bluuebunny/crossref_metadata_2025 huggingface dataset:
Home range, movement and habitat utilisation of the Carpentarian rock-rat (Zyzomys palatalis) in an isolated habitat patch
Authors: Puckey Helen , Lewis Milton , Hooper David , Michell Carrie
Published: June 2004

Radio-telemetry was used to examine the home range, movement and habitat utilisation of the critically endangered Carpentarian rock-rat (Zyzomys palatalis) in an isolated habitat patch in the Gulf of Carpentaria hinterland over a 13-month period. Two home-range estimators were used in the study, (i) minimum convex polygon (MCP) and (ii) fixed Kernel (KL), the latter also being used to estimate core areas of activity. Based on a total sample size of 21 individuals, the mean MCP home range was 11 165 m2, similar to the mean KL home range of 10 687 m2. Core areas were, on average, 11.9% of the KL home-range estimate. There was no significant difference in the size of home range or core area of males and females. Juveniles had a significantly smaller home range than adults. Home ranges and, to a lesser degree, core areas were non-exclusive, with multiple areas of overlap (averaging 41% and 38% respectively) within and between all age and gender categories, but especially between males and between juveniles. Movement frequencies showed that animals made many short forays in a central area close to the arithmetic home-range mean and far fewer long forays of distances greater than 100 m from the central area. The spatial and temporal activity of Z. palatalis was concentrated in, but not confined to, the 'valley' and 'slope' habitats, with fewer movements of rats onto the surrounding 'plateau'. Resource selection analyses showed that Z. palatalis tended to prefer valley and slope habitats over the plateau and that the proportion of point locations was significantly higher for adults in the slope habitat and for juveniles in the valley habitat. Most home ranges were centred on the ecotone between these two habitat types. Although isolated and spatially limited, these habitat patches provide high-quality resources for dense populations of Z. palatalis. This study exemplifies a species' attempt to make efficient use of a limited resource in an otherwise hostile environment. Even small declines in habitat area or quality due to their vulnerability to fire would impact upon many animals.
Genetic analysis of a population crash in brush-tailed rock-wallabies (Petrogale penicillata) from Jenolan Caves, south-eastern Australia
Authors: Eldridge Mark D. B. , Rummery Catherine , Bray Cherylin , Zenger Kyall R. , Browning Teena L. , Close Robert L.
Published: June 2004

Although the theoretical effects of a severe reduction in effective population size (i.e. a bottleneck) are well known, relatively few empirical studies of bottlenecks have been based on extensive temporally spaced samples of a population both before and after a bottleneck. Here we describe the results of one such study, utilising the Jenolan Caves (JC) population of the brush-tailed rock-wallaby (Petrogale penicillata). When first sampled in 1985 (n = 20) the JC population comprised ~90 individuals. Subsequently the population crashed, and by 1992 only seven individuals remained. In 1996 the entire population (n = 10) was again sampled. Genetic diversity in the pre- and post-crash JC population was compared using 11 polymorphic microsatellite loci and PCR–SSCP analysis of the mitochondrial DNA control region. Only a single unique control region haplotype was detected in the pre- and post-crash JC population, although variant haplotypes were present in other P. penicillata populations. Of the 35 microsatellite alleles present in the pre-crash population, nine (26%) were lost during the bottleneck. The average number of rare alleles declined by 72%, allelic diversity was reduced by 30% and average heterozygosity declined by 10%. These observations are consistent with theoretical predictions. Additional analyses revealed that a P.�penicillata female at Wombeyan Caves was the only survivor of a 1990/91 reintroduction attempt using animals from JC. Of the microsatellite alleles detected in this female, 21% (4/19) were no longer present in the post-crash JC population. Furthermore, the genetic profiles of animals from the recently discovered Taralga population indicate that they are not derived from JC stock, but represent a threatened remnant of a hitherto undetected natural P. penicillata population.
Factors influencing fledgling production in little penguins (Eudyptula minor)
Authors: Knight C. , Rogers T.
Published: June 2004

