bluuebunny/crossref_metadata_2025 huggingface dataset:
The
Arabidopsis
outward K
+
channel
GORK
is involved in regulation of stomatal movements and plant transpiration
Authors: Hosy Eric , Vavasseur Alain , Mouline Karine , Dreyer Ingo , Gaymard Frédéric , Porée Fabien , Boucherez Jossia , Lebaudy Anne , Bouchez David , Véry Anne-Aliénor , Simonneau Thierry , Thibaud Jean-Baptiste , Sentenac Hervé
Published: April 2003

Microscopic pores present in the epidermis of plant aerial organs, called stomata, allow gas exchanges between the inner photosynthetic tissue and the atmosphere. Regulation of stomatal aperture, preventing excess transpirational vapor loss, relies on turgor changes of two highly differentiated epidermal cells surrounding the pore, the guard cells. Increased guard cell turgor due to increased solute accumulation results in stomatal opening, whereas decreased guard cell turgor due to decreased solute accumulation results in stomatal closing. Here we provide direct evidence, based on reverse genetics approaches, that the
Arabidopsis GORK Shaker
gene encodes the major voltage-gated outwardly rectifying K
+
channel of the guard cell membrane. Expression of GORK dominant negative mutant polypeptides in transgenic
Arabidopsis
was found to strongly reduce outwardly rectifying K
+
channel activity in the guard cell membrane, and disruption of the
GORK
gene (T-DNA insertion knockout mutant) fully suppressed this activity. Bioassays on epidermal peels revealed that disruption of GORK activity resulted in impaired stomatal closure in response to darkness or the stress hormone azobenzenearsonate. Transpiration measurements on excised rosettes and intact plants (grown in hydroponic conditions or submitted to water stress) revealed that absence of GORK activity resulted in increased water consumption. The whole set of data indicates that
GORK
is likely to play a crucial role in adaptation to drought in fluctuating environments.
Molecular characterization of the 1,3-propanediol (1,3-PD) operon of
Clostridium butyricum
Authors: Raynaud Céline , Sarçabal Patricia , Meynial-Salles Isabelle , Croux Christian , Soucaille Philippe
Published: April 2003

The genes encoding the 1,3-propanediol (1,3-PD) operon of
Clostridium butyricum
VPI1718 were characterized from a molecular and a biochemical point of view. This operon is composed of three genes,
dhaB
1,
dhaB
2, and
dha
T. When grown in a vitamin B12-free mineral medium with glycerol as carbon source,
Escherichia coli
expressing
dhaB
1,
dhaB
2, and
dha
T produces 1,3-PD and high glycerol dehydratase and 1,3-PD dehydrogenase activities.
dhaB
1 and
dhaB
2 encode, respectively, a new type of glycerol dehydratase and its activator protein. The deduced proteins DhaB1 and DhaB2, with calculated molecular masses of 88,074 and 34,149 Da, respectively, showed no homology with the known glycerol dehydratases that are all B12 dependent but significant similarity with the pyruvate formate lyases and pyruvate formate lyases activating enzymes and their homologues. The 1,158-bp
dha
T gene codes for a 1,3-PD dehydrogenase with a calculated molecular mass of 41,558 Da, revealing a high level of identity with other DhaT proteins from natural 1,3-PD producers. The expression of the 1,3-PD operon in
C. butyricum
is regulated at the transcriptional level, and this regulation seems to involve a two-component signal transduction system DhaAS/DhaA, which may have a similar function to DhaR, a transcriptional regulator found in other natural 1,3-PD producers. The discovery of a glycerol dehydratase, coenzyme B12 independent, should significantly influence the development of an economical vitamin B12-free biological process for the production of 1,3-PD from renewable resources.
Motion generation by
Drosophila
mechanosensory neurons
Authors: Göpfert M. C. , Robert D.
Published: April 2003

In
Drosophila melanogaster
, hearing is supported by mechanosensory neurons transducing sound-induced vibrations of the antenna. It is shown here that these neurons additionally generate motions that mechanically drive the antenna and tune it to relevant sounds. Motion generation in the
Drosophila
auditory system is betrayed by the auditory mechanics; the antenna of the fly nonlinearly alters its tuning as stimulus intensity declines and oscillates spontaneously in the absence of sound. The susceptibility of auditory motion generation to mechanosensory mutations shows that motion is generated by mechanosensory neurons. Motion generation depends on molecular components of the mechanosensory transduction machinery (NompA, NompC, Btv, and TilB), apparently involving mechanical activity of ciliated dendrites and microtubule-dependent motors. Hence, in analogy to vertebrate hair cells, the mechanosensory neurons of the fly serve dual, transducing, and actuating roles, documenting a striking functional parallel between the vertebrate cochlea and the ears of
Drosophila
.
The functional analysis of insulator interactions in the
Drosophila
embryo
Authors: Majumder Parimal , Cai Haini N.
Published: April 2003

