bluuebunny/crossref_metadata_2025 huggingface dataset:
Amino acids in the Martian meteorite Nakhla
Authors: Glavin Daniel P. , Bada Jeffrey L. , Brinton Karen L. F. , McDonald Gene D.
Published: July 2002

A suite of protein and nonprotein amino acids were detected with high-performance liquid chromatography in the water- and acid-soluble components of an interior fragment of the Martian meteorite Nakhla, which fell in Egypt in 1911. Aspartic and glutamic acids, glycine, alanine, β-alanine, and γ-amino-
n
-butyric acid (γ-ABA) were the most abundant amino acids detected and were found primarily in the 6 M HCl-hydrolyzed, hot water extract. The concentrations ranged from 20 to 330 parts per billion of bulk meteorite. The amino acid distribution in Nakhla, including the
d/l
ratios (values range from <0.1 to 0.5), is similar to what is found in bacterially degraded organic matter. The amino acids in Nakhla appear to be derived from terrestrial organic matter that infiltrated the meteorite soon after its fall to Earth, although it is possible that some of the amino acids are endogenous to the meteorite. The rapid amino acid contamination of Martian meteorites after direct exposure to the terrestrial environment has important implications for Mars sample-return missions and the curation of the samples from the time of their delivery to Earth.
Flagellar determinants of bacterial sensitivity to χ-phage
Authors: Samuel Aravinthan D. T. , Pitta Thomas P. , Ryu William S. , Danese Paul N. , Leung Edward C. W. , Berg Howard C.
Published: July 2002

Bacteriophage χ is known to infect motile strains of enteric bacteria by adsorbing randomly along the length of a flagellar filament and then injecting its DNA into the bacterial cell at the filament base. Here, we provide evidence for a “nut and bolt” model for translocation of phage along the filament: the tail fiber of χ fits the grooves formed by helical rows of flagellin monomers, and active flagellar rotation forces the phage to follow the grooves as a nut follows the threads of a bolt.
Direct sequencing of bacterial and P1 artificial chromosome-nested deletions for identifying position-specific single-nucleotide polymorphisms
Authors: Chatterjee Pradeep K. , Yarnall David P. , Haneline Stephen A. , Godlevski Michele M. , Thornber Simon J. , Robinson Phil S. , Davies Heather E. , White Nicola J. , Riley John H. , Shepherd Nancy S.
Published: July 2002

A loxP-transposon retrofitting strategy for generating large nested
deletions from one end of the insert DNA in bacterial artificial
chromosomes and P1 artificial chromosomes was described recently
[Chatterjee, P. K. & Coren, J. S. (1997)
Nucleic
Acids Res
. 25, 2205–2212]. In this report, we combine this
procedure with direct sequencing of nested-deletion templates by using
primers located in the transposon end to illustrate its value for
position-specific single-nucleotide polymorphism (SNP) discovery from
chosen regions of large insert clones. A simple ampicillin sensitivity
screen was developed to facilitate identification and recovery of
deletion clones free of transduced transposon plasmid. This directed
approach requires minimal DNA sequencing, and no
in
vitro
subclone library generation; positionally oriented SNPs
are a consequence of the method. The procedure is used to discover new
SNPs as well as physically map those identified from random subcloned
libraries or sequence databases. The deletion templates, positioned
SNPs, and markers are also used to orient large insert clones into a
contig. The deletion clone can serve as a ready resource for future
functional genomic studies because each carries a mammalian
cell-specific antibiotic resistance gene from the transposon.
Furthermore, the technique should be especially applicable to the
analysis of genomes for which a full genome sequence or radiation
hybrid cell lines are unavailable.
Oryza sativa PSK
gene encodes a precursor of phytosulfokine-α, a sulfated peptide growth factor found in plants
Authors: Yang Heping , Matsubayashi Yoshikatsu , Nakamura Kenzo , Sakagami Youji
Published: July 2002

Phytosulfokine-α [PSK-α, Tyr(SO
3
H)-Ile-Tyr(SO
3
H)-Thr-Gln], a sulfated mitogenic peptide found in plants, strongly promotes proliferation of plant cells in culture at very low concentrations.
Oryza sativa PSK
(
OsPSK
) cDNA encoding a PSK-α precursor has been isolated. The cDNA is 725 base pairs long, and the 89-aa product, preprophytosulfokine, has a 22-aa hydrophobic region that resembles a cleavable leader peptide at its NH
2
terminus. The PSK-α sequence occurs only once within the precursor, close to the COOH terminus. [Ser
4
]PSK-α was secreted by transgenic rice Oc cells harboring a mutated
OsPSK
cDNA, suggesting proteolytic processing from the larger precursor, a feature commonly found in animal systems. Whereas PSK-α in conditioned medium with sense transgenic Oc cells was 1.6 times as concentrated as in the control case, antisense transgenic Oc cells produced less than 60% of the control level. Preprophytosulfokine mRNA was detected at an elevated constitutive level in rice Oc culture cells on RNA blot analysis. Although PSK-α molecules have never been identified in any intact plant, reverse transcription–PCR analysis demonstrated that
OsPSK
is expressed in rice seedlings, indicating that PSK-α may be important for plant cell proliferation both
in vitro
and
in vivo
. DNA blot analysis demonstrated that
OsPSK
homologs may occur in dicot as well as monocot plants.
Topoisomerase II drives DNA transport by hydrolyzing one ATP
Authors: Baird Cheryl L. , Harkins Timothy T. , Morris Shayne K. , Lindsley Janet E.
Published: July 2002

