bluuebunny/crossref_metadata_2025 huggingface dataset:
E-box function in a
period
gene repressed by light
Authors: Vallone Daniela , Gondi Srinivas Babu , Whitmore David , Foulkes Nicholas S.
Published: March 2004

In most organisms, light plays a key role in the synchronization of the circadian timing system with the environmental day–night cycle. Light pulses that phase-shift the circadian clock also induce the expression of
period
(
per
) genes in vertebrates. Here, we report the cloning of a zebrafish
per
gene,
zfper4
, which is remarkable in being repressed by light. We have developed an
in vivo
luciferase reporter assay for this gene in cells that contain a light-entrainable clock. High-definition bioluminescence traces have enabled us to accurately measure phase-shifting of the clock by light. We have also exploited this model to study how four E-box elements in the
zfper4
promoter regulate expression. Mutagenesis reveals that the integrity of these four E-boxes is crucial for maintaining low basal expression together with robust rhythmicity and repression by light. Importantly, in the context of a minimal heterologous promoter, the E-box elements also direct a robust circadian rhythm of expression that is significantly phase-advanced compared with the original
zfper4
promoter and lacks the light-repression property. Thus, these results reveal flexibility in the phase and light responsiveness of E-box-directed rhythmic expression, depending on the promoter context.
Morphometrics and hominoid phylogeny: Support for a chimpanzee–human clade and differentiation among great ape subspecies
Authors: Lockwood Charles A. , Kimbel William H. , Lynch John M.
Published: March 2004

Taxonomic and phylogenetic analyses of great apes and humans have identified two potential areas of conflict between molecular and morphological data: phylogenetic relationships among living species and differentiation of great ape subspecies. Here we address these problems by using morphometric data. Three-dimensional landmark data from the hominoid temporal bone effectively quantify the shape of a complex element of the skull. Phylogenetic analysis using distance-based methods corroborates the molecular consensus on African ape and human phylogeny, strongly supporting aPan–Homoclade. Phenetic differentiation of great ape subspecies is pronounced, as suggested previously by mitochondrial DNA and some morphological studies. These results show that the hominoid temporal bone contains a strong phylogenetic signal and reveal the potential for geometric morphometric analysis to shed light on phylogenetic relationships.
Pacific and Atlantic Ocean influences on multidecadal drought frequency in the United States
Authors: McCabe Gregory J. , Palecki Michael A. , Betancourt Julio L.
Published: March 2004

More than half (52%) of the spatial and temporal variance in multidecadal drought frequency over the conterminous United States is attributable to the Pacific Decadal Oscillation (PDO) and the Atlantic Multidecadal Oscillation (AMO). An additional 22% of the variance in drought frequency is related to a complex spatial pattern of positive and negative trends in drought occurrence possibly related to increasing Northern Hemisphere temperatures or some other unidirectional climate trend. Recent droughts with broad impacts over the conterminous U.S. (1996, 1999–2002) were associated with North Atlantic warming (positive AMO) and northeastern and tropical Pacific cooling (negative PDO). Much of the long-term predictability of drought frequency may reside in the multidecadal behavior of the North Atlantic Ocean. Should the current positive AMO (warm North Atlantic) conditions persist into the upcoming decade, we suggest two possible drought scenarios that resemble the continental-scale patterns of the 1930s (positive PDO) and 1950s (negative PDO) drought.
Structural basis for HLA-DQ2-mediated presentation of gluten epitopes in celiac disease
Authors: Kim Chu-Young , Quarsten Hanne , Bergseng Elin , Khosla Chaitan , Sollid Ludvig M.
Published: March 2004

Celiac disease, also known as celiac sprue, is a gluten-induced autoimmune-like disorder of the small intestine, which is strongly associated with HLA-DQ2. The structure of DQ2 complexed with an immunogenic epitope from gluten, QLQPFPQPELPY, has been determined to 2.2-Å resolution by x-ray crystallography. The glutamate at P6, which is formed by tissue transglutaminase-catalyzed deamidation, is an important anchor residue as it participates in an extensive hydrogen-bonding network involving Lys-β71 of DQ2. The gluten peptide–DQ2 complex retains critical hydrogen bonds between the MHC and the peptide backbone despite the presence of many proline residues in the peptide that are unable to participate in amide-mediated hydrogen bonds. Positioning of proline residues such that they do not interfere with backbone hydrogen bonding results in a reduction in the number of registers available for gluten peptides to bind to MHC class II molecules and presumably impairs the likelihood of establishing favorable side-chain interactions. The HLA association in celiac disease can be explained by a superior ability of DQ2 to bind the biased repertoire of proline-rich gluten peptides that have survived gastrointestinal digestion and that have been deamidated by tissue transglutaminase. Finally, surface-exposed proline residues in the proteolytically resistant ligand were replaced with functionalized analogs, thereby providing a starting point for the design of orally active agents for blocking gluten-induced toxicity.
The unique insert in myosin VI is a structural calcium–calmodulin binding site
Authors: Bahloul Amel , Chevreux Guillaume , Wells Amber L. , Martin Davy , Nolt Jocelyn , Yang Zhaohui , Chen Li-Qiong , Potier Noëlle , Van Dorsselaer Alain , Rosenfeld Steve , Houdusse Anne , Sweeney H. Lee
Published: March 2004