Chromatin boundaries or insulators modulate enhancer–promoter interactions in complex genetic loci. However, the mechanism underlying insulator activity is not known. Previous studies showed that the activity of the
Drosophila
suHw insulator is abolished by the tandem arrangement (pairing) of the insulator elements, suggesting that interactions between insulators or like elements may be involved in their enhancer-blocking mechanism. To test whether such phenomenon reflects a general property of chromatin insulators, we tested the effect of pairing on enhancer-blocking activity of 11 homologous and heterologous insulator combinations using suHw, scs, or SF1 insulators. We found that, unlike the homologous pairing of suHw, the heterologous combinations of suHw with other insulators do not reduce their enhancer-blocking activity. Rather, paired insulators exhibit a higher level of enhancer-blocking activity than either single insulator alone, suggesting that they can function independently or additively. Furthermore, the analyses of two additional chromatin boundaries, scs and SF1, in homologous or heterologous pairing with other boundary elements, also showed no reduction but rather enhancement of insulator activity. We propose that diverse mechanisms may underlie insulator activity, and selective interactions among insulators could influence their function as well as the formations of independent chromatin domains.
Evidence for a lack of DNA double-strand break repair in human cells exposed to very low x-ray doses
Authors: Rothkamm Kai , Löbrich Markus
Published: April 2003

DNA double-strand breaks (DSBs) are generally accepted to be the most biologically significant lesion by which ionizing radiation causes cancer and hereditary disease. However, no information on the induction and processing of DSBs after physiologically relevant radiation doses is available. Many of the methods used to measure DSB repair inadvertently introduce this form of damage as part of the methodology, and hence are limited in their sensitivity. Here we present evidence that foci of γ-H2AX (a phosphorylated histone), detected by immunofluorescence, are quantitatively the same as DSBs and are capable of quantifying the repair of individual DSBs. This finding allows the investigation of DSB repair after radiation doses as low as 1 mGy, an improvement by several orders of magnitude over current methods. Surprisingly, DSBs induced in cultures of nondividing primary human fibroblasts by very low radiation doses (≈1 mGy) remain unrepaired for many days, in strong contrast to efficient DSB repair that is observed at higher doses. However, the level of DSBs in irradiated cultures decreases to that of unirradiated cell cultures if the cells are allowed to proliferate after irradiation, and we present evidence that this effect may be caused by an elimination of the cells carrying unrepaired DSBs. The results presented are in contrast to current models of risk assessment that assume that cellular responses are equally efficient at low and high doses, and provide the opportunity to employ γ-H2AX foci formation as a direct biomarker for human exposure to low quantities of ionizing radiation.
A 1-Mb resolution radiation hybrid map of the canine genome
Authors: Guyon Richard , Lorentzen Travis D. , Hitte Christophe , Kim Lisa , Cadieu Edouard , Parker Heidi G. , Quignon Pascale , Lowe Jennifer K. , Renier Corinne , Gelfenbeyn Boris , Vignaux Françoise , DeFrance Hawkins B. , Gloux Stephanie , Mahairas Gregory G. , André Catherine , Galibert Francis , Ostrander Elaine A.
Published: April 2003

The purebred dog population consists of >300 partially inbred genetic isolates or breeds. Restriction of gene flow between breeds, together with strong selection for traits, has led to the establishment of a unique resource for dissecting the genetic basis of simple and complex mammalian traits. Toward this end, we present a comprehensive radiation hybrid map of the canine genome composed of 3,270 markers including 1,596 microsatellite-based markers, 900 cloned gene sequences and ESTs, 668 canine-specific bacterial artificial chromosome (BAC) ends, and 106 sequence-tagged sites. The map was constructed by using the RHDF5000-2 whole-genome radiation hybrid panel and computed by using
multimap
and
tsp/concorde
. The 3,270 markers map to 3,021 unique positions and define an average intermarker distance corresponding to 1 Mb. We also define a minimal screening set of 325 highly informative well spaced markers, to be used in the initiation of genome-wide scans. The well defined synteny between the dog and human genomes, established in part as a function of this work by the identification of 85 conserved fragments, will allow follow-up of initial findings of linkage by selection of candidate genes from the human genome sequence. This work continues to define the canine system as the method of choice in the pursuit of the genes causing mammalian variation and disease.
Three cadherin alleles associated with resistance to
Bacillus thuringiensis
in pink bollworm
Authors: Morin Shai , Biggs Robert W. , Sisterson Mark S. , Shriver Laura , Ellers-Kirk Christa , Higginson Dawn , Holley Daniel , Gahan Linda J. , Heckel David G. , Carrière Yves , Dennehy Timothy J. , Brown Judith K. , Tabashnik Bruce E.
Published: April 2003

Evolution of resistance by pests is the main threat to long-term insect control by transgenic crops that produce
Bacillus thuringiensis
(Bt) toxins. Because inheritance of resistance to the Bt toxins in transgenic crops is typically recessive, DNA-based screening for resistance alleles in heterozygotes is potentially much more efficient than detection of resistant homozygotes with bioassays. Such screening, however, requires knowledge of the resistance alleles in field populations of pests that are associated with survival on Bt crops. Here we report that field populations of pink bollworm (
Pectinophora gossypiella
), a major cotton pest, harbored three mutant alleles of a cadherin-encoding gene linked with resistance to Bt toxin Cry1Ac and survival on transgenic Bt cotton. Each of the three resistance alleles has a deletion expected to eliminate at least eight amino acids upstream of the putative toxin-binding region of the cadherin protein. Larvae with two resistance alleles in any combination were resistant, whereas those with one or none were susceptible to Cry1Ac. Together with previous evidence, the results reported here identify the cadherin gene as a leading target for DNA-based screening of resistance to Bt crops in lepidopteran pests.
Hot L1s account for the bulk of retrotransposition in the human population
Authors: Brouha Brook , Schustak Joshua , Badge Richard M. , Lutz-Prigge Sheila , Farley Alexander H. , Moran John V. , Kazazian Haig H.
Published: April 2003