DNA topoisomerase II is a homodimeric molecular machine that couples ATP usage to the transport of one DNA segment through a transient break in another segment. In the presence of a nonhydrolyzable ATP analog, the enzyme is known to promote a single turnover of DNA transport. Current models for the enzyme’s mechanism based on this result have hydrolysis of two ATPs as the last step, used only to reset the enzyme for another round of reaction. Using rapid-quench techniques, topoisomerase II recently was shown to hydrolyze its two bound ATPs in a strictly sequential manner. This result is incongruous with the models based on the nonhydrolyzable ATP analog data. Here we present evidence that hydrolysis of one ATP by topoisomerase II precedes, and accelerates, DNA transport. These results indicate that important features of this enzyme’s mechanism previously have been overlooked because of the reliance on nonhydrolyzable analogs for studying a single reaction turnover. A model for the mechanism of topoisomerase II is presented to show how hydrolysis of one ATP could drive DNA transport.
Inhibition of DNA methyltransferase stimulates the expression of signal transducer and activator of transcription 1, 2, and 3 genes in colon tumor cells
Authors: Karpf Adam R. , Peterson Peter W. , Rawlins Joseph T. , Dalley Brian K. , Yang Qian , Albertsen Hans , Jones David A.
Published: July 2002

Inhibitors of DNA methyltransferase, typified by 5-aza-2′-deoxycytidine (5-Aza-CdR), induce the expression of genes transcriptionally down-regulated byde novomethylation in tumor cells. We utilized gene expression microarrays to examine the effects of 5-Aza-CdR treatment in HT29 colon adenocarcinoma cells. This analysis revealed the induction of a set of genes that implicated IFN signaling in the HT29 cellular response to 5-Aza-CdR. Subsequent investigations revealed that the induction of this gene set correlates with the induction of signal transducer and activator of transcription (STAT) 1, 2, and 3 genes and their activation by endogenous IFN-α. These observations implicate the induction of the IFN-response pathway as a major cellular response to 5-Aza-CdR and suggests that the expression of STATs 1, 2, and 3 can be regulated by DNA methylation. Consistent with STAT’s limiting cell responsiveness to IFN, we found that 5-Aza-CdR treatment sensitized HT29 cells to growth inhibition by exogenous IFN-α2a, indicating that 5-Aza-CdR should be investigated as a potentiator of IFN responsiveness in certain IFN-resistant tumors.
Programmed cell death in castor bean endosperm is associated with the accumulation and release of a cysteine endopeptidase from ricinosomes
Authors: Schmid Markus , Simpson David , Gietl Christine
Published: July 2002

The cells of the endosperm of castor bean seeds (
Ricinus communis
) undergo programmed cell death during germination, after their oil and protein reserves have been mobilized. Nuclear DNA fragmentation first was observed at day 3 in the endosperm cells immediately adjacent to the cotyledons and progressed across to the outermost cell layers by day 5. We also detected the accumulation of small organelles known as ricinosomes, by using an antibody against a cysteine endoprotease. By the time the nuclear DNA was susceptible to heavy label by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling, the ricinosomes had released into the cytoplasm their content of cysteine endoprotease, which became activated because of the cleavage of its propeptide. The cysteine endoprotease is distinguished by a C-terminal KDEL sequence, although it is not retained in the lumen of the endoplasmic reticulum and is a marker for ricinosomes. Homologous proteases are found in the senescing tissues of other plants, including the petals of the daylily. Ricinosomes were identified in this tissue by electron microscopy and immunocytochemistry. It seems that ricinosomes are not unique to
Ricinus
and play an important role in the degradation of plant cell contents during programmed cell death.
SEM1
, a homologue of the split hand/split foot malformation candidate gene
Dss1
, regulates exocytosis and pseudohyphal differentiation in yeast
Authors: Jäntti Jussi , Lahdenranta Johanna , Olkkonen Vesa M. , Söderlund Hans , Keränen Sirkka
Published: July 2002

The exocyst is an essential multiprotein complex mediating polarized secretion in yeast. Here we describe a gene,
SEM1
, that can multicopy-suppress exocyst mutants
sec3–2
,
sec8–9
,
sec10–2
, and
sec15–1
.
SEM1
is highly conserved among eukaryotic species. Its human homologue,
DSS1
, has been suggested as a candidate gene for the split hand/split foot malformation disorder.
SEM1
is not an essential gene. However, its deletion rescued growth of the temperature-sensitive exocyst mutants
sec3–2
,
sec8–9
,
sec10–1,
and
sec15–1
at the restrictive temperature. Cell fractionation showed that Sem1p is mainly cytosolic but also associates with the microsomal fraction. In linear sucrose gradients, Sem1p cosedimented with the exocyst component Sec8p. In diploid cells that normally do not form pseudohyphae (S288C background), deletion of
SEM1
triggered pseudohyphal growth. This phenotype was abolished after reintroduction of either
SEM1
or the mouse homologue
Dss1
into the cells. In diploids that have normal capacity for pseudohyphal growth (Σ1278b background), deletion of
SEM1
enhanced filamentous growth. The functionality of both
SEM1
and
Dss1
in a differentiation process in yeast suggests that
Dss1
indeed could be the gene affected in the split hand/split foot malformation disorder. These results characterize
SEM1
as a regulator of both exocyst function and pseudohyphal differentiation and suggest a unique link between these two cellular functions in yeast.
The charged region of Hsp90 modulates the function of the N-terminal domain
Authors: Scheibel Thomas , Siegmund Heiko Ingo , Jaenicke Rainer , Ganz Peter , Lilie Hauke , Buchner Johannes
Published: July 2002