Myosin VI contains an inserted sequence that is unique among myosin superfamily members and has been suggested to be a determinant of the reverse directionality and unusual motility of the motor. It is thought that each head of a two-headed myosin VI molecule binds one calmodulin (CaM) by means of a single “IQ motif”. Using truncations of the myosin VI protein and electrospray ionization(ESI)-MS, we demonstrate that in fact each myosin VI head binds two CaMs. One CaM binds to a conventional IQ motif either with or without calcium and likely plays a regulatory role when calcium binds to its N-terminal lobe. The second CaM binds to a unique insertion between the converter region and IQ motif. This unusual CaM-binding site normally binds CaM with four Ca
2+
and can bind only if the C-terminal lobe of CaM is occupied by calcium. Regions of the MD outside of the insert peptide contribute to the Ca
2+
–CaM binding, as truncations that eliminate elements of the MD alter CaM binding and allow calcium dissociation. We suggest that the Ca
2+
-CaM bound to the unique insert represents a structural CaM, and not a calcium sensor or regulatory component of the motor. This structure is likely an integral part of the myosin VI “converter” region and repositions the myosin VI “lever arm” to allow reverse direction (minus-end) motility on actin.
Mitochondrial localization of estrogen receptor β
Authors: Yang Shao-Hua , Liu Ran , Perez Evelyn J. , Wen Yi , Stevens Stanley M. , Valencia Thomas , Brun-Zinkernagel Anne-Marie , Prokai Laszlo , Will Yvonne , Dykens James , Koulen Peter , Simpkins James W.
Published: March 2004

Estrogen receptors (ERs) are believed to be ligand-activated transcription factors belonging to the nuclear receptor superfamily, which on ligand binding translocate into the nucleus and activate gene transcription. To date, two ERs have been identified: ERα and ERβ. ERα plays major role in the estrogen-mediated genomic actions in both reproductive and nonreproductive tissue, whereas the function of ERβ is still unclear. In this study, we used immunocytochemistry, immunoblotting, and proteomics to demonstrate that ERβ localizes to the mitochondria. In immunocytochemistry studies, ERβ was detected with two ERβ antibodies and found to colocalize almost exclusively with a mitochondrial marker in rat primary neuron, primary cardiomyocyte, and a murine hippocampal cell line. The colocalization of ERβ and mitochondrial markers was identified by both fluorescence and confocal microscopy. No translocation of ERβ into the nucleus on 17β-estradiol treatment was seen by using immunocytochemistry. Immunoblotting of purified human heart mitochondria showed an intense signal of ERβ, whereas no signals for nuclear and other organelle markers were found. Finally, purified human heart mitochondrial proteins were separated by SDS/PAGE. The 50,000–65,000
M
r
band was digested with trypsin and subjected to matrix-assisted laser desorption/ionization mass spectrometric analysis, which revealed seven tryptic fragments that matched with those of ERβ. In summary, this study demonstrated that ERβ is localized to mitochondria, suggesting a role for mitochondrial ERβ in estrogen effects on this important organelle.
Predisposition to abacavir hypersensitivity conferred byHLA-B*5701and a haplotypicHsp70-Homvariant
Authors: Martin Annalise M. , Nolan David , Gaudieri Silvana , Almeida Coral Ann , Nolan Richard , James Ian , Carvalho Filipa , Phillips Elizabeth , Christiansen Frank T. , Purcell Anthony W. , McCluskey James , Mallal Simon
Published: March 2004

Susceptibility to a clinically significant drug hypersensitivity syndrome associated with abacavir use seems to have a strong genetic component. We have previously shown that the presence ofHLA-B*5701strongly predicts abacavir hypersensitivity and have identified a potential susceptibility locus within a 300-kb region between theMEGT1andC4A6loci in the central MHC. We now report the results of fine recombinant genetic mapping in an expanded patient population of 248 consecutive, fully ascertained, abacavir-exposed individuals in the Western Australian HIV Cohort Study, in which 18 cases of definite abacavir hypersensitivity (7.3%) and 230 tolerant controls were identified. Haplotype mapping within patients with allelic markers of the 57.1 ancestral haplotype suggests a susceptibility locus within the 14-kbHsp70gene cluster.HLA-B*5701was present in 94.4% of hypersensitive cases compared with 1.7% of controls (odds ratio, 960;P< 0.00001). A haplotypic nonsynonymous polymorphism ofHsp70-Hom(HspA1L, resulting from the substitution of residue M493T in the peptide-binding subunit) was found in combination withHLA-B*5701in 94.4% of hypersensitive cases and 0.4% of controls (odds ratio, 3,893;P< 0.00001). Individuals with abacavir hypersensitivity demonstrated increased monocyte tumor necrosis factor expression in response toex vivoabacavir stimulation, which was abrogated with CD8+T cell depletion. These data indicate that the concurrence ofHLA-B*5701andHsp70-Hom M493Talleles is necessary for the development of abacavir hypersensitivity, which is likely to be mediated by anHLA-B*5701-restricted immune response to abacavir.
Catalytic enantioselective intermolecular cycloadditions of 2-diazo-3,6-diketoester-derived carbonyl ylides with alkene dipolarophiles
Authors: Hodgson David M. , Brückl Tobias , Glen Rebecca , Labande Agnès H. , Selden Deborah A. , Dossetter Alexander G. , Redgrave Alison J.
Published: March 2004