An eight-year study was conducted on the breeding biology of the little penguin population at Lion Island. Forwards-selection Poisson regressions were used to determine whether variables such as year, date of lay, years since banding of each parent (indicator of age) and habitat influenced the fledgling numbers and average fledgling weight for adult pairs. 'Date of lay' provided the most significant model of fledgling numbers, while 'habitat' and 'year' as single-variable models also significantly influenced fledgling numbers. 'Date of lay' provided the most significant model of average fledgling weight. Future monitoring of the Lion Island colony therefore should focus on monitoring egg laying at the start of the breeding season, and maintaining high-quality nesting habitat.
An endoplasmic reticulum trafficking signal prevents surface expression of a voltage- and Ca
2+
-activated K
+
channel splice variant
Authors: Zarei M. M. , Eghbali M. , Alioua A. , Song M. , Knaus H.-G. , Stefani E. , Toro L.
Published: June 2004

Protein delivery to restricted plasma membrane domains is exquisitely regulated at different stages of the cell trafficking machinery. Traffic control involves the recognition of export/retention/retrieval signals in the endoplasmic reticulum (ER)/Golgi complex that will determine protein fate. A splice variant (SV), SV1, of the voltage- and Ca
2+
-activated K
+
channel α-subunit accumulates the channel in the ER, preventing its surface expression. We show that SV1 insert contains a nonbasic, hydrophobic retention/retrieval motif, CVLF, that does not interfere with proper folding and tetramerization of SV1. Localization of proteins in the ER by CVLF is independent of its position; originally, on the first internal loop, SV1 insert or CVLF perform equally well if placed at the middle or end of the α-subunit intracellular carboxyl terminus. Also, CVLF is able to restrict the traffic of an independently expressed transmembrane protein, β1-subunit. CVLF is present in proteins across species and in lower organisms. Thus, CVLF may have evolved to serve as a regulator of cellular traffic.
Species and functional group diversity independently influence biomass accumulation and its response to CO
2
and N
Authors: Reich Peter B. , Tilman David , Naeem Shahid , Ellsworth David S. , Knops Johannes , Craine Joseph , Wedin David , Trost Jared
Published: June 2004

The characteristics of plant assemblages influence ecosystem processes such as biomass accumulation and modulate terrestrial responses to global change factors such as elevated atmospheric CO
2
and N deposition, but covariation between species richness (
S
) and functional group richness (
F
) among assemblages obscures the specific role of each in these ecosystem responses. In a 4-year study of grassland species grown under ambient and elevated CO
2
and N in Minnesota, we experimentally varied plant
S
and
F
to assess their independent effects. We show here that at all CO
2
and N levels, biomass increased with
S
, even with
F
constant at 1 or 4 groups. Likewise, with
S
at 4, biomass increased as
F
varied continuously from 1 to 4. The
S
and
F
effects were not dependent upon specific species or functional groups or combinations and resulted from complementarity. Biomass increases in response to CO
2
and N, moreover, varied with time but were generally larger with increasing
S
(with
F
constant) and with increasing
F
(with
S
constant). These results indicate that
S
and
F
independently influence biomass accumulation and its response to elevated CO
2
and N.
Structure of the periplasmic component of a bacterial drug efflux pump
Authors: Higgins Matthew K. , Bokma Evert , Koronakis Eva , Hughes Colin , Koronakis Vassilis
Published: June 2004