Hsp90, an abundant heat shock protein that is highly expressed even
under physiological conditions, is involved in the folding of key
molecules of the cellular signal transduction system such as kinases
and steroid receptors. It seems to contain two chaperone sites
differing in substrate specificity. Binding of ATP or the antitumor
drug geldanamycin alters the substrate affinity of the N-terminal
chaperone site, whereas both substances show no influence on the
C-terminal one. In wild-type Hsp90 the fragments containing the
chaperone sites are connected by a highly charged linker of various
lengths in different organisms. As this linker region represents the
most striking difference between bacterial and eukaryotic Hsp90s, it
may be involved in a gain of function of eukaryotic Hsp90s. Here, we
have analyzed a fragment of yeast Hsp90 consisting of the N-terminal
domain and the charged region (N272) in comparison with the isolated
N-terminal domain (N210). We show that the charged region causes an
increase in the affinity of the N-terminal domain for nonnative protein
and establishes a crosstalk between peptide and ATP binding. Thus, the
binding of peptide to N272 decreases its affinity for ATP and
geldanamycin, whereas the ATP-binding properties of the monomeric
N-terminal domain N210 are not influenced by peptide binding. We
propose that the charged region connecting the two chaperone domains
plays an important role in regulating chaperone function of Hsp90.
Dynamics and elasticity of the fibronectin matrix in living cell culture visualized by fibronectin–green fluorescent protein
Authors: Ohashi Tomoo , Kiehart Daniel P. , Erickson Harold P.
Published: July 2002

Fibronectin (FN) forms the primitive fibrillar matrix in both embryos and healing wounds. To study the matrix in living cell cultures, we have constructed a cell line that secretes FN molecules chimeric with green fluorescent protein. These FN–green fluorescent protein molecules were assembled into a typical matrix that was easily visualized by fluorescence over periods of several hours. FN fibrils remained mostly straight, and they were seen to extend and contract to accommodate movements of the cells, indicating that they are elastic. When fibrils were broken or detached from cells, they contracted to less than one-fourth of their extended length, demonstrating that they are highly stretched in the living culture. Previous work from other laboratories has suggested that cryptic sites for FN assembly may be exposed by tension on FN. Our results show directly that FN matrix fibrils are not only under tension but are also highly stretched. This stretched state of FN is an obvious candidate for exposing the cryptic assembly sites.
A mutant of
Mycobacterium smegmatis
defective in the biosynthesis of mycolic acids accumulates meromycolates
Authors: Liu Jun , Nikaido Hiroshi
Published: July 2002

Mycolic acids are a major constituent of the mycobacterial cell wall, and they form an effective permeability barrier to protect mycobacteria from antimicrobial agents. Although the chemical structures of mycolic acids are well established, little is known on their biosynthesis. We have isolated a mycolate-deficient mutant strain of
Mycobacterium smegmatis
mc
2
-155 by chemical mutagenesis followed by screening for increased sensitivity to novobiocin. This mutant also was hypersensitive to other hydrophobic compounds such as crystal violet, rifampicin, and erythromycin. Entry of hydrophobic probes into mutant cells occurred much more rapidly than that into the wild-type cells. HPLC and TLC analysis of fatty acid composition after saponification showed that the mutant failed to synthesize full-length mycolic acids. Instead, it accumulated a series of long-chain fatty acids, which were not detected in the wild-type strain. Analysis by
1
H NMR, electrospray and electron impact mass spectroscopy, and permanganate cleavage of double bonds showed that these compounds corresponded to the incomplete meromycolate chain of mycolic acids, except for the presence of a β-hydroxyl group. This direct identification of meromycolates as precursors of mycolic acids provides a strong support for the previously proposed pathway for mycolic acid biosynthesis involving the separate synthesis of meromycolate chain and the α-branch of mycolic acids, followed by the joining of these two branches.
Neuroimaging of genesis and satiation of thirst and an interoceptor-driven theory of origins of primary consciousness
Authors: Denton Derek , Shade Robert , Zamarippa Frank , Egan Gary , Blair-West John , McKinley Michael , Lancaster Jack , Fox Peter
Published: July 2002

There are defined hypothalamic functions in the genesis of thirst, but little is known of the cortical processes subserving consciousness of thirst notwithstanding the medical disorders that occur in psychiatric illness, addiction, and the attested decline of thirst with aging. In 10 adult males, positron emission tomography scans were made (
i
) during genesis of moderate thirst by infusion of i.v. hypertonic saline 0.51 M, (
ii
) after irrigation of the mouth with water to remove the sensation of dryness, and (
iii
) 3, 14, 45, and 60 minutes after drinking water to fully satiate thirst. The correlation of regional cerebral blood flow with thirst score showed the major activation to be in the posterior cingulate. Maximum thirst sensation evoked 13 highly significant activations and 9 deactivations in cingulate and parahippocampal gyri, insula, thalamus, amygdala, and mesencephalon. It is possible that cingulate sites (Brodmann’s areas 32, 24, and 31) that persisted with wet mouth but disappeared immediately after drinking to satiation may have an important role in the consciousness of thirst. Consciousness of thirst, a primal vegetative emotion, and satiation of thirst appear to be subserved by phylogenetically ancient brain regions. This is salient to current discussion on evolutionary emergence of primary consciousness.
Mimicking dominant negative inhibition of prion replication through structure-based drug design
Authors: Perrier Véronique , Wallace Andrew C. , Kaneko Kiyotoshi , Safar Jiri , Prusiner Stanley B. , Cohen Fred E.
Published: July 2002

