bluuebunny/crossref_metadata_2025 huggingface dataset:
Sliding life enhancement of a WS2 sputtered film by ion beam mixing
Authors: Hirano Motohisa , Miyake Shojiro
Published: July 2002

The ion beam mixing effect on the tribological properties of the WS2 sputtered film is investigated. The film morphology and crystal structure related to the tribological properties are also examined. It is determined that a film so prepared has both lower friction and longer sliding life than an as-sputtered WS2 film. Scanning electron micrograph and reflection high-energy electron diffraction examinations suggest that the ion beam mixing effect can be attributed to crystalline precipitate formation resulting from recrystallization caused by the implantation.
New approach to the high quality epitaxial growth of lattice-mismatched materials
Authors: Luryi Serge , Suhir Ephraim
Published: July 2002

We have reconsidered the problem of the critical layer thickness hc for growth of strained heterolayers on lattice-mismatched substrates, using a new approach which allows us to determine the spatial distribution of stresses in a bi-material assembly and include the effects of a finite size of the sample. The possibility of dislocation-free growth of lattice-mismatched materials on porous silicon substrates is discussed as an example of a more general problem of heteroepitaxial growth on small seed pads of lateral dimension l, having a uniform crystal orientation over the entire substrate wafer. It turns out that for a given mismatch f, the critical film thickness hlc strongly depends on l, rising sharply when the latter is sufficiently small, l≲lmin. The characteristic size lmin( f ) below which, effectively, hlc( f )→∞, is determined in terms of the experimentally known (or calculated for growth on a monolithic substrate) function h∞c( f )≡hc( f ). When l≲lmin, then the entire elastic stress in the epitaxial film will be accommodated without dislocations.
Room-temperature operation of InGaAsSb/AlGaSb double heterostructure lasers near 2.2 μm prepared by molecular beam epitaxy
Authors: Chiu T. H. , Tsang W. T. , Ditzenberger J. A. , van der Ziel J. P.
Published: July 2002

In0.16Ga0.84As0.15Sb0.85/Al0.35Ga0.65Sb double heterostructure injection lasers have been grown by molecular beam epitaxy on (100) GaSb substrates. Room-temperature operation near 2.2 μm wavelength has been achieved under pulsed conditions. Low pulsed threshold current density of 4.2 kA/cm2 and a characteristic temperature of T0∼26 K have been obtained.
Electro-optic and dielectric properties of KTiOPO4
Authors: Bierlein J. D. , Arweiler C. B.
Published: July 2002

The electro-optic and dielectric properties of potassium titanyl phosphate (KTP) have been measured from dc to 1 GHz and are compared with other electro-optic materials. KTP is shown to possess a combination of properties that make it unique for a variety of electro-optic modulator applications.
Super-resolution fluorescence near-field scanning optical microscopy
Authors: Harootunian A. , Betzig E. , Isaacson M. , Lewis A.
Published: July 2002

A one-dimensional near-field scanning optical microscope (NSOM) operating in the fluorescence mode has been demonstrated. NSOM line scans of both metallic edges and fluorescent gratings have been obtained and quantitatively compared to both scanning electron micrographs and conventional optical micrographs of the same structures. The sharpness of the near-field scans indicates resolution of <100 nm.
Superconducting NbNxCy thin films fabricated with a dual ion-beam sputtering method
Authors: Lin L.-J. , Prober D. E.
Published: July 2002

We have fabricated refractory superconducting NbNxCy thin films on unheated Si substrates with a low-energy dual ion-beam sputtering technique. Films fabricated with this technique have predominantly B1 crystal structure with maximum Tc ∼13.2 K, resistivity of 80–150 μΩ cm and residual resistance ratio ∼1.0. The use of low ion-beam energies and the absence of substrate heating make this method well suited for producing NbNxCy films for superconducting microelectronic applications.
Thermal expansion and elastic properties of plasma-deposited amorphous silicon and silicon oxide films
Authors: Jansen F. , Machonkin M. A. , Palmieri N. , Kuhman D.
Published: July 2002

The thermomechanical properties of amorphous glow-discharge deposited silicon and silicon oxide films were measured as a function of film composition. Hydrogenated amorphous silicon films, deposited at 250 °C with a density of 2.0 g/cm3 and 20 at. % of hydrogen, exhibit a linear thermal expansion coefficient of about 4.4×10−6/°C and a biaxial elastic modulus of 150 GPa. The expansion coefficient of plasma-deposited silicon oxide films shows a systematic dependence on the fabrication conditions and ranges from about 10−5/°C to negative values. Strong correlations between the hydrogen concentration of the films, the film density, and thermal expansion coefficient were observed and are discussed. The biaxial elastic modulus of the oxide films is not strongly dependent on the gas ratio and is about 40 GPa, substantially smaller than the values obtained for thermally grown oxides. These differences are attributed to the effect of network terminations by hydrogen and −OH groups.
Observation of tilt boundaries in graphite by scanning tunneling microscopy and associated multiple tip effects
Authors: Albrecht T. R. , Mizes H. A. , Nogami J. , Park Sang-il , Quate C. F.
Published: July 2002