Catalyzed cascade reactions that generate molecular complexity rapidly and in an enantioselective manner are attractive methods for asymmetric synthesis. In the present article, chiral rhodium catalysts are shown to effect such a transformation by using a range of 2-diazo-3,6-diketoesters with bicyclo[2.2.1]alkenes and styrenes as reaction partners. The reactions are likely to proceed by formation of a catalyst-complexed carbonyl ylide from the diazo compound, followed by intermolecular cycloaddition with the alkene dipolarophile. It was possible to obtain high levels of asymmetric induction [up to 89% enantiomeric excess (ee) and 92% ee for the two chiral catalysts investigated]. Enantioselectivity is not highly sensitive to substituent variation at the ketone that forms the ylide; however, branching does improve ee. Observations of dipolarophile-dependent enantiofacial selectivity in the cycloadditions indicate that the dipolarophile can be intimately involved in the enantiodiscrimination process.
Responses of human visual cortex to uniform surfaces
Authors: Haynes John-Dylan , Lotto R. Beau , Rees Geraint
Published: March 2004

Surface perception is fundamental to human vision, yet most studies of visual cortex have focused on the processing of borders. We therefore investigated the responses of human visual cortex to parametric changes in the luminance of uniform surfaces by using functional MRI. Early visual areas V1 and V2/V3 showed strong and reliable increases in signal for both increments and decrements in surface luminance. Responses were significantly larger for decrements than for increments, which was fully accounted for by differences in retinal illumination arising from asymmetric pupil dynamics. Responses to both sustained and transient changes of illumination were transient. Signals in early visual cortex scaled linearly with the magnitude of change in retinal illumination, as did subjects' subjective ratings of the perceived brightness of the stimuli. Our findings show that early visual cortex responds strongly to surfaces and that perception of surface brightness is compatible with brain responses at the earliest cortical stages of processing.
A highly conserved candidate chemoreceptor expressed in both olfactory and gustatory tissues in the malaria vector
Anopheles gambiae
Authors: Pitts R. Jason , Fox A. Nicole , Zwiebel Laurence J.
Published: March 2004

Anopheles gambiae
is a highly anthropophilic mosquito responsible for the majority of malaria transmission in Africa. The biting and host preference behavior of this disease vector is largely influenced by its sense of smell, which is presumably facilitated by G protein-coupled receptor signaling [Takken, W. & Knols, B. (1999)
Annu. Rev. Entomol.
44, 131-157]. Because of the importance of host preference to the mosquitoes' ability to transmit disease, we have initiated studies intended to elucidate the molecular mechanisms underlying olfaction in
An. gambiae
. In the course of these studies, we have identified a number of genes potentially involved in signal transduction, including a family of candidate odorant receptors. One of these receptors, encoded by
GPRor7
(hereafter referred to as
AgOr7
), is remarkably similar to an odorant receptor that is expressed broadly in olfactory tissues and has been identified in
Drosophila melanogaster
and other insects [Krieger, J., Klink, O., Mohl, C., Raming, K. & Breer, H. (2003)
J. Comp. Physiol. A
189, 519-526; Vosshall, L. B., Amrein, H., Morozov, P. S., Rzhetsky, A. & Axel, R. (1999)
Cell
96, 725-736]. We have observed
AgOr7
expression in olfactory and gustatory tissues in adult
An. gambiae
and during several stages of the mosquitoes' development. Within the female adult peripheral chemosensory system, antiserum against the AgOR7 polypeptide labels most sensilla of the antenna and maxillary palp as well as a subset of proboscis sensilla. Furthermore, AgOR7 antiserum labeling is observed within the larval antenna and maxillary palpus. These results are consistent with a role for
AgOr7
in both olfaction and gustation in
An. gambiae
and raise the possibility that
AgOr7
orthologs may also be of general importance to both modalities of chemosensation in other insects.
Metagenes and molecular pattern discovery using matrix factorization
Authors: Brunet Jean-Philippe , Tamayo Pablo , Golub Todd R. , Mesirov Jill P.
Published: March 2004

We describe here the use of nonnegative matrix factorization (NMF), an algorithm based on decomposition by parts that can reduce the dimension of expression data from thousands of genes to a handful of metagenes. Coupled with a model selection mechanism, adapted to work for any stochastic clustering algorithm, NMF is an efficient method for identification of distinct molecular patterns and provides a powerful method for class discovery. We demonstrate the ability of NMF to recover meaningful biological information from cancer-related microarray data. NMF appears to have advantages over other methods such as hierarchical clustering or self-organizing maps. We found it less sensitive toa prioriselection of genes or initial conditions and able to detect alternative or context-dependent patterns of gene expression in complex biological systems. This ability, similar to semantic polysemy in text, provides a general method for robust molecular pattern discovery.
The Nash equilibrium: A perspective
Authors: Holt Charles A. , Roth Alvin E.
Published: March 2004

In 1950, John Nash contributed a remarkable one-page PNAS article that defined and characterized a notion of equilibrium for
n-
person games. This notion, now called the “Nash equilibrium,” has been widely applied and adapted in economics and other behavioral sciences. Indeed, game theory, with the Nash equilibrium as its centerpiece, is becoming the most prominent unifying theory of social science. In this perspective, we summarize the historical context and subsequent impact of Nash's contribution.
FamClash: A method for ranking the activity of engineered enzymes
Authors: Saraf Manish C. , Horswill Alexander R. , Benkovic Stephen J. , Maranas Costas D.
Published: March 2004