Recent progress determining the structure of the host-encoded prion protein (PrPC) and the role of auxiliary molecules in prion replication permits a more rational approach in the development of therapeutic interventions. Our objective is to identify a new class of lead compounds that mimic the dominant negative PrPCmutants, which inhibit an abnormal isoform (PrPSc) formation. A computational search was conducted on the Available Chemicals Directory for molecules that mimic both the spatial orientation and basic polymorphism of PrP residues 168, 172, 215, and 219, which confer dominant negative inhibition. The search revealed 1,000 potential candidates that were visually analyzed with respect to the structure of this four-residue epitope on PrPC. Sixty-three compounds were tested for inhibition of PrPScformation in scrapie-infected mouse neuroblastoma cells (ScN2a). Two compounds, Cp-60 (2-amino-6-[(2-aminophenyl)thio]-4-(2-furyl)pyridine-3,5-dicarbonitrile) and Cp-62 (N′1-({5-[(4,5-dichloro-1H-imidazol-1-yl)methyl]-2-furyl}carbonyl)-4 methoxybenzene-1-sulfonohydrazide), inhibited PrPScformation in a dose-dependent manner and demonstrated low levels of toxicity. A substructure search of the Available Chemicals Directory based on Cp-60 identified five related molecules, three of which exhibited activities comparable to Cp-60. Mimicking dominant negative inhibition in the design of drugs that inhibit prion replication may provide a more general approach to developing therapeutics for deleterious protein–protein interactions.
Regulation of protein function by native metastability
Authors: Lee Cheolju , Park Soon-Ho , Lee Min-Youn , Yu Myeong-Hee
Published: July 2002

In common globular proteins, the native form is in its most stable state. In contrast, each native form exists in a metastable state in inhibitory serpins (serine protease inhibitors) and some viral membrane fusion proteins. Metastability in these proteins is critical to their biological functions. Mutational analyses and structural examination have previously revealed unusual interactions, such as side-chain overpacking, buried polar groups, and cavities as the structural basis of the native metastability. However, the mechanism by which these structural defects regulate protein functions has not been elucidated. We report here characterization of cavity-filling mutations of α
1
-antitrypsin, a prototype serpin. Conformational stability of the molecule increased linearly with the van der Waals volume of the side chains. Increasing conformational stability is correlated with decreasing inhibitory activity. Moreover, the activity loss appears to correlate with the decrease in the rate of the conformational switch during complex formation with a target protease. These results strongly suggest that the native metastability of proteins is indeed a structural design that regulates protein functions.
The emergence of visual objects in space–time
Authors: Gepshtein Sergei , Kubovy Michael
Published: July 2002

It is natural to think that in perceiving dynamic scenes, vision takes a series of snapshots. Motion perception can ensue when the snapshots are different. The snapshot metaphor suggests two questions: (
i
) How does the visual system put together elements within each snapshot to form objects? This is the spatial grouping problem. (
ii
) When the snapshots are different, how does the visual system know which element in one snapshot corresponds to which element in the next? This is the temporal grouping problem. The snapshot metaphor is a caricature of the dominant model in the field—the sequential model—according to which spatial and temporal grouping are independent. The model we propose here is an interactive model, according to which the two grouping mechanisms are not separable. Currently, the experiments that support the interactive model are not conclusive because they use stimuli that are excessively specialized. To overcome this weakness, we created a new type of stimulus—spatiotemporal dot lattices—which allow us to independently manipulate the strength of spatial and temporal groupings. For these stimuli, sequential models make one fundamental assumption: if the spatial configuration of the stimulus remains constant, the perception of spatial grouping cannot be affected by manipulations of the temporal configuration of the stimulus. Our data are inconsistent with this assumption.
Kinetochore reproduction in animal evolution: Cell biological explanation of karyotypic fission theory
Authors: Kolnicki Robin L.
Published: July 2002

Karyotypic fission theory of Todd offers an explanation for the diverse range of diploid numbers of many mammalian taxa. Theoretically, a full complement of acrocentric chromosomes can be introduced into a population by chromosomal fission. Subsequent inheritance of ancestral chromosomes and paired fission derivatives potentially generates a diploid range from the ancestral condition to double its number of chromosomes. Although it is undisputed that both chromosomal fission and fusion (“Robertsonian rearrangements”) have significantly contributed to karyological diversity, it is generally assumed that independent events, the fission of single chromosomes or the fusion of two chromosomes, are the sources of such change. The karyotypic fission idea by contrast posits that all mediocentric chromosomes simultaneously fission. Here I propose a specific cell biological mechanism for Todd's karyotypic fission concept, “kinetochore reproduction theory,” where a complete set of dicentric chromatids is synthesized during gametogenesis, and kinetochore protein dephosphorylation regulates dicentric chromatid segregation. Three postulates of kinetochore reproduction theory are: (
i
) breakage of dicentric chromosomes between centromere pairs forms acrocentric derivatives, (
ii
)
de novo
capping of newly synthesized acrocentric ends with telomeric DNA stabilizes these derivatives, and (
iii
) mitotic checkpoints regulate chromosomal disjunction to generate fissioned karyotypes. Subsequent chromosomal rearrangement, especially pericentric inversion, increases the probability of genetic isolation amongst incipient sympatric species polytypic for fission-generated acrocentric autosomes. This mechanism obviates the requirement for numerous independent Robertsonian rearrangements and neatly accounts for mammalian karyotype evolution as exemplified in analyses of Carnivora, Artiodactyla, and Primates.
Visuo-tactile cross-modal associations in cortical somatosensory cells
Authors: Zhou Yong-Di , Fuster Joaquín M.
Published: July 2002