Tilt boundaries have been observed on the (0001) surface of graphite by scanning tunneling microscopy (STM). Rotation angles about the c axis of 6.5°, 8°, and 19° were found, indicating no preferential orientation of grains in the basal plane of graphite. The grain boundary region between crystallites appears disordered with a width varying between 10 and 100 Å. Moiré patterns are observed near grain boundaries when multiple tips scanning over different grains contribute to the image simultaneously. Such images support the theory that multiple isolated tips, occasionally hundreds of angstroms apart, can contribute to STM images.
The B2 element of the
Saccharomyces cerevisiae
ARS1
origin of replication requires specific sequences to facilitate pre-RC formation
Authors: Wilmes Gwendolyn M. , Bell Stephen P.
Published: July 2002

The minimal requirements for a eukaryotic origin of
replication are an initiator binding site and a region of helically
unstable DNA [DNA unwinding element (DUE)]. Budding yeast origins
consist of modular elements, and one of these elements, B2, has been
proposed to act as a DUE. To test this hypothesis, we screened for
sequences that function at the B2 element of
ARS1
. We
found that the B2 element required A-rich sequences, but that the
function of these identified sequences did not correlate with helical
instability. Instead, the sequences that substituted fully for B2
function showed similarity to the ARS consensus sequence (ACS). The ACS
is the binding site for the initiator origin recognition complex (ORC),
but the selected sequences are not strong ORC binding sites
in
vitro
. Nonfunctional B2 sequences show a corresponding loss in
Mcm2-7p origin association. The function of these mutant sequences is
rescued by Cdc6p overexpression. We propose that the B2 element
requires specific sequences to bind a component of the pre-RC.
Expression of a splice variant of the receptor for GHRH in 3T3 fibroblasts activates cell proliferation responses to GHRH analogs
Authors: Kiaris Hippokratis , Schally Andrew V. , Busto Rebeca , Halmos Gabor , Artavanis–Tsakonas Spyros , Varga Jozsef L.
Published: July 2002

The stimulatory effects of growth hormone-releasing hormone (GHRH)
and the antiproliferative action of GHRH antagonists have been
demonstrated in various cancers, but the receptors that mediate these
responses are not clearly identified. Recently, we reported that
human cancer cell lines express splice variants (SVs) of the receptors
for GHRH. SV1 exhibits the greatest similarity to the pituitary GHRH
receptor and is most likely to be functional. To ascertain whether SV1
mediates mitogenic effects on nonpituitary tissues, we expressed SV1 in
3T3 mouse fibroblasts and studied the properties of the transfected
cells. Radioligand binding assays with
125
I-labeled GHRH
antagonist JV-1–42 detected high affinity
(
K
d
= 0.58 ± 0.17 nM) binding
sites for GHRH with a maximal binding capacity
(
B
max
) of 103 ± 17.4 fmol/mg of
membrane protein in 3T3 cells transfected with pcDNA3-SV1, whereas the
control cells transfected with the empty vector did not show any GHRH
binding. Cell proliferation studies showed that cells expressing SV1
are much more sensitive to GHRH analogs than the pcDNA3 controls. Thus,
the expression of SV1 augments the stimulatory responses to
GHRH(1–29)NH
2
or GHRH agonist JI-38 and inhibitory
responses to GHRH antagonist JV-1–38 as compared with pcDNA3 controls.
The stimulation of SV1-expressing cells by GHRH or JI-38 is followed by
an increase in cAMP production, but no GH release occurs. Vasoactive
intestinal peptide had no effect, and its antagonist JV-1–53 did not
inhibit the proliferation of SV1-expressing cells stimulated by GHRH.
Our results suggest that SV1 could mediate responses of nonpituitary
cells and various tumors to GHRH and GHRH antagonists. The presence of
SV1 in several human cancer cell lines provides a rationale for
antitumor therapy based on the blockade of this receptor by specific
GHRH antagonists.
HOX
genes in the sepiolid squid
Euprymna scolopes
: Implications for the evolution of complex body plans
Authors: Callaerts Patrick , Lee Patricia N. , Hartmann Britta , Farfan Claudia , Choy Darrett W. Y. , Ikeo Kazuho , Fischbach Karl-Friedrich , Gehring Walter J. , de Couet H. Gert
Published: July 2002

Molluscs display a rich diversity of body plans ranging from the wormlike appearance of aplacophorans to the complex body plan of the cephalopods with highly developed sensory organs, a complex central nervous system, and cognitive abilities unrivaled among the invertebrates. The aim of the current study is to define molecular parameters relevant to the developmental evolution of cephalopods by using the sepiolid squid
Euprymna scolopes
as a model system. Using PCR-based approaches, we identified one anterior, one paralog group 3, five central, and two posterior group
Hox
genes. The deduced homeodomain sequences of the
E. scolopes
Hox
cluster genes are most similar to known annelid, brachiopod, and nemertean
Hox
gene homeodomain sequences. Our results are consistent with the presence of a single
Hox
gene cluster in cephalopods. Our data also corroborate the proposed existence of a differentiated
Hox
gene cluster in the last common ancestor of Bilaterians. Furthermore, our phylogenetic analysis and in particular the identification of
Post
-
1
and
Post
-
2
homologs support the Lophotrochozoan clade.
Calmodulin kinase is a molecular switch for cardiac excitation –contraction coupling
Authors: Wu Yuejin , Colbran Roger J. , Anderson Mark E.
Published: July 2002