This article introduces the computational procedure FamClash for analyzing incompatibilities in engineered protein hybrids by using protein family sequence data. All pairs of residue positions in the sequence alignment that conserve the property triplet of charge, volume, and hydrophobicity are first identified, and significant deviations are denoted as residue–residue clashes. This approach moves beyond earlier efforts aimed at solely classifying hybrids as functional or nonfunctional by correlating the rank ordering of these hybrids based on their activity levels. Experimental testing of this approach was performed in parallel to assess the predictive ability of FamClash. As a model system, single-crossover ITCHY (incremental truncation for the creation of hybrid enzymes) libraries were prepared from theEscherichia coliandBacillus subtilisdihydrofolate reductases, and the activities of functional hybrids were determined. Comparisons of the predicted clash map as a function of crossover position revealed good agreement with activity data, reproducing the observed V shape and matching the location of a local peak in activity.
A Fenton reaction at the endoplasmic reticulum is involved in the redox control of hypoxia-inducible gene expression
Authors: Liu Qing , Berchner-Pfannschmidt Utta , Möller Ulrike , Brecht Martina , Wotzlaw Christoph , Acker Helmut , Jungermann Kurt , Kietzmann Thomas
Published: March 2004

It has been proposed that hydroxyl radicals (·OH) generated in a perinuclear iron-dependent Fenton reaction are involved in O
2
-dependent gene expression. Thus, it was the aim of this study to localize the cellular compartment in which the Fenton reaction takes place and to determine whether scavenging of ·OH can modulate hypoxia-inducible factor 1 (HIF-1)-dependent gene expression. The Fenton reaction was localized by using the nonfluorescent dihydrorhodamine (DHR) 123 that is irreversibly oxidized to fluorescent rhodamine 123 while scavenging ·OH together with gene constructs allowing fluorescent labeling of mitochondria, endoplasmic reticulum (ER), Golgi apparatus, peroxisomes, or lysosomes. A 3D two-photon confocal laser scanning microscopy showed ·OH generation in distinct hot spots of perinuclear ER pockets. This ER-based Fenton reaction was strictly pO
2
-dependent. Further colocalization experiments showed that the O
2
-sensitive transcription factor HIF-1α was present at the ER under normoxia, whereas HIF-1α was present only in the nucleus under hypoxia. Inhibition of the Fenton reaction by the ·OH scavenger DHR attenuated HIF-prolyl hydroxylase activity and interaction with von Hippel–Lindau protein, leading to enhanced HIF-1α levels, HIF-1α transactivation, and activated expression of the HIF-1 target genes plasminogen activator inhibitor 1 and heme oxygenase 1. Further, ·OH scavenging appeared to enhance redox factor 1 (Ref-1) binding and, thus, recruitment of p300 to the transactivation domain C because mutation of the Ref-1 binding site cysteine 800 abolished DHR-induced transactivation. Thus, the localized Fenton reaction appears to impact the expression of hypoxia-regulated genes by means of HIF-1α stabilization and coactivator recruitment.
From paragraph to graph: Latent semantic analysis for information visualization
Authors: Landauer Thomas K. , Laham Darrell , Derr Marcia
Published: March 2004

Most techniques for relating textual information rely on intellectually created links such as author-chosen keywords and titles, authority indexing terms, or bibliographic citations. Similarity of the semantic content of whole documents, rather than just titles, abstracts, or overlap of keywords, offers an attractive alternative. Latent semantic analysis provides an effective dimension reduction method for the purpose that reflects synonymy and the sense of arbitrary word combinations. However, latent semantic analysis correlations with human text-to-text similarity judgments are often empirically highest at ≈300 dimensions. Thus, two- or three-dimensional visualizations are severely limited in what they can show, and the first and/or second automatically discovered principal component, or any three such for that matter, rarely capture all of the relations that might be of interest. It is our conjecture that linguistic meaning is intrinsically and irreducibly very high dimensional. Thus, some method to explore a high dimensional similarity space is needed. But the 2.7 × 107projections and infinite rotations of, for example, a 300-dimensional pattern are impossible to examine. We suggest, however, that the use of a high dimensional dynamic viewer with an effective projection pursuit routine and user control, coupled with the exquisite abilities of the human visual system to extract information about objects and from moving patterns, can often succeed in discovering multiple revealing views that are missed by current computational algorithms. We show some examples of the use of latent semantic analysis to support such visualizations and offer views on future needs.
Long-term human response to uncertain environmental conditions in the Andes
Authors: Dillehay Tom D. , Kolata Alan L.
Published: March 2004

Human interaction with the physical environment has increasingly transformed Earth-system processes. Reciprocally, climate anomalies and other processes of environmental change of natural and anthropogenic origin have been affecting, and often disrupting, societies throughout history. Transient impact events, despite their brevity, can have significant long-term impact on society, particularly if they occur in the context of ongoing, protracted environmental change. Major climate events can affect human activities in critical conjunctures that shape particular trajectories of social development. Here we report variable human responses to major environmental events in the Andes with a particular emphasis on the period from anno Domini 500–1500 on the desert north coast of Perú. We show that preindustrial agrarian societies implemented distinct forms of anticipatory response to environmental change and uncertainty. We conclude that innovations in production strategies and agricultural infrastructures in these indigenous societies reflect differential social response to both transient (El Niño–Southern Oscillation events) and protracted (desertification) environmental change.
A nascent polypeptide domain that can regulate translation elongation
Authors: Fang Peng , Spevak Christina C. , Wu Cheng , Sachs Matthew S.
Published: March 2004