Although LINE-1 (long interspersed nucleotide element-1, L1) retrotransposons comprise 17% of the human genome, an exhaustive search of the December 2001 “freeze” of the haploid human genome working draft sequence (95% complete) yielded only 90 L1s with intact ORFs. We demonstrate that 38 of 86 (44%) L1s are polymorphic as to their presence in human populations. We cloned 82 (91%) of the 90 L1s and found that 40 of the 82 (49%) are active in a cultured cell retrotransposition assay. From these data, we predict that there are 80–100 retrotransposition-competent L1s in an average human being. Remarkably, 84% of assayed retrotransposition capability was present in six highly active L1s (hot L1s). By comparison, four of five full-length L1s involved in recent human insertions had retrotransposition activity comparable to the six hot L1s in the human genome working draft sequence. Thus, our data indicate that most L1 retrotransposition in the human population stems from hot L1s, with the remaining elements playing a lesser role in genome plasticity.
Analysis of gene expression in the developing mouse retina
Authors: Díaz Elva , Yang Yee Hwa , Ferreira Todd , Loh Kenneth C. , Okazaki Yasushi , Hayashizaki Yoshihide , Tessier-Lavigne Marc , Speed Terence P. , Ngai John
Published: April 2003

In the visual system, differential gene expression underlies development of the anterior–posterior and dorsal–ventral axes. Here we present the results of a microarray screen to identify genes differentially expressed in the developing retina. We assayed gene expression in nasal (anterior), temporal (posterior), dorsal, and ventral embryonic mouse retina. We used a statistical method to estimate gene expression between different retina regions. Genes were clustered according to their expression pattern and were ranked within each cluster. We identified groups of genes expressed in gradients or with restricted patterns of expression as verified by
in situ
hybridization. A common theme for the identified genes is the differential expression in the dorsal-ventral axis. By analyzing gene expression patterns, we provide insight into the molecular organization of the developing retina.
Predicted correspondence between species abundances and dendrograms of niche similarities
Authors: Sugihara George , Bersier Louis-Félix , Southwood T. Richard E. , Pimm Stuart L. , May Robert M.
Published: April 2003

We examine a hypothesized relationship between two descriptions of community structure: the niche-overlap dendrogram that describes the ecological similarities of species and the pattern of relative abundances. Specifically, we examine the way in which this relationship follows from the niche hierarchy model, whose fundamental assumption is a direct connection between abundances and underlying hierarchical community organization. We test three important, although correlated, predictions of the niche hierarchy model and show that they are upheld in a set of 11 communities (encompassing fishes, amphibians, lizards, and birds) where both abundances and dendrograms were reported. First, species that are highly nested in the dendrogram are on average less abundant than species from branches less subdivided. Second, and more significantly, more equitable community abundances are associated with more evenly branched dendrogram structures, whereas less equitable abundances are associated with less even dendrograms. This relationship shows that abundance patterns can give insight into less visible aspects of community organization. Third, one can recover the distribution of proportional abundances seen in assemblages containing two species by treating each branch point in the dendrogram as a two-species case. This reconstruction cannot be achieved if abundances and the dendrogram are unrelated and suggests a method for hierarchically decomposing systems. To our knowledge, this is the first test of a species abundance model based on nontrivial predictions as to the origins and causes of abundance patterns, and not simply on the goodness-of-fit of distributions.
Protein
O
-fucosyltransferase 1 is an essential component of Notch signaling pathways
Authors: Shi Shaolin , Stanley Pamela
Published: April 2003

Notch receptor signaling regulates cell growth and differentiation, and core components of Notch signaling pathways are conserved from
Drosophila
to humans. Fringe glycosyltransferases are crucial modulators of Notch signaling that act on epidermal growth factor (EGF)-like repeats in the Notch receptor extracellular domain. The substrate of Fringe is EGF-
O
-fucose and the transfer of fucose to Notch by protein
O
-fucosyltransferase 1 is necessary for Fringe to function.
O
-fucose also occurs on Cripto and on Notch ligands. Here we show that mouse embryos lacking protein
O
-fucosyltransferase 1 die at midgestation with severe defects in somitogenesis, vasculogenesis, cardiogenesis, and neurogenesis. The phenotype is similar to that of embryos lacking downstream effectors of all Notch signaling pathways such as presenilins or RBP-Jκ, and is different from Cripto, Notch receptor, Notch ligand, or Fringe null phenotypes. Protein
O
-fucosyltransferase 1 is therefore an essential core member of Notch signaling pathways in mammals.
Altered positive selection due to corecognition of floppy peptide/MHC II conformers supports an integrative model of thymic selection
Authors: Viret Christophe , He Xin , Janeway Charles A.
Published: April 2003