Multidrug resistance among Gram-negative bacteria is conferred by three-component membrane pumps that expel diverse antibiotics from the cell. These efflux pumps consist of an inner membrane transporter such as the AcrB proton antiporter, an outer membrane exit duct of the TolC family, and a periplasmic protein known as the adaptor. We present the x-ray structure of the MexA adaptor from the human pathogen
Pseudomonas aeruginosa
. The elongated molecule contains three linearly arranged subdomains; a 47-Å-long α-helical hairpin, a lipoyl domain, and a six-stranded β-barrel. In the crystal, hairpins of neighboring MexA monomers pack side-by-side to form twisted arcs. We discuss the implications of the packing of molecules within the crystal. On the basis of the structure and packing, we suggest a model for the key periplasmic interaction between the outer membrane channel and the adaptor protein in the assembled drug efflux pump.
Selective facilitation of sexual solicitation in the female rat by a melanocortin receptor agonist
Authors: Pfaus James G. , Shadiack Annette , Van Soest Tanya , Tse Maric , Molinoff Perry
Published: June 2004

Disorders of sexual desire affect an estimated 30% of women in North America and Europe, with etiologies based on interpersonal, personal, and physiological factors. There are currently no pharmacological agents approved for use in the treatment of female sexual dysfunction. This is due, in part, to a focus on the effects of experimental drugs on reflexive components of sexual behavior, such as lordosis, in animal models. Here we report that PT-141, a peptide analogue of α-melanocyte-stimulating hormone that binds to central melanocortin receptors, selectively stimulates solicitational behaviors in the female rat. This occurs without affecting lordosis, pacing, or other sexual behaviors. PT-141 did not cause generalized motor activation, nor did it affect the perception of sexual reward. A selective pharmacological effect on appetitive sexual behavior in female rats has not been reported previously, and indicates that central melanocortin systems are important in the regulation of female sexual desire. Accordingly, PT-141 may be the first identified pharmacological agent with the capability to treat female sexual desire disorders.
Silent information regulator 2 potentiates Foxo1-mediated transcription through its deacetylase activity
Authors: Daitoku Hiroaki , Hatta Mitsutoki , Matsuzaki Hitomi , Aratani Satoko , Ohshima Takayuki , Miyagishi Makoto , Nakajima Toshihiro , Fukamizu Akiyoshi
Published: June 2004

Longevity regulatory genes include the Forkhead transcription factor FOXO and the NAD-dependent histone deacetylase silent information regulator 2 (Sir2). Genetic studies demonstrate that Sir2 acts to extend lifespan in
Caenorhabditis elegans
upstream of DAF-16, a member of the FOXO family, in the insulin-like signaling pathway. However, the molecular mechanisms underlying the requirement of DAF-16 activity in Sir2-mediated longevity remain unknown. Here we show that reversible acetylation of Foxo1 (also known as FKHR), the mouse DAF-16 ortholog, modulates its transactivation function. cAMP-response element-binding protein (CREB)-binding protein binds and acetylates Foxo1 at the K242, K245, and K262 residues, the modification of which is involved in the attenuation of Foxo1 as a transcription factor. Conversely, Sir2 binds and deacetylates Foxo1 at residues acetylated by cAMP-response element-binding protein-binding protein. Sir2 is recruited to insulin response sequence-containing promoter and increases the expression of manganese superoxide dismutase and p27
kip1
in a deacetylase-activity-dependent manner. Our findings establish Foxo1 as a direct and functional target for Sir2 in mammalian systems.
Methicillin-resistantStaphylococcus aureusin hospitals and the community: Stealth dynamics and control catastrophes
Authors: Cooper B. S. , Medley G. F. , Stone S. P. , Kibbler C. C. , Cookson B. D. , Roberts J. A. , Duckworth G. , Lai R. , Ebrahim S.
Published: June 2004

Methicillin-resistantStaphylococcus aureus(MRSA) represents a serious threat to the health of hospitalized patients. Attempts to reduce the spread of MRSA have largely depended on hospital hygiene and patient isolation. These measures have met with mixed success: although some countries have almost eliminated MRSA or remained largely free of the organism, others have seen substantial increases despite rigorous control policies. We use a mathematical model to show how these increases can be explained by considering both hospital and community reservoirs of MRSA colonization. We show how the timing of the intervention, the level of resource provision, and chance combine to determine whether control measures succeed or fail. We find that even control measures able to repeatedly prevent sustained outbreaks in the short-term can result in long-term control failure resulting from gradual increases in the community reservoir. If resources do not scale with MRSA prevalence, isolation policies can fail “catastrophically.”
A community of unknown, endophytic fungi in western white pine
Authors: Ganley Rebecca J. , Brunsfeld Steven J. , Newcombe George
Published: June 2004