The evolutionarily conserved fungal arginine attenuator peptide (AAP), as a nascent peptide, stalls the translating ribosome in response to the presence of a high concentration of the amino acid arginine. Here we examine whether the AAP maintains regulatory function in fungal, plant, and animal cell-free translation systems when placed as a domain near the N terminus or internally within a large polypeptide. Pulse–chase analyses of the radiolabeled polypeptides synthesized in these systems indicated that wild-type AAP functions at either position to stall polypeptide synthesis in response to arginine. Toeprint analyses performed to map the positions of stalled ribosomes on transcripts introduced into the fungal system revealed that ribosome stalling required translation of the AAP coding sequence. The positions of the stalled ribosomes were consistent with the sizes of the radiolabeled polypeptide intermediates. These findings demonstrate that an internal polypeptide domain in a nascent chain can regulate eukaryotic translational elongation in response to a small molecule. Apparently the peptide-sensing features are conserved in fungal, plant, and animal ribosomes. These data provide precedents for translational strategies that would allow domains within nascent polypeptide chains to modulate gene expression.
Expression profiling with arrays of randomly disrupted genes in mouse embryonic stem cells leads to
in vivo
functional analysis
Authors: Matsuda Eishou , Shigeoka Toshiaki , Iida Ryuji , Yamanaka Shinya , Kawaichi Masashi , Ishida Yasumasa
Published: March 2004

DNA arrays are capable of profiling the expression patterns of many genes in a single experiment. After finding a gene of interest in a DNA array, however, labor-intensive gene-targeting experiments sometimes must be performed for the
in vivo
analysis of the gene function. With random gene trapping, on the other hand, it is relatively easy to disrupt and retrieve hundreds of genes/gene candidates in mouse embryonic stem (ES) cells, but one could overlook potentially important gene-disruption events if only the nucleotide sequences and not the expression patterns of the trapped DNA segments are analyzed. To combine the benefits of the above two experimental systems, we first created ≈900 genetrapped mouse ES cell clones and then constructed arrays of cDNAs derived from the disrupted genes. By using these arrays, we identified a novel gene predominantly expressed in the mouse brain, and the corresponding ES cell clone was used to produced mice homozygous for the disrupted allele of the gene. Detailed analysis of the knockout mice revealed that the gene trap vector completely abolished gene expression downstream of its integration site. Therefore, identification of a gene or novel gene candidate with an interesting expression pattern by using this type of DNA array immediately allows the production of knockout mice from an ES cell clone with a disrupted allele of the sequence of interest.
Ataxin 1, a
SCA1
neurodegenerative disorder protein, is functionally linked to the silencing mediator of retinoid and thyroid hormone receptors
Authors: Tsai Chih-Cheng , Kao Hung-Ying , Mitzutani Akifumi , Banayo Ester , Rajan Harini , McKeown Michael , Evans Ronald M.
Published: March 2004

Ataxin 1 (Atx1) is a foci-forming polyglutamine protein of unknown function, whose mutant form causes type 1 spinocerebellar ataxia in humans and exerts neurotoxicity in transgenic mouse and fly expressing mutant Atx1. In this study, we demonstrate that Atx1 interacts with the transcriptional corepressor SMRT (silencing mediator of retinoid and thyroid hormone receptors) and with histone deacetylase 3. Atx1 binds chromosomes and mediates transcriptional repression when tethered to DNA. Interaction with SMRT-related factors is a conserved feature of Atx1, because Atx1 also binds SMRTER, a
Drosophila
cognate of SMRT. Significantly, mutant Atx1 forms aggregates in
Drosophila
, and such mutant Atx1-mediated aggregates sequester SMRTER. Consistently, the neurodegenerative eye phenotype caused by mutant Atx1 is enhanced by a
Smrter
mutation and, conversely, is suppressed by a chromosomal duplication that contains the wild type
Smrter
gene. Together, our results suggest that Atx1 is a transcriptional factor whose mutant form exerts its deleterious effects in part by perturbing corepressor-dependent transcriptional pathways.
The transforming activity of Wnt effectors correlates with their ability to induce the accumulation of mammary progenitor cells
Authors: Liu Bob Y. , McDermott Sean P. , Khwaja Shariq S. , Alexander Caroline M.
Published: March 2004

Ectopic activation of the Wnt signaling pathway is highly oncogenic for many human tissues. Here, we show that ectopic Wnt signaling increases the effective stem cell activity in mouse mammary glands
in vivo
. Furthermore, Wnt effectors induce the accumulation of mouse mammary epithelial progenitors (assayed by Hoechst dye exclusion, a surrogate stem cell marker, side population cells) both
in vivo
and
in vitro
. The longevity of stem cells makes them good candidate tumor precursors, and we propose that Wnt-induced progenitor amplification is likely to be key to tumor initiation. In support of this notion, mammary glands from a tumor-resistant strain of mice (carrying a null mutation in syndecan-1) contain fewer side population cells. When this strain is crossed to mice that overexpress effectors of the β-catenin/T cell factor Wnt pathway, the amplification of progenitors is reduced, together with all subsequent events of tumor development. We propose that the growth dynamic of the stem cell fraction is a major determinant of tumor susceptibility.
Waves and aggregation patterns in myxobacteria
Authors: Igoshin Oleg A. , Welch Roy , Kaiser Dale , Oster George
Published: March 2004

Under starvation conditions, a population of myxobacteria aggregates to build a fruiting body whose shape is species-specific and within which the cells sporulate. Early in this process, cells often pass through a “ripple phase” characterized by traveling linear, concentric, and spiral waves. These waves are different from the waves observed during slime mold aggregation that depend on diffusible morphogens, because myxobacteria communicate by direct contact. The difference is most dramatic when waves collide: rather than annihilating one another, myxobacterial waves appear to pass through one another unchanged. Under certain conditions, the spacing and location of the nascent fruiting bodies is determined by the wavelength and pattern of the waves. Later in fruiting body development, waves are replaced by streams of cells that circulate around small initial aggregates enlarging and rounding them. Still later, pairs of motile aggregates coalesce to form larger aggregates that develop into fruiting bodies. Here we present a mathematical model that quantitatively explains these wave and aggregation phenomena.
Capillarity-driven assembly of two-dimensional cellular carbon nanotube foams
Authors: Chakrapani Nirupama , Wei Bingqing , Carrillo Alvaro , Ajayan Pulickel M. , Kane Ravi S.
Published: March 2004