Thymocytes bearing the Eα52-68/I-Abcomplex-specific 1H3.1 αβ T cell antigen receptor are positively selected in Ab-Ep [Ab-Ep transgenic, invariant chain (Ii)−/−, I-Aβb−/−] mice, where I-Abmolecules present only Eα52-68. Although Ii reintroduction led to deletion, I-Aβbreintroduction disrupted positive selection. T cell antigen receptor transgenic Ab-Ep I-Aβb+mice had a large thymus with an increased absolute number of CD4+CD8+cells and no overt signs of deletion. Unlike Ab-Ep Ii+antigen-presenting cells, Ab-Ep I-Aβb+antigen-presenting cells did not activate 1H3.1 T cells. However, their capacity to present Eα52-68 was intact. Thus, positive selection of 1H3.1 thymocytes on the tight compact Eα52-68/I-Abcomplex is neutralized by the corecognition of loose compact self-peptide/I-Abconformers that do not interfere with the cognate activation of mature 1H3.1 T cells. The data support the notion that the integration of distinct signals generated by the simultaneous recognition of multiple self-peptide/MHC complexes directs intrathymic selection of T cells.
Inhibition of rat corneal angiogenesis by a nuclease-resistant RNA aptamer specific for angiopoietin-2
Authors: White Rebekah R. , Shan Siqing , Rusconi Christopher P. , Shetty Geetha , Dewhirst Mark W. , Kontos Christopher D. , Sullenger Bruce A.
Published: April 2003

Angiopoietin-2 (Ang2) appears to be a naturally occurring antagonist of the endothelial receptor tyrosine kinase Tie2, an important regulator of vascular stability. Destabilization of the endothelium by Ang2 is believed to potentiate the actions of proangiogenic growth factors. To investigate the specific role of Ang2 in the adult vasculature, we generated a nuclease-resistant RNA aptamer that binds and inhibits Ang2 but not the related Tie2 agonist, angiopoietin-1. Local delivery of this aptamer but not a partially scrambled mutant aptamer inhibited basic fibroblast growth factor-mediated neovascularization in the rat corneal micropocket angiogenesis assay. These
in vivo
data directly demonstrate that a specific inhibitor of Ang2 can act as an antiangiogenic agent.
Identifying residue–residue clashes in protein hybrids by using a second-order mean-field approach
Authors: Moore Gregory L. , Maranas Costas D.
Published: April 2003

In this article, a second-order mean-field-based approach is introduced for characterizing the complete set of residue–residue couplings consistent with a given protein structure. This information is subsequently used to classify protein hybrids with respect to their potential to be functional based on the presence/absence and severity of clashing residue–residue interactions. First, atomistic representations of both the native and denatured states are used to calculate rotamer–backbone, rotamer–intrinsic, and rotamer–rotamer conformational energies. Next, this complete conformational energy table is coupled with a second-order mean-field description to elucidate the probabilities of all possible rotamer–rotamer combinations in a minimum Helmholtz free-energy ensemble. Computational results for the dihydrofolate reductase family reveal correlation in substitution patterns between not only contacting but also distal second-order structural elements. Residue–residue clashes in hybrid proteins are quantified by contrasting the ensemble probabilities of protein hybrids against the ones of the original parental sequences. Good agreement with experimental data is demonstrated by superimposing these clashes against the functional crossover profiles of bidirectional incremental truncation libraries for
Escherichia coli
and human glycinamide ribonucleotide transformylases.
Total chemical synthesis of a functional interacting protein pair: The protooncogene H-Ras and the Ras-binding domain of its effector c-Raf1
Authors: Becker Christian F. W. , Hunter Christie L. , Seidel Ralf , Kent Stephen B. H. , Goody Roger S. , Engelhard Martin
Published: April 2003

Generation of biological function by chemical methods is potentially of great importance for the understanding and targeting of physiological processes. Chemical synthesis of proteins offers the ability to alter the properties of target protein molecules in a tailor-made fashion. In the present work it is demonstrated that this methodology can be expanded to the elucidation of protein–protein interactions as exemplified by the complete chemical synthesis of the protooncogene product H-Ras as well as of the Ras-binding domain (RBD) of its effector c-Raf1. The 166-aa polypeptide chain of H-Ras was synthesized by native chemical ligation of three unprotected peptide segments. Similarly, the 81-aa RBD was prepared by ligation of two peptide segments. Both RBD and Ras displayed functional and spectroscopic properties indistinguishable from their recombinant forms as judged by CD spectroscopy and from transient kinetic measurements of the Ras–RBD interaction as well as from nucleotide replacement reactions in Ras. An unnatural amino acid bearing a nitrobenzofurazan side chain was introduced into position 91 of the RBD, providing unique fluorescence properties. The association transient of nitrobenzofurazan labeled with Ras⋅guanosine 5′-β,γ-imidotriphosphate showed a slow phase that had not been detected in earlier work by using other signals.
γ
2
subunit of G protein heterotrimer is an N-end rule ubiquitylation substrate
Authors: Hamilton Maria H. , Cook Lana A. , McRackan Theodore R. , Schey Kevin L. , Hildebrandt John D.
Published: April 2003

Heterotrimeric G proteins transduce signals from activated transmembrane G protein-coupled receptors to appropriate downstream effectors within cells. Signaling specificity is achieved in part by the specific α, β, and γ subunits that compose a given heterotrimer. Additional structural and functional diversity in these subunits is generated at the level of posttranslational modification, offering alternate regulatory mechanisms for G protein signaling. Presented here is the identification of a variant of the γ
2
subunit of G protein heterotrimer purified from bovine brain and the demonstration that this RDTASIA γ
2
variant, containing unique amino acid sequence at its N terminus, is a substrate for ubiquitylation and degradation via the N-end rule pathway. Although N-end-dependent degradation has been shown to have important functions in peptide import, chromosome segregation, angiogenesis, and cardiovascular development, the identification of cellular substrates in mammalian systems has remained elusive. The isolation of RDTASIA γ
2
from a native tissue represents identification of a mammalian N-end rule substrate from a physiological source, and elucidates a mechanism for the targeting of G protein γ subunits for ubiquitylation and degradation.
Reversal of developmental restrictions in neural crest lineages: Transition from Schwann cells to glial-melanocytic precursors
in vitro
Authors: Dupin Elisabeth , Real Carla , Glavieux-Pardanaud Corinne , Vaigot Pierre , Le Douarin Nicole M.
Published: April 2003