Recent studies show that cells in the somatosensory cortex are involved in the short-term retention of tactile information. In addition, some somatosensory cells appear to retain visual information that has been associated with the touch of an object. The presence of such cells suggests that nontactile stimuli associated with touch have access to cortical neuron networks engaged in the haptic sense. Thus, we inferred that somatosensory cells would respond to behaviorally associated visual and tactile stimuli. To test this assumption, single units were recorded from the anterior parietal cortex (Brodmann's areas 3a, 3b, 1, and 2) of monkeys performing a visuo-haptic delay task, which required the memorization of a visual cue for a tactile choice. Most cells responding to that cue responded also to the corresponding object presented for tactile choice. Significant correlations were observed in some cells between their differential reactions to tactile objects and their differential reactions to the associated visual cues. Some cells were recorded in both the cross-modal task and a haptic unimodal task, where the animal had to retain a tactile cue for a tactile choice. In most of these cells, correlations were observed between stimulus-related firing in corresponding cue periods of the two tasks. These findings suggest that cells in somatosensory cortex are the components of neuronal networks representing tactile information. Associated visual stimuli may activate such networks through visuo-haptic associations established by behavioral training.
Independent adaptation to riverine habitats allowed survival of ancient cetacean lineages
Authors: Cassens Insa , Vicario Saverio , Waddell Victor G. , Balchowsky Heather , Van Belle Daniel , Ding Wang , Fan Chen , Mohan R. S. Lal , Simões-Lopes Paulo C. , Bastida Ricardo , Meyer Axel , Stanhope Michael J. , Milinkovitch Michel C.
Published: July 2002

The four species of “river dolphins” are associated with six
separate great river systems on three subcontinents and have been
grouped for more than a century into a single taxon based on their
similar appearance. However, several morphologists recently questioned
the monophyly of that group. By using phylogenetic analyses of
nucleotide sequences from three mitochondrial and two nuclear genes, we
demonstrate with statistical significance that extant river dolphins
are not monophyletic and suggest that they are relict species whose
adaptation to riverine habitats incidentally insured their survival
against major environmental changes in the marine ecosystem or the
emergence of Delphinidae.
Dynamics and folding of single two-stranded coiled-coil peptides studied by fluorescent energy transfer confocal microscopy
Authors: Talaga David S. , Lau Wai Leung , Roder Heinrich , Tang Jianyong , Jia Yiwei , DeGrado William F. , Hochstrasser Robin M.
Published: July 2002

We report single-molecule measurements on the folding and
unfolding conformational equilibrium distributions and dynamics of a
disulfide crosslinked version of the two-stranded coiled coil from
GCN4. The peptide has a fluorescent donor and acceptor at the N termini
of its two chains and a Cys disulfide near its C terminus. Thus,
folding brings the two N termini of the two chains close together,
resulting in an enhancement of fluorescent resonant energy transfer.
End-to-end distance distributions have thus been characterized under
conditions where the peptide is nearly fully folded (0 M urea),
unfolded (7.4 M urea), and in dynamic exchange between folded and
unfolded states (3.0 M urea). The distributions have been compared for
the peptide freely diffusing in solution and deposited onto aminopropyl
silanized glass. As the urea concentration is increased, the mean
end-to-end distance shifts to longer distances both in free solution
and on the modified surface. The widths of these distributions indicate
that the molecules are undergoing millisecond conformational
fluctuations. Under all three conditions, these fluctuations gave
nonexponential correlations on 1- to 100-ms time scale. A component of
the correlation decay that was sensitive to the concentration of urea
corresponded to that measured by bulk relaxation kinetics. The
trajectories provided effective intramolecular diffusion coefficients
as a function of the end-to-end distances for the folded and unfolded
states. Single-molecule folding studies provide information concerning
the distributions of conformational states in the folded, unfolded, and
dynamically interconverting states.
In vitro
cloning of complex mixtures of DNA on microbeads: Physical separation of differentially expressed cDNAs
Authors: Brenner Sydney , Williams Steven R. , Vermaas Eric H. , Storck Thorsten , Moon Keith , McCollum Christie , Mao Jen-I , Luo Shujun , Kirchner James J. , Eletr Sam , DuBridge Robert B. , Burcham Timothy , Albrecht Glenn
Published: July 2002

We describe a method for cloning nucleic acid molecules onto the surfaces of 5-μm microbeads rather than in biological hosts. A unique tag sequence is attached to each molecule, and the tagged library is amplified. Unique tagging of the molecules is achieved by sampling a small fraction (1%) of a very large repertoire of tag sequences. The resulting library is hybridized to microbeads that each carry ≈10
6
strands complementary to one of the tags. About 10
5
copies of each molecule are collected on each microbead. Because such clones are segregated on microbeads, they can be operated on simultaneously and then assayed separately. To demonstrate the utility of this approach, we show how to label and extract microbeads bearing clones differentially expressed between two libraries by using a fluorescence-activated cell sorter (FACS). Because no prior information about the cloned molecules is required, this process is obviously useful where sequence databases are incomplete or nonexistent. More importantly, the process also permits the isolation of clones that are expressed only in given tissues or that are differentially expressed between normal and diseased states. Such clones then may be spotted on much more cost-effective, tissue- or disease-directed, low-density planar microarrays.
Inversion of the chordate body axis: Are there alternatives?
Authors: Gerhart J.
Published: July 2002