Signaling between cell membrane-bound L-type Ca
2+
channels (LTCC) and ryanodine receptor Ca
2+
release
channels (RyR) on sarcoplasmic reticulum (SR) stores grades
excitation–contraction coupling (ECC) in striated muscle. A physical
connection regulates LTCC and RyR in skeletal muscle, but the molecular
mechanism for coordinating LTCC and RyR in cardiomyocytes, where this
physical link is absent, is unknown. Calmodulin kinase
(CaMK) has characteristics suitable for an ECC coordinating molecule:
it is activated by Ca
2+
/calmodulin, it
regulates LTCC and RyR, and it is enriched in the vicinity of LTCC and
RyR. Intact cardiomyocytes were studied under conditions where CaMK
activity could be controlled independently of intracellular
Ca
2+
by using an engineered Ca
2+
-independent
form of CaMK and a highly specific CaMK inhibitory peptide. CaMK
reciprocally enhanced L-type Ca
2+
current and reduced
release of Ca
2+
from the SR while increasing SR
Ca
2+
content. These findings support the hypothesis that
CaMK is required to functionally couple LTCC and RyR during cardiac
ECC.
A Gaussian-chain model for treating residual charge–charge interactions in the unfolded state of proteins
Authors: Zhou Huan-Xiang
Published: July 2002

Characterization of the unfolded state is essential for understanding the protein folding problem. In the unfolded state, a protein molecule samples vastly different conformations. Here I present a simple theoretical method for treating residual charge–charge interactions in the unfolded state. The method is based on modeling an unfolded protein as a Gaussian chain. After sampling over all conformations, the electrostatic interaction energy between two charged residues (separated by
l
peptide bonds) is given by
W
= 332(6/π)
1/2
[1 −
π
1/2
x
exp(
x
2
)erfc(
x
)]/
ɛd
, where
d
=
bl
1/2
+
s
and
x
=
κd
/6
1/2
. In unfolded barnase, the residual interactions lead to downward pK
a
shifts of ≈0.33 unit, in agreement with experiment. pK
a
shifts in the unfolded state significantly affect pH dependence of protein folding stability, and the predicted effects agree very well with experimental results on barnase and four other proteins. For T4 lysozyme, the charge reversal mutation K147E is found to stabilize the unfolded state even more than the folded state (1.39 vs. 0.46 kcal/mol), leading to the experimentally observed result that the mutation is net destabilizing for the folding. The Gaussian-chain model provides a quantitative characterization of the unfolded state and may prove valuable for elucidating the energetic contributions to the stability of thermophilic proteins and the energy landscape of protein folding.
Headpiece domain of dematin is required for the stability of the erythrocyte membrane
Authors: Khanna Richie , Chang Seon H. , Andrabi Shaida , Azam Mohammad , Kim Anthony , Rivera Alicia , Brugnara Carlo , Low Philip S. , Liu Shih-Chun , Chishti Athar H.
Published: July 2002

Dematin is an actin-binding and bundling protein of the erythrocyte membrane skeleton. Dematin is localized to the spectrin–actin junctions, and its actin-bundling activity is regulated by phosphorylation of cAMP-dependent protein kinase. The carboxyl terminus of dematin is homologous to the “headpiece” domain of villin, an actin-bundling protein of the microvillus cytoskeleton. The headpiece domain contains an actin-binding site, a cAMP-kinase phosphorylation site, plays an essential role in dematin self-assembly, and bundles F-actin
in vitro
. By using homologous recombination in mouse embryonic stem cells, the headpiece domain of dematin was deleted to evaluate its function
in vivo
. Dematin headpiece null mice were viable and born at the expected Mendelian ratio. Hematological evaluation revealed evidence of compensated anemia and spherocytosis in the dematin headpiece null mice. The headpiece null erythrocytes were osmotically fragile, and ektacytometry/micropore filtration measurements demonstrated reduced deformability and filterability.
In vitro
membrane stability measurements indicated significantly greater membrane fragmentation of the dematin headpiece null erythrocytes. Finally, biochemical characterization, including the vesicle/cytoskeleton dissociation, spectrin self-association, and chemical crosslinking measurements, revealed a weakened membrane skeleton evidenced by reduced association of spectrin and actin to the plasma membrane. Together, these results provide evidence for the physiological significance of dematin and demonstrate a role for the headpiece domain in the maintenance of structural integrity and mechanical properties of erythrocytes
in vivo
.
Pillared honeycomb nanoarchitectures formed on solid surfaces by the self-assembly of lipid-packaged one-dimensional Pt complexes
Authors: Chang-Soo Lee , Kimizuka Nobuo
Published: July 2002

A highly lipophilic polyion complex [Pt(en)
2
][PtCl
2
(en)
2
](1)
4
(en, 1,2-diaminoethane) is prepared from one-dimensional mixed valence Pt
II
/Pt
IV
complex and newly designed chiral amphiphile 1. The powdery sample showed purple color, which is a result of the mixed valence absorption of the linear chlorobridged complex (Pt
II
-Cl-Pt
IV
-Cl-)
n
. When the lipid complex is dispersed in dichloromethane, purple-colored dispersion is obtained at 0°C, whereas the color disappears after heating the solution to 21°C. The observed thermochromism is reversible with respect to the temperature changes and is ascribed to the reversible dissociation and reassembly of the self-assembling inorganic wires. Casting of the 0°C-purple dispersion on solid substrates affords honeycomb nanostructures in addition to the nanowires with the width of about 20 nm. The honeycomb patterns seem to be templated by the condensed water droplets that are formed and aligned on the rapidly evaporating dichloromethane solution. On the other hand, more regular honeycomb structures are exclusively obtained by casting the 21°C-colorless solution. These observations indicate that the ordered honeycomb structures are obtainable on solid surfaces by the self-assembly of molecularly dispersed components [Pt(en)
2
](1)
2
and
trans
-[PtCl
2
(en)
2
](1)
2.
Very interestingly, formation of double-layered honeycomb nanostructure is observed by scanning electron microscopy. The unit hexagons and pillars of the honeycombs are made of nanowires that are hierarchically assembled from the lipid-packaged Pt
II
/Pt
IV
complexes. The surface self-organization of lipophilic inorganic complexes has a potential to fabricate novel nanoarchitectures with conjugated electronic structures.
Nuclear entry and CRM1-dependent nuclear export of the Rous sarcoma virus Gag polyprotein
Authors: Scheifele Lisa Z. , Garbitt Rachel A. , Rhoads Jonathan D. , Parent Leslie J.
Published: July 2002