In vertebrate embryos, diversification of the lineages arising from the neural crest (NC) is controlled to a large extent by environmental factors. In previous work, we showed that endothelin 3 (ET3) peptide favors the development of glial and melanocytic NC precursors
in vitro
. This factor is also capable of inducing proliferation of cultured epidermal pigment cells and their conversion to glia. ET3 therefore strongly promotes the emergence of melanocytic and glial phenotypes from precursors and acts on the maintenance of these phenotypes. In the present work, we explored the capacity of ET3 to reprogram glial cells into melanocytes. Schwann cells expressing glial-specific markers [such as the Schwann cell myelin protein (SMP)] were isolated from sciatic nerves of quail embryos and cultured
in vitro
. We found that ET3 promotes cell growth and sequential expression of melanocyte differentiation markers in cultures of purified SMP-expressing cells, whereas it had no significant effect on SMP-negative cells from the same nerves. Moreover, we provide evidence for the transition of differentiated Schwann cells to melanocytes in clonal cultures. This transition involves the production of a mixed progeny of melanoblasts/melanocytes, glia, and cells bearing differentiation markers of both phenotypes. Therefore, Schwann cells exposed to ET3 transdifferentiate to melanocytes through reversion to the stage of bipotent glial-melanocytic NC precursors. These findings show that NC-derived pigment and glial cells are phenotypically unstable
in vitro
and may undergo reversal of precursor hierarchy to function as bipotent stem cells.
The role of side-chain interactions in the early steps of aggregation: Molecular dynamics simulations of an amyloid-forming peptide from the yeast prion Sup35
Authors: Gsponer Jörg , Haberthür Urs , Caflisch Amedeo
Published: April 2003

Understanding the early steps of aggregation at atomic detail might be crucial for the rational design of therapeutics preventing diseases associated with amyloid deposits. In this paper, aggregation of the heptapeptide GNNQQNY, from the N-terminal prion-determining domain of the yeast protein Sup35, was studied by 20 molecular dynamics runs for a total simulation time of 20 μs. The simulations generate in-register parallel packing of GNNQQNY β-strands that is consistent with x-ray diffraction and Fourier transform infrared data. The statistically preferred aggregation pathway does not correspond to a purely downhill profile of the energy surface because of the presence of enthalpic barriers that originate from out-of-register interactions. The parallel β-sheet arrangement is favored over the antiparallel because of side-chain contacts; in particular, stacking interactions of the tyrosine rings and hydrogen bonds between amide groups. No ordered aggregation was found in control simulations with the mutant sequence SQNGNQQRG in accord with experimental data and the strong sequence dependence of aggregation.
Speech–sound-selective auditory impairment in children with autism: They can perceive but do not attend
Authors: Čeponienė R. , Lepistö T. , Shestakova A. , Vanhala R. , Alku P. , Näätänen R. , Yaguchi K.
Published: April 2003

In autism, severe abnormalities in social behavior coexist with aberrant attention and deficient language. In the attentional domain, attention to people and socially relevant stimuli is impaired the most. Because socially meaningful stimulus events are physically complex, a deficiency in sensory processing of complex stimuli has been suggested to contribute to aberrant attention and language in autism. This study used event-related brain potentials (ERP) to examine the sensory and early attentional processing of sounds of different complexity in high-functioning children with autism. Acoustically matched simple tones, complex tones, and vowels were presented in separate oddball sequences, in which a repetitive “standard” sound was occasionally replaced by an infrequent “deviant” sound differing from the standard in frequency (by 10%). In addition to sensory responses, deviant sounds elicited an ERP index of automatic sound-change discrimination, the mismatch negativity, and an ERP index of attentional orienting, the P3a. The sensory sound processing was intact in the high-functioning children with autism and was not affected by sound complexity or “speechness.” In contrast, their involuntary orienting was affected by stimulus nature. It was normal to both simple- and complex-tone changes but was entirely abolished by vowel changes. These results demonstrate that, first, auditory orienting deficits in autism cannot be explained by sensory deficits and, second, that orienting deficit in autism might be speech–sound specific.
Modular organization of internal models of tools in the human cerebellum
Authors: Imamizu Hiroshi , Kuroda Tomoe , Miyauchi Satoru , Yoshioka Toshinori , Kawato Mitsuo
Published: April 2003

Human capabilities in dexterously manipulating many different tools suggest modular neural organization at functional levels, but anatomical modularity underlying the capabilities has yet to be demonstrated. Although modularity in phylogenetically older parts of the cerebellum is well known, comparable modularity in the lateral cerebellum for cognitive functions remains unknown. We investigated these issues by functional MRI (fMRI) based on our previous findings of a cerebellar internal model of a tool. After subjects intensively learned to manipulate two novel tools (the rotated mouse whose cursor appeared at a rotated position, and the velocity mouse whose cursor velocity was proportional to the mouse position), they could easily switch between the two. The lateral and posterior cerebellar activities for the two different tools were spatially segregated, and their overlaps were <10%, even at low statistical thresholds. Activities of the rotated mouse were more anterior and lateral than the velocity mouse activities. These results were consistent with predictions by the MOdular Selection And Identification Controller (MOSAIC) model that multiple internal models compete to partition sensory-motor experiences and their outputs are linearly combined for a particular context.
Telomere attrition and Chk2 activation in human heart failure
Authors: Oh Hidemasa , Wang Sam C. , Prahash Arun , Sano Motoaki , Moravec Christine S. , Taffet George E. , Michael Lloyd H. , Youker Keith A. , Entman Mark L. , Schneider Michael D.
Published: April 2003