The endophytic fungi of woody plants may be diverse as often claimed, and likewise, they may be functionally novel as demonstrated in a few studies. However, the endophyte taxa that are most frequently reported tend to belong to fungal groups composed of morphologically similar endophytes and parasites. Thus, it is plausible that endophytes are known (i.e., described) parasites in a latent phase within the host. If this null hypothesis were true, endophytes would represent neither additional fungal diversity distinct from parasite diversity nor a symbiont community likely to be novel ecologically. To be synonymous with parasites of the host, endophytes should at least be most closely related to those same parasites. Here we report that seven distinct parasites of
Pinus monticola
do not occur as endophytes. The majority of endophytes of
P. monticola
(90% of 2,019 cultures) belonged to one fungal family, the Rhytismataceae. However, not a single rhytismataceous endophyte was found to be most closely related by sequence homology to the three known rhytismataceous parasites of
P. monticola
. Similarly, neither endophytic
Mycosphaerella
nor endophytic
Rhizosphaera
isolates were most closely related to known parasites of
P. monticola
. Morphologically, the endophytes of
P. monticola
can be confounded with the parasites of the same host. However, they are actually most closely related to, but distinct from, parasites of other species of
Pinus
. If endophytes are generally unknown species, then estimates of 1 million endophytes (i.e., approximately 1 in 14 of all species of life) seem reasonable.
Desmin-related cardiomyopathy in transgenic mice: A cardiac amyloidosis
Authors: Sanbe Atsushi , Osinska Hanna , Saffitz Jeffrey E. , Glabe Charles G. , Kayed Rakez , Maloyan Alina , Robbins Jeffrey
Published: June 2004

An R120G missense mutation in the small heat shock protein α-B-crystallin (CryAB
R120G
) causes desmin-related cardiomyopathy (DRM). DRM is characterized by the formation of aggregates containing CryAB and desmin, and it can be recapitulated in transgenic mice by cardiac-specific expression of the mutant protein. In this article, we show that expression of CryAB
R120G
leads to the formation of electron-dense bodies characteristic of the DRMs and identify these bodies as aggresomes, which are characteristic of the neurodegenerative diseases. Cardiomyocytes transfected with adenovirus containing CryAB
R120G
establish the necessity and sufficiency of CryAB
R120G
expression for aggresome formation. The commonality of these aggresomes with oligomeric protein aggregates found in the amyloid-related degenerative diseases was corroborated by the presence of high levels of amyloid oligomers that may represent a primary toxic species in the amyloid diseases. These oligomeric amyloid intermediates are present also in cardiomyocytes derived from many human dilated and hypertrophic cardiomyopathies.
Labeling of fusion proteins with synthetic fluorophores in live cells
Authors: Keppler Antje , Pick Horst , Arrivoli Claudio , Vogel Horst , Johnsson Kai
Published: June 2004

A general approach for the sequential labeling of fusion proteins of O
6
-alkylguanine-DNA alkyltransferase (AGT) with different fluorophores in mammalian cells is presented. AGT fusion proteins with different localizations in the cell can be labeled specifically with different fluorophores, and the fluorescence labeling can be used for applications such as multicolor analysis of dynamic processes and fluorescence resonance energy transfer measurements. The facile access to a variety of different AGT substrates as well as the specificity of the labeling reaction should make the approach an important tool to study protein function in live cells.
An
in vivo
assay identifies changes in residue accessibility on mechanosensitive channel gating
Authors: Bartlett Jessica L. , Levin Gal , Blount Paul
Published: June 2004