The retroviral Gag polyprotein directs budding from the plasma membrane of infected cells. Until now, it was believed that Gag proteins of type C retroviruses, including the prototypic oncoretrovirus Rous sarcoma virus, were synthesized on cytosolic ribosomes and targeted directly to the plasma membrane. Here we reveal a previously unknown step in the subcellular trafficking of the Gag protein, that of transient nuclear localization. We have identified a targeting signal within the N-terminal matrix domain that facilitates active nuclear import of the Gag polyprotein. We also found that Gag is transported out of the nucleus through the CRM1 nuclear export pathway, based on observations that treatment of virus-expressing cells with leptomycin B resulted in the redistribution of Gag proteins from the cytoplasm to the nucleus. Internal deletion of the C-terminal portion of the Gag p10 region resulted in the nuclear sequestration of Gag and markedly diminished budding, suggesting that the nuclear export signal might reside within p10. Finally, we observed that a previously described matrix mutant, Myr1E, was insensitive to the effects of leptomycin B, apparently bypassing the nuclear compartment during virus assembly. Myr1E has a defect in genomic RNA packaging, implying that nuclear localization of Gag might be involved in viral RNA interactions. Taken together, these findings provide evidence that nuclear entry and egress of the Gag polyprotein are intrinsic components of the Rous sarcoma virus assembly pathway.
Prion protein: Evolution caught en route
Authors: Tompa P. , Tusnády G. E. , Cserző M. , Simon I.
Published: July 2002

The prion protein displays a unique structural ambiguity in that it
can adopt multiple stable conformations under physiological conditions.
In our view, this puzzling feature resulted from a sudden environmental
change in evolution when the prion, previously an integral membrane
protein, got expelled into the extracellular space. Analysis of known
vertebrate prions unveils a primordial transmembrane protein encrypted
in their sequence, underlying this relocalization hypothesis.
Apparently, the time elapsed since this event was insufficient to
create a “minimally frustrated” sequence in the new milieu,
probably due to the functional constraints set by the importance of the
very flexibility that was created in the relocalization. This scenario
may explain why, in a structural sense, the prion protein is still en
route toward becoming a foldable globular protein.
In deep trouble: Habitat selection constrained by multiple enemies in zooplankton
Authors: Decaestecker Ellen , De Meester Luc , Ebert Dieter
Published: July 2002

Habitat selection behavior is an important predator-avoidance strategy for many organisms. Its particular expression is often explained as the result of a tradeoff between avoiding antagonists and acquiring resources. However, there is need for a broader perspective on this behavior, as organisms are often simultaneously involved in complex antagonistic relationships with multiple types of enemies. We show experimentally that a tradeoff between predator and parasite avoidance may be important in the evolution of habitat selection behavior in the waterflea,Daphnia magna. In this species, negatively phototactic clones suffer less from visually hunting predators by residing in deeper and darker portions of the water column during the day. However, this behavior increases the risk of parasitic infections when theDaphniaare exposed to pond sediments containing parasite spores. Positively phototactic clones, which are at a higher risk of predation, are less exposed to parasite spores in the sediment and consequently suffer less from parasitic infection. We show that the increased risk of infection remains even if the animals change their phototactic behavior on exposure to chemical cues from fish. This tradeoff highlights a substantial cost of predator-induced changes in habitat selection behavior. Tradeoffs caused by multiple enemies may explain genetic polymorphism for habitat selection behavior in many natural populations.
Galactose metabolism is essential for the African sleeping sickness parasite
Trypanosoma brucei
Authors: Roper Janine R. , Güther Maria Lucia S. , Milne Kenneth G. , Ferguson Michael A. J.
Published: July 2002

The tsetse fly-transmitted protozoan parasite
Trypanosoma brucei
is the causative agent of human African sleeping sickness and the cattle disease Nagana. The bloodstream form of the parasite uses a dense cell-surface coat of variant surface glycoprotein to escape the innate and adaptive immune responses of the mammalian host and a highly glycosylated transferrin receptor to take up host transferrin, an essential growth factor. These glycoproteins, as well as other flagellar pocket, endosomal, and lysosomal glycoproteins, are known to contain galactose. The parasite is unable to take up galactose, suggesting that it may depend on the action of UDP-glucose 4′-epimerase for the conversion of UDP-Glc to UDP-Gal and subsequent incorporation of galactose into glycoconjugates via UDP-Gal-dependent galactosyltransferases. In this paper, we describe the cloning of
T. brucei galE
, encoding
T. brucei
UDP-Glc-4′-epimerase, and functional characterization by complementation of a
galE
-deficient
Escherichia coli
mutant and enzymatic assay of recombinant protein. A tetracycline-inducible conditional
galE
null mutant of
T. brucei
was created using a transgenic parasite expressing the
TETR
tetracycline repressor protein gene. Withdrawal of tetracycline led to a cessation of cell division and substantial cell death, demonstrating that galactose metabolism in
T. brucei
proceeds via UDP-Glc-4′-epimerase and is essential for parasite growth. After several days without tetracycline, cultures spontaneously recovered. These cells were shown to have undergone a genetic rearrangement that deleted the
TETR
gene. The results show that enzymes and transporters involved in galactose metabolism may be considered as potential therapeutic targets against African trypanosomiasis.
Molecular dynamics simulations of protein folding from the transition state
Authors: Gsponer Jörg , Caflisch Amedeo
Published: July 2002