The “postmitotic” phenotype in adult cardiac muscle exhibits similarities to replicative senescence more generally and constitutes a barrier to effective restorative growth in heart disease. Telomere dysfunction is implicated in senescence and apoptotic signaling but its potential role in heart disorders is unknown. Here, we report that cardiac apoptosis in human heart failure is associated specifically with defective expression of the telomere repeat- binding factor TRF2, telomere shortening, and activation of the DNA damage checkpoint kinase, Chk2. In cultured cardiomyocytes, interference with either TRF2 function or expression triggered telomere erosion and apoptosis, indicating that cell death can occur via this pathway even in postmitotic, noncycling cells; conversely, exogenous TRF2 conferred protection from oxidative stress.
In vivo
, mechanical stress was sufficient to down-regulate TRF2, shorten telomeres, and activate Chk2 in mouse myocardium, and transgenic expression of telomerase reverse transcriptase conferred protection from all three responses. Together, these data suggest that apoptosis in chronic heart failure is mediated in part by telomere dysfunction and suggest an essential role for TRF2 even in postmitotic cells.
Dispersal of NK homeobox gene clusters in amphioxus and humans
Authors: Luke Graham N. , Castro L. Filipe C. , McLay Kirsten , Bird Christine , Coulson Alan , Holland Peter W. H.
Published: April 2003

The
Drosophila melanogaster
genome has six physically clustered NK-related homeobox genes in just 180 kb. Here we show that the NK homeobox gene cluster was an ancient feature of bilaterian animal genomes, but has been secondarily split in chordate ancestry. The NK homeobox gene clusters of amphioxus and vertebrates are each split and dispersed at two equivalent intergenic positions. From the ancestral NK gene cluster, only the Tlx–Lbx and NK3–NK4 linkages have been retained in chordates. This evolutionary pattern is in marked contrast to the Hox and ParaHox gene clusters, which are compact in amphioxus and vertebrates, but have been disrupted in
Drosophila
.
Extracellular polysaccharides mediate pilus retraction during social motility of
Myxococcus
xanthus
Authors: Li Yinuo , Sun Hong , Ma Xiaoyuan , Lu Ann , Lux Renate , Zusman David , Shi Wenyuan
Published: April 2003

Myxococcus xanthus
is a Gram-negative bacterium with a complex life cycle that includes vegetative swarming and fruiting-body formation. Social (S)-motility (coordinated movement of large cell groups) requires both type IV pili and fibrils (extracellular matrix material consisting of polysaccharides and protein). Little is known about the role of this extracellular matrix, or fibril material, in pilus-dependent motility. In this study, mutants lacking fibril material and, therefore, S-motility were found to be hyperpiliated. We demonstrated that addition of fibril material resulted in pilus retraction and rescued this phenotype. The fibril material was further examined to determine the component(s) that were responsible for triggering pilus retraction. Protein-free fibril material was found to be highly active in correcting hyperpiliation. However, the amine sugars present in hydrolyzed fibril material, e.g., glucosamine and
N
-acetylglucosamine (GlcNAc) had no effect on fibril
−
mutants, but, interestingly, cause hyperpiliation in wild-type cells. In contrast, chitin, a natural GlcNAc polymer, was found to restore pilus retraction in hyperpiliated mutants, indicating that a polysaccharide containing amine sugars is likely required for pilus retraction. These data suggest that the interaction of type IV pili with amine-containing polysaccharides on cell and slime-trail surfaces may trigger pilus retraction, resulting in S-motility and slime-trailing behaviors.
Moth chemosensory protein exhibits drastic conformational changes and cooperativity on ligand binding
Authors: Campanacci Valérie , Lartigue Audrey , Hällberg B. Martin , Jones T. Alwyn , Giudici-Orticoni Marie-Therèse , Tegoni Mariella , Cambillau Christian
Published: April 2003

Chemosensory proteins (CSPs) have been proposed to transport hydrophobic chemicals from air to olfactory or taste receptors. They have been isolated from several sensory organs of a wide range of insect species. The x-ray structure of CSPMbraA6, a 112-aa antennal protein from the moth
Mamestra brassicae (Mbra
), was shown to exhibit a novel type of α-helical fold. We have performed a structural and binding study of CSPMbraA6 to get some insights into its possible molecular function. Tryptophan fluorescence quenching demonstrates the ability of CSPMbraA6 to bind several types of semio-chemicals or surrogate ligands with μM
K
d
. Its crystal structure in complex with one of these compounds, 12-bromo-dodecanol, reveals extensive conformational changes on binding, resulting in the formation of a large cavity filled by three ligand molecules. Furthermore, binding cooperativity was demonstrated for some ligands, suggesting a stepwise binding. The peculiar rearrangement of CSPMbraA6 conformation and the cooperativity phenomenon might trigger the recognition of chemicals by receptors and induce subsequent signal transduction.
A rapidly evolving MYB-related protein causes species isolation in
Drosophila
Authors: Barbash Daniel A. , Siino Dominic F. , Tarone Aaron M. , Roote John
Published: April 2003