Capillary forces arising during the evaporation of liquids from dense carbon nanotube arrays are used to reassemble the nanotubes into two-dimensional contiguous cellular foams. The stable nanotube foams can be elastically deformed, transferred to other substrates, or floated out to produce free-standing macroscopic fabrics. The lightweight cellular foams made of condensed nanotubes could have applications as shock-absorbent structural reinforcements and elastic membranes. The ability to control the length scale, orientation, and shape of the cellular structures and the simplicity of the assembly process make this a particularly attractive system for studying pattern formation in ordered media.
The neural F-box protein NFB42 mediates the nuclear export of the herpes simplex virus type 1 replication initiator protein (UL9 protein) after viral infection
Authors: Eom Chi-Yong , Heo Won Do , Craske Madeleine L. , Meyer Tobias , Lehman I. Robert
Published: March 2004

The neural F-box 42-kDa protein (NFB42) is a component of the SCF
NFB42
E3 ubiquitin ligase that is expressed in all major areas of the brain; it is not detected in nonneuronal tissues. We previously identified NFB42 as a binding partner for the herpes simplex virus 1 (HSV-1) UL9 protein, the viral replication-initiator, and showed that coexpression of NFB42 and UL9 in human embryonic kidney (293T) cells led to a significant decrease in the level of UL9 protein. We have now found that HSV-1 infection promotes the shuttling of NFB42 between the cytosol and the nucleus in both 293T cells and primary hippocampal neurons, permitting NFB42 to bind to the phosphorylated UL9 protein, which is localized in the nucleus. This interaction mediates the export of the UL9 protein from the nucleus to the cytosol, leading to its ubiquitination and degradation via the 26S proteasome. Because the intranuclear localization of the UL9 protein, along with other viral and cellular factors, is an essential step in viral DNA replication, degradation of the UL9 protein in neurons by means of nuclear export through its specific interaction with NFB42 may prevent active replication and promote neuronal latency of HSV-1.
Effect of hydrogen-switchable mirrors on the Casimir force
Authors: Iannuzzi Davide , Lisanti Mariangela , Capasso Federico
Published: March 2004

We present systematic measurements of the Casimir force between a gold-coated plate and a sphere coated with a hydrogen-switchable mirror. Hydrogen-switchable mirrors are shiny metals that can become transparent upon hydrogenation. Despite such a dramatic change of the optical properties of the sphere, we did not observe any significant decrease of the Casimir force after filling the experimental apparatus with hydrogen. This counterintuitive result can be explained by the Lifshitz theory that describes the Casimir attraction between metallic and dielectric materials.
Isoprenoid biosynthesis in the diatoms
Rhizosolenia setigera
(Brightwell) and
Haslea ostrearia
(Simonsen)
Authors: Massé Guillaume , Belt Simon T. , Rowland Steven J. , Rohmer Michel
Published: March 2004

Isoprenoid biosynthesis in the widespread diatomaceous algae,
Rhizosolenia setigera
(Brightwell) and
Haslea ostrearia
(Simonsen), results not only in the production of diterpenoids, triterpenoids, and sterols but, unusually for diatoms, also in the production of sesterterpenoids. By using
13
C and
2
H isotopic labeling techniques followed by NMR and mass spectrometry, specific inhibition of mevalonate (MVA) and methylerythritol (MEP) pathways, and comparison with the natural
13
C/
12
C isotope ratios of the lipids, the different biosynthetic pathways of the sesterterpenes and other isoprenoids have now been determined. Surprisingly, whereas the sesterterpenes (Δ
7(20)
-haslenes) in
R. setigera
were made by the MVA pathway, as were the related triterpenoid rhizenes and desmosterol, in
H. ostrearia
the structurally similar Δ
6(17)
-haslenes and the major sterol, 24-ethylcholest-5-en-3β-ol, were instead biosynthesized by the MEP route. Phytol was biosynthesized in both diatoms by the MEP route. Subfractionation of
R. setigera
cells revealed that although phytol was located in the chloroplasts, the haslenes, rhizenes, and sterols were present in the cytoplasm. The observations described here for
R. setigera
and
H. ostrearia
show that terpenoid biosynthesis in diatoms is species-dependent and cannot simply be grouped according to structural type. Triterpenes appear to be made by the MVA route as in higher plants, whereas sesterterpenes and sterols can be made by either the MVA or MEP routes. In neither organism were the isoprenoids biosynthesized by leucine metabolism. Sesterterpene and triterpene biosynthesis in diatoms has not been investigated previously.
A nuclear translation-like factor eIF4AIII is recruited to the mRNA during splicing and functions in nonsense-mediated decay
Authors: Ferraiuolo Maria A. , Lee Chung-Sheng , Ler Lian Wee , Hsu Jeanne L. , Costa-Mattioli Mauro , Luo Ming-Juan , Reed Robin , Sonenberg Nahum
Published: March 2004