One major morphological difference between chordates and annelids
or arthropods is the opposite orientation of the nerve cord and heart.
A long-standing proposal is that the chordate axis evolved by inverting
the body of an ancestor with the annelid/arthropod orientation.
However, the data can also be explained by a common ancestor with
diffuse dorsoventral organization, followed by oppositely directed
condensation of the nerve cord and relocation of the heart in the two
lines.
A TSG101/MDM2 regulatory loop modulates MDM2 degradation and MDM2/p53 feedback control
Authors: Li Limin , Liao Jian , Ruland Jürgen , Mak Tak W. , Cohen Stanley N.
Published: July 2002

The p53 tumor suppressor protein and the MDM2 oncoprotein form a
feedback-control loop that up-regulates cellular MDM2 production,
blocks p53 activity, and promotes p53 decay.
tsg101
was
discovered as a gene whose deficiency results in neoplastic
transformation of NIH 3T3 cells and the ability to generate metastatic
tumors in nude mice. Its protein product contains a domain, Ubc,
characteristic of the catalytic domain of ubiquitin conjugase (E2)
enzymes but lacking an active-site cysteine crucial for ubiquitin
conjugase activity. Here we report that TSG101 participates with MDM2
in an autoregulatory loop that modulates the cellular levels of both
proteins, and also of p53, by affecting protein decay. We show that the
Ubc domain of TSG101 interferes with ubiquitination of MDM2, that
TSG101 inhibits MDM2 decay and elevates its steady-state level, and
that these events are associated with down-regulation of p53 protein.
Conversely, pulse–chase and Western blot experiments in wild-type and
mutant fibroblasts indicate that elevation of MDM2 by overexpression of
wild-type p53, by amplification of the endogenous
MDM2
gene, or by transfection of MDM2-expressing
constructs promotes TSG101 loss, which we show occurs by 26S
proteasome-dependent decay. Our results identify TSG101 as both a
regulator of, and target of, MDM2/p53 circuitry.
Religion and attitudes towards nature in Britain
Authors: Hayes Bernadette G. , Marangudakis Manussos
Published: July 2002

Religious institutions have been identified as important conduits in shaping social attitudes toward nature and the environment. Using Lynn White's historical thesis that Judeo‐Christianity has cherished the domination of nature (‘dominion’ belief) by humans as our frame of reference, this article examines the impact of religion, specifically Abrahamic and Judeo–Christian beliefs, on environmental attitudes in Britain. Based on the 1993 British Social Attitudes Survey, a nationally representative sample of the adult population in Britain, the multivariate results of this paper suggest that: (a) there is no significant difference between Christians and non‐Christians concerning environmental attitudes; (b) Roman Catholics are the most sceptic toward nature among Christian denominations; and (c) irrespective of religious identification, the two most notable and consistent factors in determining prodominion attitudes in Britain are educational attainment and particularly levels of scientific knowledge about the natural environment.
From disorganized capitalism to transnational fine tuning?: recent trends in wage development, industrial relations, and ‘work’ as a sociological category
Authors: Hasse Raimund , Leiulfsrud Håkon
Published: July 2002

The disorganization thesis concentrates upon globalization and market dynamics, which are believed to trigger the breakdown of any kind of institutional structures. The diversity of capitalism approach, by contrast, places much emphasis on the persistence of distinct paths of national economies. Referring to comparative data from the OECD and other sources it is shown that some variables indicate a robustness of national styles of capitalism. Others hint at resemblance: e.g.there is a striking synchronization of the overall and sectoral wage development, there is a significant decrease in industrial disputes, and the class composition tends to become more similar. A move beyond the disorganization thesis and diversity of capitalism approach is suggested. Special attention should be paid to the profound impacts of transnational institutions and knowledge carriers in the form of experts and guidelines.
The Impact of Dwelling Quality and Neighbourhood Quality on the Estimation of the Income Elasticity of Demand for Rental Housing in Jeddah, Saudi Arabia
Authors: Maatoug Gandil Mohammad
Published: July 2002

This study provides new evidence on the estimation of the income elasticity of demand for rental housing based on individual interviews with 405 households. It looks specifically at the impact of both dwelling and neighbourhood quality aspects on the evaluation of this elasticity. The study finds that the income elasticity of demand for rental housing in Jeddah is 0.519. When the sample is restricted to include Saudi households only, the elasticity estimate is 0.465. These values are within the range of previous empirical estimates. The results also indicate that for a range of various quality measures, the income elasticity is estimated to be less than unity. But, our suggested quality aspects were found to exert sizeable differences on the magnitude of this elasticity for the full sample (from 0.351 to 0.802) as well as for the Saudi sample (from 0.382 to 0.603). Stratified income elasticity estimates for Saudis were also found to be generally less than those for the whole sample.
National numeracy initiatives in England and Wales: a comparative study of policy
Authors: Jones Dylan V.
Published: July 2002