Matings among different species of animals or plants often result in sterile or lethal hybrids. Identifying the evolutionary forces that create hybrid incompatibility alleles is fundamental to understanding the process of speciation, but very few such alleles have been identified, particularly in model organisms that are amenable to experimental manipulation. We report here the cloning of the first, to our knowledge,
Drosophila melanogaster
gene involved in hybrid incompatibilities,
Hybrid male rescue
(
Hmr
).
Hmr
causes lethality and female sterility in hybrids among
D. melanogaster
and its sibling species. We have found that
Hmr
encodes a protein with homology to a family of MYB-related DNA-binding transcriptional regulators. The HMR protein has evolved both amino acid substitutions and insertions and deletions at an extraordinarily high rate between
D. melanogaster
and its sibling species, including in its predicted DNA-binding domain. Our results suggest that hybrid lethality may result from disruptions in gene regulation, and we also propose that rapid evolution may be a hallmark of speciation genes in general.
Kidney-specific inactivation of the KIF3A subunit of kinesin-II inhibits renal ciliogenesis and produces polycystic kidney disease
Authors: Lin Fangming , Hiesberger Thomas , Cordes Kimberly , Sinclair Angus M. , Goldstein Lawrence S. B. , Somlo Stefan , Igarashi Peter
Published: April 2003

Polycystic kidney disease (PKD) is the most common genetic cause of renal failure in humans. Several proteins that are encoded by genes associated with PKD have recently been identified in primary cilia in renal tubular epithelia. These findings have suggested that abnormalities in cilia formation and function may play a role in the pathogenesis of PKD. To directly determine whether cilia are essential to maintain tubular integrity, we conditionally inactivated KIF3A, a subunit of kinesin-II that is essential for cilia formation, in renal epithelia. Constitutive inactivation of KIF3A produces abnormalities of left–right axis determination and embryonic lethality. Here we show that tissue-specific inactivation of KIF3A in renal tubular epithelial cells results in viable offspring with normal-appearing kidneys at birth. Cysts begin to develop in the kidney at postnatal day 5 and cause renal failure by postnatal day 21. The cyst epithelial cells lack primary cilia and exhibit increased proliferation and apoptosis, apical mislocalization of the epidermal growth factor receptor, increased expression of β-catenin and c-Myc, and inhibition of p21
CIP1
. These results demonstrate that the absence of renal cilia produces both the clinical and cell biological findings associated with PKD. Most generally, the phenotype of
Kif3a
mutant mice suggests a role for primary cilia in the maintenance of lumen-forming epithelial differentiation.
Noncovalent functionalization of carbon nanotubes for highly specific electronic biosensors
Authors: Chen Robert J. , Bangsaruntip Sarunya , Drouvalakis Katerina A. , Wong Shi Kam Nadine , Shim Moonsub , Li Yiming , Kim Woong , Utz Paul J. , Dai Hongjie
Published: April 2003

Novel nanomaterials for bioassay applications represent a rapidly progressing field of nanotechnology and nanobiotechnology. Here, we present an exploration of single-walled carbon nanotubes as a platform for investigating surface–protein and protein–protein binding and developing highly specific electronic biomolecule detectors. Nonspecific binding on nanotubes, a phenomenon found with a wide range of proteins, is overcome by immobilization of polyethylene oxide chains. A general approach is then advanced to enable the selective recognition and binding of target proteins by conjugation of their specific receptors to polyethylene oxide-functionalized nanotubes. This scheme, combined with the sensitivity of nanotube electronic devices, enables highly specific electronic sensors for detecting clinically important biomolecules such as antibodies associated with human autoimmune diseases.
Activation of archaeal transcription by recruitment of the TATA-binding protein
Authors: Ouhammouch Mohamed , Dewhurst Robert E. , Hausner Winfried , Thomm Michael , Geiduschek E. Peter
Published: April 2003

The hyperthermophilic archaeon
Methanococcus jannaschii
encodes two
p
utative
t
ranscription
r
egulators, Ptr1 and Ptr2, that are members of the Lrp/AsnC family of bacterial transcription regulators. In contrast, this archaeon's RNA polymerase and core transcription factors are of eukaryotic type. Using the
M. jannaschii
high-temperature
in vitro
transcription system, we show that Ptr2 is a potent transcriptional activator, and that it conveys its stimulatory effects on its cognate eukaryal-type transcription machinery from an upstream activating region composed of two Ptr2-binding sites. Transcriptional activation is generated, at least in part, by Ptr2-mediated recruitment of the TATA-binding protein to the promoter.
An obligate pollination mutualism and reciprocal diversification in the tree genus
Glochidion
(Euphorbiaceae)
Authors: Kato Makoto , Takimura Atsushi , Kawakita Atsushi
Published: April 2003

Highly coevolved pollination mutualism accompanied by reciprocal diversification has been known in only two plant genera,
Ficus
(Moraceae) and
Yucca
(Agavaceae), which are pollinated exclusively by obligate seed-parasitic wasps and moths, respectively. An additional, highly diversified, species-specific pollination mutualism between a monoecious tree genus,
Glochidion
(Euphorbiaceae), and a moth genus,
Epicephala
(Gracillariidae), is presented here. At night, the small female moth actively deposits pollen on the cryptic stigma of the female flower by using its proboscis, then oviposits into the style. The moth larva infests only a portion of the developing seeds within fruit. We confirmed that at least three
Glochidion
species are pollinated only by their respective seed-parasitic moth species, which could be distinguished by genitalic morphology and mitochondrial DNA sequences. These results and widespread evidence of limited seed infestation by the moths associated with
Glochidion
species suggest that speciation based on the highly specialized
Glochidion
stylar structure and moth oviposition behavior have promoted species diversification in
Glochidion
and its pollinators.
Yeast DNA polymerase η makes functional contacts with the DNA minor groove only at the incoming nucleoside triphosphate
Authors: Washington M. Todd , Wolfle William T. , Spratt Thomas E. , Prakash Louise , Prakash Satya
Published: April 2003