Putative transition-state ensemble (TSE) conformations of src SH3 were identified by monitoring the deviation from the experimental φ values along molecular dynamics (MD) simulations of unfolding. Sixty MD trajectories (for a total of about 7 μs) were then started from the putative TSE. About one-half of the 60 runs reached the folded state while unfolding was observed in the remaining half of the runs. This result validates φ-value analysis as an approach to obtain structural information on the transition state. It also demonstrates that an atomic resolution description of the TSE can be extracted from MD simulations. All conformations in the TSE have the central three-stranded β-sheet formed in agreement with experimental data. An elongation of strand β2 as well as nonnative side-chain interactions between the diverging turn and the distal hairpin are observed. The simulation results indicate that the tight packing of the side chains between the diverging turn and the distal hairpin is a necessary condition for rapid folding. Contacts between residues in the most structured element of the TSE, the central β-sheet, are kinetically more important than those between the N- and C-terminal strands.
Altered pain responses in mice lacking α
1E
subunit of the voltage-dependent Ca
2+
channel
Authors: Saegusa Hironao , Kurihara Takashi , Zong Shuqin , Minowa Osamu , Kazuno An-a , Han Wenhua , Matsuda Yoshihiro , Yamanaka Hitomi , Osanai Makoto , Noda Tetsuo , Tanabe Tsutomu
Published: July 2002

α
1
subunit of the voltage-dependent Ca
2+
channel is essential for channel function and determines the functional specificity of various channel types. α
1E
subunit was originally identified as a neuron-specific one, but the physiological function of the Ca
2+
channel containing this subunit (α
1E
Ca
2+
channel) was not clear compared with other types of Ca
2+
channels because of the limited availability of specific blockers. To clarify the physiological roles of the α
1E
Ca
2+
channel, we have generated α
1E
mutant (α
1E
−/−) mice by gene targeting. The
lacZ
gene was inserted in-frame and used as a marker for α
1E
subunit expression. α
1E
−/− mice showed reduced spontaneous locomotor activities and signs of timidness, but other general behaviors were apparently normal. As involvement of α
1E
in pain transmission was suggested by localization analyses with 5-bromo-4-chloro-3-indolyl β-
d
-galactopyranoside staining, we conducted several pain-related behavioral tests using the mutant mice. Although α
1E
+/− and α
1E
−/− mice exhibited normal pain behaviors against acute mechanical, thermal, and chemical stimuli, they both showed reduced responses to somatic inflammatory pain. α
1E
+/− mice showed reduced response to visceral inflammatory pain, whereas α
1E
−/− mice showed apparently normal response compared with that of wild-type mice. Furthermore, α
1E
−/− mice that had been presensitized with a visceral noxious conditioning stimulus showed increased responses to a somatic inflammatory pain, in marked contrast with the wild-type mice in which long-lasting effects of descending antinociceptive pathway were predominant. These results suggest that the α
1E
Ca
2 +
channel controls pain behaviors by both spinal and supraspinal mechanisms.
Survival without recovery after mass extinctions
Authors: Jablonski David
Published: July 2002

Because many survivors of mass extinctions do not participate in postrecovery diversifications, and therefore fall into a pattern that can be termed “Dead Clade Walking” (DCW), the effects of mass extinctions extend beyond the losses observed during the event itself. Analyses at two taxonomic levels provide a first-order test of the prevalence of DCWs by using simple and very conservative operational criteria. For four of the Big Five mass extinctions of the Phanerozoic, the marinegenerathat survived the extinction suffered ≈10–20% attrition in the immediately following geologic stage that was significantly greater than the losses sustained in preextinction stages. The stages immediately following the three Paleozoic mass extinctions also account for 17% of allorder-level losses in marine invertebrates over that interval, which is, again, significantly greater than that seen for the other stratigraphic stages (no orders are lost immediately after the end-Triassic or end-Cretaceous mass extinctions). DCWs are not evenly distributed among four regional molluscan time-series following the end-Cretaceous extinction, demonstrating the importance of spatial patterns in recovery dynamics. Although biotic interactions have been invoked to explain the differential postextinction success of clades, such hypotheses must be tested against alternatives that include stochastic processes in low-diversity lineages—which is evidently not a general explanation for the ordinal DCW patterns, because postextinction fates are not related to the size of extinction bottlenecks in Paleozoic orders—and ongoing physical environmental changes.
Glucagon receptor activates extracellular signal-regulated protein kinase 1/2 via cAMP-dependent protein kinase
Authors: Jiang Youwei , Cypess Aaron M. , Muse Evan D. , Wu Cui-Rong , Unson Cecilia G. , Merrifield R. B. , Sakmar Thomas P.
Published: July 2002