The introduction of the National Numeracy Strategy (NNS) in England has had a very significant effect on the teaching of numeracy in English primary schools. The NNS, however, does not apply in Wales where the Welsh Office decided to adopt a less prescriptive approach and encouraged local education authorities (LEAs), in partnership with their schools, to develop their own, locally based, numeracy initiatives. This article outlines the way in which recent numeracy policies have developed in England and Wales and compares their main strands. The article suggests that the decision not to adopt the NNS in Wales was potentially one of the most significant educational policy decisions taken in Cardiff during recent years. The article also highlights the fact that these parallel approaches offer an ideal opportunity for evaluating the impact and effectiveness of such alternative strategies as levers for large‐scale reform, and that such examination could usefully inform future policy‐making both within and beyond the curricular and geographical limits of the contexts considered here.
Beyond Living with Drugs: an interview with Mike Gossop
Authors: ALLSOP STEVE J.
Published: July 2002

Living with Drugs [1] has now been printed in four editions, a rare achievement for any text in the drug field. Many consider the book, which clearly enunciates the history, debate and controversies in the drug field, to be an essential read for researchers, clinicians, educators and students. Gossop was recently in Australia as a National Drug Strategy Visiting Fellow. This provided the opportunity to explore the author's views on a number of key policy, clinical and research issues.
Alcohol dehydrogenase ADH2–1 and ADH2–2 allelic isoforms in the Russian population correlate with type of alcoholic disease
Authors: Ogurtsov Pavel P. , Garmash Irina V. , Miandina Galina I. , Guschin Alexander E. , Itkes Alexander V. , Moiseev Valentin S.
Published: July 2002

The frequency ADH2‐2 allele in the Moscow urban population and a correlation between the ADH2‐2 allele, alcoholic dependence without cirrhosis, symptomatic alcoholic cirrhosis and status on hepatitis B and C infection have been studied. One hundred and twenty‐three inhabitants of Moscow (50 healthy donors, 36 patients with alcoholic cirrhosis (subdivided into infected and uninfected by HBV and/or HCV) and 37 patients with alcoholic dependence) of a similar age/sex and drinking pattern have been analysed. The frequency of 41% for ADH2‐2 allele is characteristic for an urban Moscow population. This value is intermediate between that found for Asian peoples and for Central and Western Europe. There is a negative correlation between the ADH2‐2 allele and alcohol misuse (both alcoholic dependence and alcoholic cirrhosis). This correlation is expressed more in alcoholic dependence. In spite of the possession of the ADH2‐2 allele (or genotype ADH2‐1/2), alcohol misuse increases the risk of cirrhosis. At the same time, positive status for active hepatitis B, C or combined infection B + C (replication markers HBV‐DNA or HCV‐RNA) increases the risk for symptomatic alcoholic cirrhosis in alcohol abusing patients, independently of ADH2 genotype.
Concentrations of transferrin and carbohydrate‐deficient transferrin in postmortem human brain from alcoholics
Authors: DODD PETER R. , ECKERT ALLISON L. , FLETCHER LINDA M. , KRIL JILLIAN J. , HARPER CLIVE G. , HALLIDAY JUNE W.
Published: July 2002

Transferrin (T f) and its carbohydrate‐deficient isoform (CDT) were measured by radioimmunoassay in phosphate‐buffered saline extracts of two informative areas of cerebral cortex tissue obtained at autopsy from alcoholics without other associated disease (n = 4); alcoholics with cirrhosis of the liver (n = 4) and agematched controls (n = 4). Total T f was also measured in two informative cortical areas from five dementia cases. All cases were male. Total immunoreactive T f was assayed directly in the extract, CDT immunoreactivity in the concentrated eluate after the sialylated form was removed by passing through DEAE‐Sephacel at pH 5.65. Brain CDT averaged 10% of total T f overall. Although replicate extractions of individual samples gave consistent assays for both substances, there was wide variation both between different cortical areas from a given case and between cases within groups. There were no significant differences between total T f levels in uncomplicated alcoholics, dementia cases and controls, but cirrhotic alcoholics gave significantly higher values. The CDT: T f ratio was not increased in the brains of either group of alcoholics compared to controls. Whereas the serum CDT: T f ratio is an excellent marker of recent alcohol consumption, brain T f and CDT concentrations do not mark alcoholism nor dementia, and their biological variability diminishes their usefulness as disease indices. However, brain T f may be a marker of cirrhosis‐induced changes.
Frequency of the A1/A2 alleles of the D2 dopamine receptor (DRD2) gene in a British, Caucasian control group screened to exclude alcoholism and heavy drinking
Authors: TURNER ADRIAN , LAWRENCE JACOB , CHIH‐HUI CHEN ANDREW , COOK CHRISTOPHER , GURLING HUGH
Published: July 2002

Abstract Recent reports of a genetic association between restriction fragment length polymorphisms at the D2 dopamine receptor locus (DRD2) on chromosome 11q and alcoholism have suggested involvement of the D2 receptor protein in the aetiology of alcoholism, smoking and possibly other disorders. These allelic association findings have been criticized on the basis of the possible confounding of ethnic with disease differences, between allele frequencies of subjects and controls, and the fact that some of the control groups were not screened to exclude alcoholism or heavy drinking. We have observed the frequency of the A1 and A2 alleles in a population of 307 Caucasian British individuals screened for alcohol consumption of less than the UK Royal College of Psychiatrists’ recommended sensible drinking limits for males and females and who also failed to qualify for a diagnosis of alcoholism according to the Research Diagnostic Criteria (RDC). The frequency of the A1 allele was found to be 0.20, which is slightly higher than most of the other screened Caucasian control groups from Europe and the United States. The allele and genotype frequencies in our sample are a resource for comparison with samples of alcoholics from the United Kingdom which have been selected on the basis of British ancestry and for residence in London. When we combined our new control data with that of the previous Caucasian control samples we found a significantly higher frequency of A1A1 homozygotes among the unscreened than the screened controls, suggesting that the DRD2 locus may be involved in drinking variation among the general population.
Kinetic Characteristics of ATP Hydrolysis by a Detergent‐Solubilized Alkaline Phosphatase From Rat Osseous Plate
Authors: Demenis Marlene A. , Leone Francisco A.
Published: July 2002