DNA polymerase η (Polη) functions in the proficient bypass of a variety of DNA lesions. Relative to the replicative polymerases, Polη has a greater tolerance for distorted DNA geometries and possesses a low fidelity. X-ray crystal structures and studies with nucleotide analogs have implicated interactions with the DNA minor groove as being crucial for the high fidelity of replicative DNA polymerases. To determine whether Polη also makes such functionally important contacts with the DNA minor groove, here we examine the effects on Polη-catalyzed nucleotide incorporation when 3-deazaguanine, a base analog that lacks the ability to form minor-groove hydrogen bonds with the protein, is substituted for guanine at various positions in the DNA. From these studies, we conclude that Polη makes only a single functional contact with the DNA minor groove at the position of the incoming nucleotide; in this regard, Polη differs from high-fidelity DNA polymerases that are unable to replicate through DNA lesions. These results help explain the proficient ability of Polη for bypassing distorting DNA lesions.
Effector CD8 T cells possess suppressor function after 4-1BB and Toll-like receptor triggering
Authors: Myers Lara , Takahashi Chikara , Mittler Robert S. , Rossi Robert J. , Vella Anthony T.
Published: April 2003

To better understand how innate and adaptive immune responses interact with each other, we combined 4-1BB T cell costimulation with specific adjuvants to determine whether these treatments would influence specific T cell expansion and functionin vivo. In the presence of 4-1BB ligation and Toll-like receptor 3 (TLR)3 and/or TLR4 triggering, CD8 T cell clonal expansion and survival was augmented profoundly. Specific T cells primedin vivowith TLR ligands responded normally toin vitrorecall stimulus, but, surprisingly, copriming with 4-1BB costimulation significantly impaired the recall response even though many more specific effector T cells were rescuedin vivo. Here, we demonstrate that the rescued CD8 T cells suppressed CD4 T cell proliferation via a type β transforming growth factor-dependent mechanism. Thus, 4-1BB and TLR ligands induce survival of specific effector CD8 T cells with suppressive recall potential, which may explain the dual role that 4-1BB activation plays in mediating tumor clearance and prevention of autoimmune disease.
Semiquinone radical intermediate in catecholic estrogen-mediated cytotoxicity and mutagenesis: Chemoprevention strategies with antioxidants
Authors: Samuni Ayelet M. , Chuang Eric Y. , Krishna Murali C. , Stein William , DeGraff William , Russo Angelo , Mitchell James B.
Published: April 2003

Modulation of the cytotoxicity and mutagenicity of 4-hydroxyestradiol (4-OHE
2
), an oxidative metabolite of estrogen, by antioxidants was assessed in human MCF7 cells and TK-6 lymphoblast cells. The cytotoxicity of the catecholic estrogens was potentiated by depletion of intracellular glutathione and was independent of oxygen concentration. Agents such as the nitroxide Tempol can facilitate the oxidation of the semiquinone to the Q and enhanced 4-OHE
2
cytotoxicity. Conversely, reducing agents such as ascorbate, cysteine, and 1,4-dihydroxytetramethylpiperidine (THP) protected against cytotoxicity and decreased mutation induction, presumably by reducing the semiquinone to the hydroquinone. Our results support the proposition that oxidation of the semiquinone to the corresponding Q is crucial in eliciting the deleterious effects of catecholic estrogens. Furthermore, because the deleterious effects of 4-OHE
2
were abrogated by dietary and synthetic antioxidants, our results would support the chemopreventive use of diets rich in reducing substances (vitamins and added synthetic antioxidants) as a means of decreasing the risks associated with estrogen exposure and developing of breast cancer.
Genetic changes occurring in established tumors rapidly stimulate new antibody responses
Authors: Spiotto Michael T. , Reth Michael A. , Schreiber Hans
Published: April 2003

Cancer patients and tumor-bearing mice possess serum antibodies that recognize antigens expressed by cancer cells at the time of diagnosis. After diagnosis, cancers progress to more aggressive stages, most often by acquiring new genetic changes that can give rise to new proteins, some of which are antigenic. However, at these relatively later stages of tumor growth, it remains unclear whether, when, and how a host can generate
de novo
antibody responses against these newly appearing tumor antigens. To this end, we used a tamoxifen-regulated Cre-loxP system, MerCreMer, to induce genetic recombination in cancer cells of well-established tumors, resulting in increased enhanced green fluorescence protein (EGFP) expression. These late tumor-bearing mice generated specific IgG antibodies against EGFP within 3 wk after antigen induction. Mice generated these antibody responses in the presence of preexisting anti-tumor antibody responses. Preexisting CD4
+
T cell responses to already expressed tumor antigens likely enhanced antibody responses to the induced EGFP antigen. By analogy, new antibody responses in cancer patients may identify genetic changes occurring in a growing tumor and indicate imminent tumor progression.