We prepared a stable cell line expressing the glucagon receptor to characterize the effect of G
s
-coupled receptor stimulation on extracellular signal-regulated protein kinase 1/2 (ERK1/2) activity. Glucagon treatment of the cell line caused a dose-dependent increase in cAMP concentration, activation of cAMP-dependent protein kinase (PKA), and transient release of intracellular calcium. Glucagon treatment also caused rapid dose-dependent phosphorylation and activation of mitogen-activated protein kinase kinase/ERK kinase (MEK1/2) and ERK1/2. Inhibition of either PKA or MEK1/2 blocked ERK1/2 activation by glucagon. However, no significant activation of several upstream activators of MEK, including Ras, Rap1, and Raf, was observed in response to glucagon treatment. In addition, chelation of intracellular calcium reduced glucagon-mediated ERK1/2 activation. In transient transfection experiments, glucagon receptor mutants that bound glucagon but failed to increase intracellular cAMP and calcium concentrations showed no glucagon-stimulated ERK1/2 phosphorylation. We conclude that glucagon-induced MEK1/2 and ERK1/2 activation is mediated by PKA and that an increase in intracellular calcium concentration is required for maximal ERK activation.
Inhibition of proteasomal degradation by the Gly-Ala repeat of Epstein–Barr virus is influenced by the length of the repeat and the strength of the degradation signal
Authors: Dantuma Nico P. , Heessen Stijn , Lindsten Kristina , Jellne Marianne , Masucci Maria G.
Published: July 2002

The Gly-Ala repeat (GAr) of the Epstein–Barr virus nuclear antigen-1 is a transferable element that inhibits
in cis
ubiquitin/proteasome-dependent proteolysis. We have investigated this inhibitory activity by using green fluorescent protein-based reporters that have been targeted for proteolysis by N end rule or ubiquitin-fusion degradation signals, resulting in various degrees of destabilization. Degradation of the green fluorescent protein substrates was inhibited on insertion of a 25-aa GAr, but strongly destabilized reporters were protected only partially. Protection could be enhanced by increasing the length of the repeat. However, reporters containing the Ub-R and ubiquitin-fusion degradation signals were degraded even in the presence of a 239-aa GAr. In accordance, insertion of a powerful degradation signal relieved the blockade of proteasomal degradation in Epstein–Barr virus nuclear antigen-1. Our findings suggest that the turnover of natural substrates may be finely tuned by GAr-like sequences that counteract targeting signals for proteasomal destruction.
DNA microarray reveals changes in gene expression of shear stressed human umbilical vein endothelial cells
Authors: McCormick Susan M. , Eskin Suzanne G. , McIntire Larry V. , Teng Christina L. , Lu Chiung-Mei , Russell Christopher G. , Chittur Krishnan K.
Published: July 2002

Using DNA microarray screening (GeneFilter 211, Research Genetics, Huntsville, AL) of mRNA from primary human umbilical vein endothelial cells (HUVEC), we identified 52 genes with significantly altered expression under shear stress [25 dynes/cm
2
for 6 or 24 h (1 dyne = 10 μN), compared with matched stationary controls]; including several genes not heretofore recognized to be shear stress responsive. We examined mRNA expression of nine genes by Northern blot analysis, which confirmed the results obtained on DNA microarrays. Thirty-two genes were up-regulated (by more than 2-fold), the most enhanced being cytochromes P450 1A1 and 1B1, zinc finger protein EZF/GKLF, glucocorticoid-induced leucine zipper protein, argininosuccinate synthase, and human prostaglandin transporter. Most dramatically decreased (by more than 2-fold) were connective tissue growth factor, endothelin-1, monocyte chemotactic protein-1, and spermidine/spermine N1-acetyltransferase. The changes observed suggest several potential mechanisms for increased NO production under shear stress in endothelial cells.
A calcium influx is triggered and propagates in the zygote as a wavefront during
in vitro
fertilization of flowering plants
Authors: Antoine A. F. , Faure J.-E. , Cordeiro S. , Dumas C. , Rougier M. , Feijó J. A.
Published: July 2002

In this paper, we report direct measurement of an influx of extracellular Ca
2+
induced by gamete fusion in flowering plants. This result was obtained during maize
in vitro
fertilization with the use of an extracellular Ca
2+
-selective vibrating probe. Ca
2+
influx recorded at the surface of isolated egg cells, with or without adhesion of a male sperm cell, was close to zero and stable over time. Gamete fusion, however, triggered a Ca
2+
influx in the vicinity of the sperm entry site with a delay of 1.8 ± 0.6 sec. The Ca
2+
influx spread subsequently through the whole egg cell plasma membrane as a wavefront, progressing at an estimated rate of 1.13 μm⋅sec
−1
. Once established, Ca
2+
influx intensities were sustained, monotonic and homogeneous over the whole egg cell, with an average peak influx of 14.92 pmol⋅cm
−2
⋅sec
−1
and an average duration of 24.4 min. The wavefront spread of channel activation correlates well with the cytological modifications induced by fertilization, such as egg cell contraction, and with the cytosolic Ca
2+
(
c
[Ca
2+
]) elevation previously reported. Calcium influx was inhibited effectively by gadolinium, possibly implicating mechanosensitive channels. Furthermore, artificial influxes created by incubation with Ca
2+
ionophores mimicked some aspects of egg activation. Taken together, these results suggest that, during fertilization in higher plants, gamete membrane fusion starts the first embryonic events by channel opening and Ca
2+
influx. In turn,
c
[Ca
2+
] may work as a trigger and possibly a space and time coordinator of many aspects of egg activation.
The magnitude distribution of declustered earthquakes in Southern California
Authors: Knopoff Leon
Published: July 2002

