte1 was impaired. The mitotic exit defect of cdc5-10 Δlte1 Δgic1 Δgic2 cells was rescued by inactivation of the inhibiting Bfa1-Bub2 GTPase-activating protein. Moreover, Gic1 bound directly to Bub2 and prevented binding of the GTPase Tem1 to Bub2. We propose that in anaphase the Cdc42-regulated Gic proteins trigger mitotic exit by interfering with Bfa1-Bub2 GTPase-activating protein function.
Trans-SNARE interactions elicit Ca2+ efflux from the yeast vacuole lumen
Authors: Merz Alexey J. , Wickner William T.
Published: January 2004

Ca2+ transients trigger many SNARE-dependent membrane fusion events. The homotypic fusion of yeast vacuoles occurs after a release of lumenal Ca2+. Here, we show that trans-SNARE interactions promote the release of Ca2+ from the vacuole lumen. Ypt7p–GTP, the Sec1p/Munc18-protein Vps33p, and Rho GTPases, all of which function during docking, are required for Ca2+ release. Inhibitors of SNARE function prevent Ca2+ release. Recombinant Vam7p, a soluble Q-SNARE, stimulates Ca2+ release. Vacuoles lacking either of two complementary SNAREs, Vam3p or Nyv1p, fail to release Ca2+ upon tethering. Mixing these two vacuole populations together allows Vam3p and Nyv1p to interact in trans and rescues Ca2+ release. Sec17/18p promote sustained Ca2+ release by recycling SNAREs (and perhaps other limiting factors), but are not required at the release step itself. We conclude that trans-SNARE assembly events during docking promote Ca2+ release from the vacuole lumen.
Activation of Arterial Wall Dendritic Cells and Breakdown of Self-tolerance in Giant Cell Arteritis
Authors: Ma-Krupa Wei , Jeon Myung-Shin , Spoerl Silvia , Tedder Thomas F. , Goronzy Jörg J. , Weyand Cornelia M.
Published: January 2004

Giant cell arteritis (GCA) is a granulomatous and occlusive vasculitis that causes blindness, stroke, and aortic aneurysm. CD4+ T cells are selectively activated in the adventitia of affected arteries. In human GCA artery–severe combined immunodeficiency (SCID) mouse chimeras, depletion of CD83+ dendritic cells (DCs) abrogated vasculitis, suggesting that DCs are critical antigen-presenting cells in GCA. Healthy medium-size arteries possessed an indigenous population of DCs at the adventitia–media border. Adoptive T cell transfer into temporal artery–SCID mouse chimeras demonstrated that DCs in healthy arteries were functionally immature, but gained T cell stimulatory capacity after injection of lipopolysaccharide. In patients with polymyalgia rheumatica (PMR), a subclinical variant of GCA, adventitial DCs were mature and produced the chemokines CCL19 and CCL21, but vasculitic infiltrates were lacking. Human histocompatibility leukocyte antigen class II–matched healthy arteries, PMR arteries, and GCA arteries were coimplanted into SCID mice. Immature DCs in healthy arteries failed to stimulate T cells, but DCs in PMR arteries could attract, retain, and activate T cells that originated from the GCA lesions. We propose that in situ maturation of DCs in the adventitia is an early event in the pathogenesis of GCA. Activation of adventitial DCs initiates and maintains T cell responses in the artery and breaks tissue tolerance in the perivascular space.
AIRE Functions As an E3 Ubiquitin Ligase
Authors: Uchida Daisuke , Hatakeyama Shigetsugu , Matsushima Akemi , Han Hongwei , Ishido Satoshi , Hotta Hak , Kudoh Jun , Shimizu Nobuyoshi , Doucas Vassilis , Nakayama Keiichi I. , Kuroda Noriyuki , Matsumoto Mitsuru
Published: January 2004

Autoimmune regulator (AIRE) gene mutation is responsible for the development of autoimmune-polyendocrinopathy-candidiasis ectodermal dystrophy, an organ-specific autoimmune disease with monogenic autosomal recessive inheritance. AIRE is predominantly expressed in medullary epithelial cells of the thymus and is considered to play important roles in the establishment of self-tolerance. AIRE contains two plant homeodomain (PHD) domains, and the novel role of PHD as an E3 ubiquitin (Ub) ligase has just emerged. Here we show that the first PHD (PHD1) of AIRE mediates E3 ligase activity. The significance of this finding was underscored by the fact that disease-causing missense mutations in the PHD1 (C311Y and P326Q) abolished its E3 ligase activity. These results add a novel enzymatic function for AIRE and suggest an indispensable role of the Ub proteasome pathway in the establishment of self-tolerance, in which AIRE is involved.
Medullary Epithelial Cells of the Human Thymus Express a Highly Diverse Selection of Tissue-specific Genes Colocalized in Chromosomal Clusters
Authors: Gotter Jörn , Brors Benedikt , Hergenhahn Manfred , Kyewski Bruno
Published: January 2004

Promiscuous expression of tissue-specific self-antigens in the thymus imposes T cell tolerance and protects from autoimmune diseases, as shown in animal studies. Analysis of promiscuous gene expression in purified stromal cells of the human thymus at the single and global gene level documents the species conservation of this phenomenon. Medullary thymic epithelial cells overexpress a highly diverse set of genes (>400) including many tissue-specific antigens, disease-associated autoantigens, and cancer-germline genes. Although there are no apparent structural or functional commonalities among these genes and their products, they cluster along chromosomes. These findings have implications for human autoimmune diseases, immuno-therapy of tumors, and the understanding of the nature of this unorthodox regulation of gene expression.
DNA Polymerase η Is Involved in Hypermutation Occurring during Immunoglobulin Class Switch Recombination
Authors: Faili Ahmad , Aoufouchi Said , Weller Sandra , Vuillier Françoise , Stary Anne , Sarasin Alain , Reynaud Claude-Agnès , Weill Jean-Claude
Published: January 2004

Base substitutions, deletions, and duplications are observed at the immunoglobulin locus in DNA sequences involved in class switch recombination (CSR). These mutations are dependent upon activation-induced cytidine deaminase (AID) and present all the characteristics of the ones observed during V gene somatic hypermutation, implying that they could be generated by the same mutational complex. It has been proposed, based on the V gene mutation pattern of patients with the cancer-prone xeroderma pigmentosum variant (XP-V) syndrome who are deficient in DNA polymerase η (pol η), that this enzyme could be responsible for a large part of the mutations occurring on A/T bases. Here we show, by analyzing switched memory B cells from two XP-V patients, that pol η is also an A/T mutator during CSR, in both the switch region of tandem repeats as well as upstream of it, thus suggesting that the same error-prone translesional polymerases are involved, together with AID, in both processes.