In eukaryotes, a surveillance mechanism known as nonsense-mediated decay (NMD) degrades the mRNA when a premature-termination codon (PTC) is present. NMD requires translation to read the frame of the mRNA and detect the PTC. During pre-mRNA splicing, the exon–exon junction complex (EJC) is recruited to a region 20–24 nt upstream of the exon junction on the mature mRNA. The presence of a PTC upstream from the EJC elicits NMD. Eukaryotic initiation factor 4A (eIF4A) III is a nuclear protein that interacts physically or functionally with translation initiation factors eIF4G and eIF4B, respectively, and shares strikingly high identity with the initiation factors eIF4AI/II. Here we show that siRNA against eIF4AIII, but not against eIF4AI/II, inhibits NMD. Moreover, eIF4AIII, but not eIF4AI, is specifically recruited to the EJC during splicing. The observations that eIF4AIII is loaded onto the mRNA during splicing in the nucleus, has properties related to a translation initiation factor, and functions in NMD raises the possibility that eIF4AIII substitutes for eIF4AI/II during NMD.
Structural basis of androgen receptor binding to selective androgen response elements
Authors: Shaffer Paul L. , Jivan Arif , Dollins D. Eric , Claessens Frank , Gewirth Daniel T.
Published: March 2004

Steroid receptors bind as dimers to a degenerate set of response elements containing inverted repeats of a hexameric half-site separated by 3 bp of spacer (IR3). Naturally occurring selective androgen response elements have recently been identified that resemble direct repeats of the hexameric half-site (ADR3). The 3D crystal structure of the androgen receptor (AR) DNA-binding domain bound to a selective ADR3 reveals an unexpected head-to-head arrangement of the two protomers rather than the expected head-to-tail arrangement seen in nuclear receptors bound to response elements of similar geometry. Compared with the glucocorticoid receptor, the DNA-binding domain dimer interface of the AR has additional interactions that stabilize the AR dimer and increase the affinity for nonconsensus response elements. This increased interfacial stability compared with the other steroid receptors may account for the selective binding of AR to ADR3 response elements.
Conformational control of Bax localization and apoptotic activity by Pro168
Authors: Schinzel Anna , Kaufmann Thomas , Schuler Martin , Martinalbo Jorge , Grubb David , Borner Christoph
Published: March 2004

In healthy cells, Bax resides inactive in the cytosol because its COOH-terminal transmembrane region (TMB) is tucked into a hydrophobic pocket. During apoptosis, Bax undergoes a conformational change involving NH2-terminal exposure and translocates to mitochondria to release apoptogenic factors. How this process is regulated remains unknown. We show that the TMB of Bax is both necessary and sufficient for mitochondrial targeting. However, its availability for targeting depends on Pro168 located within the preceding loop region. Pro168 mutants of Bax lack apoptotic activity, cannot rescue the apoptosis-resistant phenotype of Bax/Bak double knockout cells, and are retained in the cytosol even in response to apoptotic stimuli. Moreover, the mutants have their NH2 termini exposed. We propose that Pro168 links the NH2 and the COOH terminus of Bax and is required for COOH-terminal release and mitochondrial targeting once this link is broken.
VCIP135 acts as a deubiquitinating enzyme during p97–p47-mediated reassembly of mitotic Golgi fragments
Authors: Wang Yanzhuang , Satoh Ayano , Warren Graham , Meyer Hemmo H.
Published: March 2004

The AAA-ATPase p97/Cdc48 functions in different cellular pathways using distinct sets of adapters and other cofactors. Together with its adaptor Ufd1–Npl4, it extracts ubiquitylated substrates from the membrane for subsequent delivery to the proteasome during ER-associated degradation. Together with its adaptor p47, on the other hand, it regulates several membrane fusion events, including reassembly of Golgi cisternae after mitosis. The finding of a ubiquitin-binding domain in p47 raises the question as to whether the ubiquitin–proteasome system is also involved in membrane fusion events. Here, we show that p97–p47-mediated reassembly of Golgi cisternae requires ubiquitin, but is not dependent on proteasome-mediated proteolysis. Instead, it requires the deubiquitinating activity of one of its cofactors, VCIP135, which reverses a ubiquitylation event that occurs during mitotic disassembly. Together, these data reveal a cycle of ubiquitylation and deubiquitination that regulates Golgi membrane dynamics during mitosis. Furthermore, they represent the first evidence for a proteasome-independent function of p97/Cdc48.
Markers for Detergent-resistant Lipid Rafts Occupy Distinct and Dynamic Domains in Native Membranes
Authors: Wilson Bridget S. , Steinberg Stanly L. , Liederman Karin , Pfeiffer Janet R. , Surviladze Zurab , Zhang Jun , Samelson Lawrence E. , Yang Li-hong , Kotula Paul G. , Oliver Janet M.
Published: March 2004

Lipid rafts isolated by detergent extraction and sucrose gradient fractionation from mast cells are enriched for the glycosylphosphatidylinositol-linked protein Thy-1, the ganglioside GM1, palmitoylated LAT, and cross-linked IgE receptors, FcϵRI. This study addresses the relationship of fractionation data to the organization of raft markers in native membranes. Immunogold labeling and electron microscopy shows there is little or no colocalization of the raft markers Thy-1, GM1, and LAT with each other or with FcϵRI on native membrane sheets prepared from unstimulated cells. External cross-linking of Thy-1 promotes coclustering of Thy-1 with LAT, but not with GM1. Thy-1 and LAT clusters occur on membrane regions without distinctive features. In contrast, external cross-linking of FcϵRI and GM1 causes their redistribution to electron-dense membrane patches independently of each other and of Thy-1. The distinctive patches that accumulate cross-linked FcϵRI and GM1 also accumulate osmium, a stain for unsaturated lipids, and are sites for coated vesicle budding. Electron microscopy reveals a more complex and dynamic topographical organization of membrane microdomains than is predicted by biochemical analysis of detergent-resistant membranes.
The Integrin β1 Subunit Transmembrane Domain Regulates Phosphatidylinositol 3-Kinase-dependent Tyrosine Phosphorylation of Crk-associated Substrate
Authors: Armulik Annika , Velling Teet , Johansson Staffan
Published: March 2004