The binned distribution densities of magnitudes in both the complete and the declustered catalogs of earthquakes in the Southern California region have two significantly different branches with crossover magnitude nearM= 4.8. In the case of declustered earthquakes, theb-values on the two branches differ significantly from each other by a factor of about two. The absence of self-similarity across a broad range of magnitudes in the distribution of declustered earthquakes is an argument against the application of an assumption of scale-independence to models of main-shock earthquake occurrence, and in turn to the use of such models to justify the assertion that earthquakes are unpredictable. The presumption of scale-independence for complete local earthquake catalogs is attributable, not to a universal process of self-organization leading to future large earthquakes, but to the universality of the process that produces aftershocks, which dominate complete catalogs.
Symbiosis-specific expression of
Rhizobium etli casA
encoding a secreted calmodulin-related protein
Authors: Xi Chuanwu , Schoeters Eric , Vanderleyden Jos , Michiels Jan
Published: July 2002

Symbiosis between
Rhizobium
and its leguminous host
requires elaborate communication between the partners throughout the
interaction process. A calmodulin-like protein, termed
calsymin, was identified in
Rhizobium etli
; a
calmodulin-related protein in a Gram-negative bacterium had
not been described previously. Calsymin possesses three repeated
homologous domains. Each domain contains two predicted EF-hand
Ca
2+
-binding motifs. Ca
2+
-binding activity of
calsymin was demonstrated on purified protein.
R. etli
efficiently secretes calsymin without N-terminal cleavage of the
protein. The gene encoding calsymin,
casA
, is
exclusively expressed during colonization and infection of
R.
etli
with the host. Expression of
casA
is
controlled by a repressor protein, termed CasR, belonging to the TetR
family of regulatory proteins. Mutation of the
casA
gene
affects the development of bacteroids during symbiosis and symbiotic
nitrogen fixation.
Retrieval of memory for fear-motivated training initiates extinction requiring protein synthesis in the rat hippocampus
Authors: Vianna Monica R. M. , Szapiro German , McGaugh James L. , Medina Jorge H. , Izquierdo Ivan
Published: July 2002

Evidence that protein synthesis inhibitors induce amnesia in a variety of species and learning paradigms indicates that the consolidation of newly acquired information into stable memories requires the synthesis of new proteins. Because extinction of a response also requires acquisition of new information, extinction, like original learning, would be expected to require protein synthesis. The present experiments examined the involvement of protein synthesis in the hippocampus in the extinction of a learned fear-based response known to involve the hippocampus. Rats were trained in a one-trial inhibitory avoidance task in which they received footshock after stepping from a small platform to a grid floor. They were then given daily retention tests without footshock. The inhibitory response (e.g., remaining on the platform) gradually extinguished with repeated testing over several days. Footshock administered in a different context, instead of a retention test, prevented the extinction. Infusions of the protein synthesis inhibitor anisomycin (80 μg) into the CA1 region of the hippocampus (bilaterally) 10 min before inhibitory avoidance training impaired retention on all subsequent tests. Anisomycin infused into the hippocampus immediately after the 1st retention test blocked extinction of the response. Infusions administered before the 1st retention test induced a temporary (i.e., 1 day) reduction in retention performance and blocked subsequent extinction. These findings are consistent with other evidence that anisomycin blocks both the consolidation of original learning and extinction.
Mass spectrometry and immobilized enzymes for the screening of inhibitor libraries
Authors: Cancilla Mark T. , Leavell Michael D. , Chow Jason , Leary Julie A.
Published: July 2002

A technique has been developed to rapidly screen enzyme inhibitor
candidates from complex mixtures, such as those created by
combinatorial synthesis. Inhibitor libraries are screened by using
immobilized enzyme technologies and electrospray ionization ion
cyclotron resonance mass spectrometry. The library mixture is first
sprayed into the mass spectrometer, and compounds are identified. The
library is subsequently incubated with the immobilized enzyme of
interest under the correct conditions (buffer, pH, temperature) by
using an excess of enzyme to ensure a surplus of sites for ligand
binding. The immobilized enzyme/inhibitor mixture is centrifuged, and
an aliquot of supernatant is again analyzed by electrospray ionization
mass spectrometry. Potential inhibitors are quickly identified by
comparison of the spectra before and after incubation with the
immobilized enzyme. Non-inhibitors show no change in ion intensity
after incubation, whereas weak inhibitors exhibit a visible decrease in
ion abundance. Once inhibitor candidates have been identified, the
library is reinjected into the mass spectrometer, and tandem mass
spectrometry is used to determine the structure of the inhibitor
candidates as needed. This method has been successfully demonstrated by
identifying inhibitors of the enzymes pepsin and glutathione
S
-transferase from a 19- and 17-component library,
respectively. It is further shown that the immobilized enzyme can be
recycled and reused for continuous screening of additional new
libraries without adding additional enzyme.
RNA polymerase III transcription factor TFIIIC2 is overexpressed in ovarian tumors
Authors: Winter Andrew G. , Sourvinos George , Allison Simon J. , Tosh Kerrie , Scott Pamela H. , Spandidos Demetrios A. , White Robert J.
Published: July 2002