Polidocanol‐solubilized alkaline phosphatase was purified to homogeneity with a specific activity of 822.3 U/mg. In the absence of Mg2+ and Ca2+ ions and at pH 9.4, the enzyme hydrolyzed ATP in a manner that could be represented by biphasic curves with V = 94.3 U/mg, K0.5 = 17.2 muM, and n = 1.8 and V = 430.3 U/mg, K0.5 = 3.2 muM, and n = 3.2 for high‐ and low‐affinity sites, respectively. In the presence of saturating concentrations of Mg2+ or Ca2+ ions, the hydrolysis of ATP also followed biphasic curves. However, the specific activity increased to as much as 1,000 U/mg, whereas the K0.5 and n values remained almost unchanged. In the presence of nonsaturating concentrations of metal ions, the hydrolysis of ATP was similar to that observed in the absence of these ions, but with a marked decrease in K0.5 values. At pH 7.5, the enzyme also hydrolyzed ATP with K0.5 = 8.1 muM and V = 719.8 U/mg. Apparently, alkaline phosphatase was able to hydrolyze ATP in vivo, either at pH 7.5 or pH 9.4. These data contribute to the knowledge of the biological properties of skeletal alkaline phosphatase and suggest that this enzyme may have a high‐affinity binding site for ATP at alkaline pH.
Identification of a Novel Marker for Primordial Smooth Muscle and Its Differential Expression Pattern in Contractile vs Noncontractile Cells
Authors: Hungerford Jill E. , Hoeffler James P. , Bowers Chauncey W. , Dahm Lisa M. , Falchetto Rocco , Shabanowitz Jeffrey , Hunt Donald F. , Little Charles D.
Published: July 2002

The assembly of the vessel wall from its cellular and extracellular matrix components is an essential event in embryogenesis. Recently, we used the descending aorta of the embryonic quail to define the morphological events that initiate the formation of a multilayered vessel wall from a nascent endothelial cell tube (Hungerford, J.E., G.K. Owens, W.S. Argraves, and C.D. Little. 1996. Dev. Biol. 178:375–392). We generated an mAb, 1E12, that specifically labels smooth muscle cells from the early stages of development to adulthood. The goal of our present study was to characterize further the 1E12 antigen using both cytological and biochemical methods. The 1E12 antigen colocalizes with the actin cytoskeleton in smooth muscle cells grown on planar substrates in vitro; in contrast, embryonic vascular smooth muscle cells in situ contain 1E12 antigen that is distributed in threadlike filaments and in cytoplasmic rosette-like patterns. Initial biochemical analysis shows that the 1E12 mAb recognizes a protein, Mr = 100,000, in lysates of adult avian gizzard. An additional polypeptide band, Mr = 40,000, is also recognized in preparations of lysate, when stronger extraction conditions are used. We have identified the 100-kD polypeptide as smooth muscle α-actinin by tandem mass spectroscopy analysis. The 1E12 antibody is an IgM isotype. To prepare a more convenient 1E12 immunoreagent, we constructed a single chain antibody (sFv) using recombinant protein technology. The sFv recognizes a single 100-kD protein in gizzard lysates. Additionally, the recombinant antibody recognizes purified smooth muscle α-actinin. Our results suggest that the 1E12 antigen is a member of the α-actinin family of cytoskeletal proteins; furthermore, the onset of its expression defines a primordial cell restricted to the smooth muscle lineage.
Antagonism of Cell Adhesion by an α-Catenin Mutant, and of the Wnt-signaling Pathway by α-Catenin in Xenopus Embryos
Authors: Sehgal Ravinder N.M. , Gumbiner Barry M. , Reichardt Louis F.
Published: July 2002

In Xenopus laevis development, β-catenin plays an important role in the Wnt-signaling pathway by establishing the Nieuwkoop center, which in turn leads to specification of the dorsoventral axis. Cadherins are essential for embryonic morphogenesis since they mediate calcium-dependent cell–cell adhesion and can modulate β-catenin signaling. α-catenin links β-catenin to the actin-based cytoskeleton. To study the role of endogenous α-catenin in early development, we have made deletion mutants of αN-catenin. The binding domain of β-catenin has been mapped to the NH2-terminal 210 amino acids of αN-catenin. Overexpression of mutants lacking the COOH-terminal 230 amino acids causes severe developmental defects that reflect impaired calcium-dependent blastomere adhesion. Lack of normal adhesive interactions results in a loss of the blastocoel in early embryos and ripping of the ectodermal layer during gastrulation. The phenotypes of the dominant-negative mutants can be rescued by coexpressing full-length αN-catenin or a mutant of β-catenin that lacks the internal armadillo repeats.

We next show that coexpression of αN-catenin antagonizes the dorsalizing effects of β-catenin and Xwnt-8. This can be seen phenotypically, or by studying the effects of expression on the downstream homeobox gene Siamois. Thus, α-catenin is essential for proper morphogenesis of the embryo and may act as a regulator of the intracellular β-catenin signaling pathway in vivo.