MscL is a mechanosensitive channel of large conductance that functions as an “emergency release valve,” allowing bacteria to survive acute hypoosmotic stress. Although
Escherichia coli
MscL is the best-studied mechanosensitive channel, structural rearrangements occurring during gating remain disputed. Introduction of a charged residue into the pore of MscL was shown to result in a reduced-viability phenotype. Here, we probe for residues in the transmembrane domains that are exposed to the aqueous environment in the presence and absence of hypoosmotic shock by reacting a charged sulfhydryl reagent with substituted cysteines. Subsequent analysis of cell viability allows for an assessment of residues exposed in the closed and opening states
in vivo
. The results suggest that the crystal structure of MscL derived from the
Mycobacterium tuberculosis
orthologue may reflect a nearly closed rather than fully closed state and support a clockwise rotation of the pore-forming first transmembrane domain on gating.
Six nonnodulating plant mutants defective for Nod factor-induced transcriptional changes associated with the legume-rhizobia symbiosis
Authors: Mitra Raka M. , Shaw Sidney L. , Long Sharon R.
Published: June 2004

As the legume-rhizobia symbiosis is established, the plant recognizes bacterial-signaling molecules, Nod factors (NFs), and initiates transcriptional and developmental changes within the root to allow bacterial invasion and the construction of a novel organ, the nodule. Plant mutants defective in nodule initiation (Nod
-
) are thought to have defects in NF-signal transduction. However, it is unknown whether WT plants respond to NF-independent bacterial-derived signals or whether Nod
-
plant mutants show defects in global symbiosis-associated gene expression. To characterize plant gene expression in the establishment of the symbiosis, we used an Affymetrix oligonucleotide microarray representing 9,935
Medicago truncatula
expressed sequences. We identified 46 sequences that are differentially expressed in plants exposed for 24 h to WT
Sinorhizobium meliloti
or to the invasion defective
S. meliloti
mutant,
exoA
. Eight of these genes encode nucleolar proteins, which are implicated in ribosome biogenesis. We also identified differentially expressed transcription factors, signaling components, defense response proteins, stress response proteins, and several previously uncharacterized genes. NF appears both necessary and sufficient to induce most changes. Six of seven Nod
-
M. truncatula
mutants (
nfp
,
dmi1
,
dmi2
,
dmi3
,
nsp1
, and
nsp2
) showed no transcriptional response to
S. meliloti
, suggesting that the encoded proteins are required for initiating new transcription. The Nod
-
mutant
hcl
, however, exhibits a reduced transcriptional response to
S. meliloti
, indicating that the machinery responsible for initiating new transcription is at least partially functional in this mutant.
Preferred peptide backbone conformations in the unfolded state revealed by the structure analysis of alanine-based (AXA) tripeptides in aqueous solution
Authors: Eker Fatma , Griebenow Kai , Cao Xiaolin , Nafie Laurence A. , Schweitzer-Stenner Reinhard
Published: June 2004

We have combined Fourier transform IR, polarized Raman spectroscopy, and vibrational CD measurements of the amide I′ band profile of alanyl-X-alanine tripeptides in
2
H
2
O to obtain the dihedral angles of their central amino acid residue. X represents glycine, valine, methionine, histidine, serine, proline, lysine, leucine, tryptophan, tyrosine, and phenylalanine. The experimental data were analyzed by means of a recently developed algorithm, which exploits the excitonic coupling between the amide modes of the two peptide groups. The results were checked by measuring the respective electronic CD spectra. The investigated peptides can be sorted into three classes. Valine, phenylalanine, tryptophan, histidine, and serine predominantly adopt an extended β-strand conformation. Cationic lysine and proline prefer a polyproline II-like structure. Alanine, methionine, glycine, and leucine populate these two conformations with comparable probability. Our results are in variance with the prediction of the random-coil model, but supportive of Flory's isolated-pair hypothesis. We combined the obtained structural propensities of the investigated residues and similar information about other residues in the literature (i.e., glutamate, aspartate, isoleucine, and glutamine) to predict possible conformations of the monomeric amyloid β peptide Aβ
1–42
in aqueous solution, which reproduces results from most recent spectroscopic studies. Thus, it is demonstrated that the unfolded state of peptides can be understood in terms of the intrinsic structural propensities of their amino acid residues.
Activity-based probes for the proteomic profiling of metalloproteases
Authors: Saghatelian Alan , Jessani Nadim , Joseph Arul , Humphrey Mark , Cravatt Benjamin F.
Published: June 2004