Most transformed cells display abnormally high levels of RNA
polymerase (pol) III transcripts. Although the full significance of
this is unclear, it may be fundamental because healthy cells use two
key tumor suppressors to restrain pol III activity. We present the
first evidence that a pol III transcription factor is overexpressed in
tumors. This factor, TFIIIC2, is a histone acetyltransferase that is
required for synthesis of most pol III products, including tRNA and 5S
rRNA. TFIIIC2 is a complex of five polypeptides, and mRNAs encoding
each of these subunits are overexpressed in human ovarian carcinomas;
this may explain the elevated TFIIIC2 activity that is found
consistently in the tumors. Deregulation in these cancers is unlikely
to be a secondary response to rapid proliferation, because there is
little or no change in TFIIIC2 mRNA levels when actively cycling cells
are compared with growth-arrested cells in culture. Using purified
factors, we show that raising the level of TFIIIC2 is sufficient to
stimulate pol III transcription in ovarian cell extracts. The data
suggest that overexpression of TFIIIC2 contributes to the abnormal
abundance of pol III transcripts in ovarian tumors.
Effect of the protein import machinery at the mitochondrial surface on precursor stability
Authors: Huang Shihai , Murphy Sherry , Matouschek Andreas
Published: July 2002

Many biological processes require proteins to undergo
conformational changes at the surface of membranes. For example, some
precursor proteins unfold at the surface of mitochondria and
chloroplasts before translocation into the organelles, and toxins such
as colicin A unfold to the molten globule state at bacterial surfaces
before inserting into the cell membrane. It is commonly thought that
the membrane surfaces and the associated protein machinery destabilize
the substrate proteins and that this effect is required for membrane
insertion or translocation. One of the best characterized translocation
processes is protein import into mitochondria. By measuring the
contributions of individual interactions within a model protein to its
stability at the mitochondrial surface and in free solution, we show
here that the mitochondrial surface neither induces the molten globule
state in this protein nor preferentially destabilizes any type of
interaction (e.g., hydrogen bonds, nonpolar, etc.) within the protein.
Because it is not possible to measure absolute protein stability at the
surface of mitochondria, we determined the stability of a tightly
associated protein–protein complex at the mitochondrial import site as
a model of the stability of a protein. We found the binding constants
of the protein–protein complex at the mitochondrial surface and in
free solution to be identical. Our results demonstrate that the
mitochondrial surface does not destabilize importing precursor proteins
in its vicinity.
Differential roles of phosphorylation in the formation of transcriptional active RNA polymerase I
Authors: Fath Stephan , Milkereit Philipp , Peyroche Gerald , Riva Michel , Carles Christophe , Tschochner Herbert
Published: July 2002

Regulation of rDNA transcription depends on the formation
and dissociation of a functional complex between RNA polymerase I (pol
I) and transcription initiation factor Rrn3p. We analyzed whether
phosphorylation is involved in this molecular switch. Rrn3p is a
phosphoprotein that is predominantly phosphorylated
in
vivo
when it is not bound to pol I.
In vitro
,
Rrn3p is able both to associate with pol I and to enter the
transcription cycle in its nonphosphorylated form. By contrast,
phosphorylation of pol I is required to form a stable pol I-Rrn3p
complex for efficient transcription initiation. Furthermore,
association of pol I with Rrn3p correlates with a change in the
phosphorylation state of pol I
in vivo
. We suggest that
phosphorylation at specific sites of pol I is a prerequisite for proper
transcription initiation and that phosphorylation/dephosphorylation
of pol I is one possibility to modulate cellular rDNA transcription
activity.
The archetype STYX/dead-phosphatase complexes with a spermatid mRNA-binding protein and is essential for normal sperm production
Authors: Wishart Matthew J. , Dixon Jack E.
Published: July 2002

Differentiation of spermatids into spermatozoa is regulated via phosphorylated RNA-binding proteins that modulate the expression of stage-specific mRNAs. We demonstrate that the phosphoserine, -threonine or -tyrosine, interaction protein, Styx, complexes with a testicular RNA-binding protein and is essential for normal spermiogenesis. Ablation of
Styx
expression in mouse disrupts round and elongating spermatid development, resulting in a >1,000-fold decrease in spermatozoa production. Moreover, Styx
−/−
males are infertile because of structural head abnormalities in residual epididymal sperm. Immunoprecipitation of Styx with Crhsp-24, a phosphorylated RNA-binding protein implicated in translational repression of histone mRNAs, provides a strategy for regulating posttranscriptional gene expression.
Time-resolved x-ray diffraction reveals multiple conformations in the M–N transition of the bacteriorhodopsin photocycle
Authors: Oka Toshihiko , Yagi Naoto , Fujisawa Tetsuro , Kamikubo Hironari , Tokunaga Fumio , Kataoka Mikio
Published: July 2002

We measured the M–N transition of wild-type
bacteriorhodopsin (pH 9, 10°C) by time-resolved
x-ray diffraction study at SPring8 BL45XU-A. We confirmed the
accumulation of M and N intermediates by absorbance measurements, and
we found that the time resolution of x-ray diffraction experiments (244
ms) was sufficient to resolve the M–N transition. From the x-ray
diffraction data, three components were decomposed by singular value
decomposition analysis. The existence of three components in the
M→N→BR reaction revealed that BR changes its structure during the
M–N transition. Moreover, the difference Fourier maps of reconstituted
fast and slow decay components clearly showed that the electron density
distributions of the F helix changes in the M–N transition. The
observed structural change at the F helix will increase access of the
Schiff base and D96 to the cytoplasmic surface and facilitate the
proton transfer steps that begin with the decay of the M state.