Our previous studies on the transmembrane domain of human integrin subunits have shown that a conserved basic amino acid in both subunits of integrin heterodimers is positioned in the plasma membrane in the absence of interacting proteins. To investigate the possible functional role of the lipid-embedded lysine in the mouse integrin β1 subunit, this amino acid was replaced with leucine, and the mutated β1 subunit (β1AK756L) was stably expressed in β1-deficient GD25 cells. The extracellular domain of β1AK756Lintegrins possesses a competent conformation for ligand binding as determined by the ability to mediate cell adhesion, and by the presence of the monoclonal antibody 9EG7 epitope. However, the spreading of GD25-β1AK756Lcells on fibronectin and laminin-1 was impaired, and the rate of migration of GD25-β1AK756Lcells on fibronectin was reduced compared with GD25-β1A cells. Phosphorylation of tyrosines in focal adhesion kinase (FAK) and the Y416 in c-Src in response to β1AK756L-mediated adhesion was similar to that induced by wild-type β1. The tyrosine phosphorylation level of paxillin, a downstream target of FAK/Src, was unaffected by the β1 mutation, whereas tyrosine phosphorylation of CAS was strongly reduced. The results demonstrate that CAS is a target for phosphorylation both by FAK-dependent and -independent pathways after integrin ligation. The latter pathway was inhibited by wortmannin and LY294002, implicating that it required an active phosphatidylinositol 3-kinase. Furthermore, the K756L mutation in the β1 subunit was found to interfere with β1-induced activation of Akt. The results from this study identify phosphatidylinositol 3-kinase as an early component of a FAK-independent integrin signaling pathway triggered by the membrane proximal part of the β1 subunit.
RalA Activation at Nascent Lamellipodia of Epidermal Growth Factor-stimulated Cos7 Cells and Migrating Madin-Darby Canine Kidney Cells
Authors: Takaya Akiyuki , Ohba Yusuke , Kurokawa Kazuo , Matsuda Michiyuki
Published: March 2004

RalA, a member of the Ras-family GTPases, regulates various cellular functions such as filopodia formation, endocytosis, and exocytosis. On epidermal growth factor (EGF) stimulation, activated Ras recruits guanine nucleotide exchange factors (GEFs) for RalA, followed by RalA activation. By using fluorescence resonance energy transfer-based probes for RalA activity, we found that the EGF-induced RalA activation in Cos7 cells was restricted at the EGF-induced nascent lamellipodia, whereas under a similar condition both Ras activation and Ras-dependent translocation of Ral GEFs occurred more diffusely at the plasma membrane. This EGF-induced RalA activation was not observed when lamellipodial protrusion was suppressed by a dominant negative mutant of Rac1, a GTPase-activating protein for Cdc42, inhibitors of phosphatidylinositol 3-kinase, or inhibitors of actin polymerization. On the other hand, EGF-induced lamellipodial protrusion was inhibited by microinjection of the RalA-binding domains of RalBP1 and Sec5. Furthermore, we found that RalA activity was high at the lamellipodia of migrating Madin-Darby canine kidney cells and that the migration of Madin-Darby canine kidney cells was perturbed by the microinjection of RalBP1–RalA-binding domain. Thus, RalA activation is required for the induction of lamellipodia, and conversely, lamellipodial protrusion seems to be required for the RalA activation, suggesting the presence of a positive feedback loop between RalA activation and lamellipodial protrusion. Our observation also demonstrates that the spatial regulation of RalA is conducted by a mechanism distinct from the temporal regulation conducted by Ras-dependent plasma membrane recruitment of Ral guanine nucleotide exchange factors.
Tissue-specific Expression and Dynamic Organization of SR Splicing Factors inArabidopsis
Authors: Fang Yuda , Hearn Stephen , Spector David L.
Published: March 2004

The organization of the pre-mRNA splicing machinery has been extensively studied in mammalian and yeast cells and far less is known in living plant cells and different cell types of an intact organism. Here, we report on the expression, organization, and dynamics of pre-mRNA splicing factors (SR33, SR1/atSRp34, and atSRp30) under control of their endogenous promoters in Arabidopsis. Distinct tissue-specific expression patterns were observed, and differences in the distribution of these proteins within nuclei of different cell types were identified. These factors localized in a cell type-dependent speckled pattern as well as being diffusely distributed throughout the nucleoplasm. Electron microscopic analysis has revealed that these speckles correspond to interchromatin granule clusters. Time-lapse microscopy revealed that speckles move within a constrained nuclear space, and their organization is altered during the cell cycle. Fluorescence recovery after photobleaching analysis revealed a rapid exchange rate of splicing factors in nuclear speckles. The dynamic organization of plant speckles is closely related to the transcriptional activity of the cells. The organization and dynamic behavior of speckles in Arabidopsis cell nuclei provides significant insight into understanding the functional compartmentalization of the nucleus and its relationship to chromatin organization within various cell types of a single organism.