Metalloproteases (MPs) are a large and diverse class of enzymes implicated in numerous physiological and pathological processes, including tissue remodeling, peptide hormone processing, and cancer. MPs are tightly regulated by multiple posttranslational mechanisms
in vivo
, hindering their functional analysis by conventional genomic and proteomic methods. Here we describe a general strategy for creating activity-based proteomic probes for MPs by coupling a zinc-chelating hydroxamate to a benzophenone photocrosslinker, which promote selective binding and modification of MP active sites, respectively. These probes labeled active MPs but not their zymogen or inhibitor-bound counterparts and were used to identify members of this enzyme class up-regulated in invasive cancer cells and to evaluate the selectivity of MP inhibitors in whole proteomes. Interestingly, the matrix metalloproteinase inhibitor GM6001 (ilomastat), which is currently in clinical development, was found to also target the neprilysin, aminopeptidase, and dipeptidylpeptidase clans of MPs. These results demonstrate that MPs can display overlapping inhibitor sensitivities despite lacking sequence homology and stress the need to evaluate MP inhibitors broadly across this enzyme class to develop agents with suitable target selectivities
in vivo
. Activity-based profiling offers a powerful means for conducting such screens, as this approach can be carried out directly in whole proteomes, thereby facilitating the discovery of disease-associated MPs concurrently with inhibitors that selectively target these proteins.
Coupling of retinal isomerization to the activation of rhodopsin
Authors: Patel Ashish B. , Crocker Evan , Eilers Markus , Hirshfeld Amiram , Sheves Mordechai , Smith Steven O.
Published: June 2004

Activation of the visual pigment rhodopsin is caused by 11-cis to -trans isomerization of its retinal chromophore. High-resolution solid-state NMR measurements on both rhodopsin and the metarhodopsin II intermediate show how retinal isomerization disrupts helix interactions that lock the receptor off in the dark. We made 2D dipolar-assisted rotational resonance NMR measurements between
13
C-labels on the retinal chromophore and specific
13
C-labels on tyrosine, glycine, serine, and threonine in the retinal binding site of rhodopsin. The essential aspects of the isomerization trajectory are a large rotation of the C20 methyl group toward extracellular loop 2 and a 4- to 5-Å translation of the retinal chromophore toward transmembrane helix 5. The retinal–protein contacts observed in the active metarhodopsin II intermediate suggest a general activation mechanism for class A G protein-coupled receptors involving coupled motion of transmembrane helices 5, 6, and 7.
Delivery of therapeutic levels of heparin and low-molecular-weight heparin through a pulmonary route
Authors: Qi Yiwei , Zhao Ganlin , Liu Dongfang , Shriver Zachary , Sundaram Mallik , Sengupta Shiladitya , Venkataraman Ganesh , Langer Robert , Sasisekharan Ram
Published: June 2004

Although heparin and low-molecular-weight heparins (LMWH) have been widely used clinically as anticoagulants, their broader use has been limited by the lack of noninvasive delivery methods for this class of molecules. In this study, we demonstrate an efficient, rapid, and reproducible delivery system for heparin through the lungs that is not confined to particles of a certain geometric or aerodynamic diameter. Importantly, blood levels after intrapulmonary administration of either heparin or LMWH were comparable to that of s.c. administration but are characterized by a more rapid onset of action (
t
1/2
= 40 min vs. 2.5 h, respectively). Furthermore, we show in animal models, that inhaled heparin species efficiently inhibit diseases such as thrombosis and emphysema, and that the repetitive inhalation of formulated LMWH results in no observable toxicity from the delivery of reproducible systemic levels of heparin or LMWH.
Expression of butyrophilin (Btn1a1) in lactating mammary gland is essential for the regulated secretion of milk–lipid droplets
Authors: Ogg Sherry L. , Weldon Anne K. , Dobbie Lorraine , Smith Andrew J. H. , Mather Ian H.
Published: June 2004

Butyrophilin 1a1 (Btn1a1), which is a member of the Ig superfamily, is highly expressed in the lactating mammary gland and is secreted into milk in association with lipid droplets. To determine the potential function of Btn1a1 in milk secretion, we ablated
Btn1a1
in mice and analyzed the lactation phenotype of homozygous (
Btn1a1
-/-
) animals. Two mutant mouse lines were generated in which expression of Btn1a1 was either disrupted or eliminated, respectively. The regulated secretion of milk–lipid droplets was severely compromised in both mutant mouse lines in comparison to wild-type animals. Large pools of triacylglycerol accumulated in the cytoplasm of secretory cells, and lipid droplets escaped from the apical surface with disrupted outer membranes. Luminal spaces became engorged with unstable lipid droplets, which coalesced to form large aggregates. The amount of lipid (wt/vol) was elevated, on average by 50%, during the first 10 days of lactation, and the diameter of the droplets was up to seven times larger than the normal diameter. In contrast, there was no significant difference between wild-type and null animals in the relative amounts of skim-milk proteins secreted from Golgi-derived secretory vesicles. Approximately half the pups suckling
Btn1a1
-/-
animals died within the first 20 days, and weaning weights for the surviving pups were 60–80% of those suckling wild-type mice. Thus, expression of
Btn1a1
is essential for the regulated secretion of milk–lipid droplets. We speculate that Btn1a1 functions either as a structural protein or as a signaling receptor by binding to xanthine dehydrogenase/oxidase.
Microviscometry reveals reduced blood viscosity and altered shear rate and shear stress profiles in microvessels after hemodilution
Authors: Long David S. , Smith Michael L. , Pries Axel R. , Ley Klaus , Damiano Edward R.
Published: June 2004

We show that many salient hemodynamic flow properties, which have been difficult or impossible to assess in microvessels
in vivo
, can be estimated by using microviscometry and fluorescent microparticle image velocimetry in microvessels >20 μm in diameter. Radial distributions in blood viscosity, shear stress, and shear rate are obtained and used to predict axial pressure gradient, apparent viscosity, and endothelial-cell surface-layer thickness
in vivo
. Based solely on microparticle image velocimetry data, which are readily obtainable during the course of most intravital microscopy protocols from systemically injected particle tracers, we show that the microviscometric method consistently predicted a reduction in local and apparent blood viscosity after isovolemic hemodilution. Among its clinical applications, hemodilution is a procedure that is used to treat various pathologies that require reduction in peripheral vascular-flow resistance. Our results are directly relevant in this context because they suggest that the fractional decrease in systemic hematocrit is ≈25–35% greater than the accompanying fractional decrease in microvascular-flow resistance
in vivo
. In terms of its fundamental usefulness, the microviscometric method provides a comprehensive quantitative analysis of microvascular hemodynamics that has applications in broad areas of medicine and physiology and is particularly relevant to quantitative studies of angiogenesis, tumor growth, leukocyte adhesion, vascular-flow resistance, tissue perfusion, and endothelial-cell mechanotransduction.
Engineered DsbC chimeras catalyze both protein oxidation and disulfide-bond isomerization in
Escherichia coli
: Reconciling two competing pathways
Authors: Segatori Laura , Paukstelis Paul J. , Gilbert Hiram F. , Georgiou George
Published: June 2004

In the
Escherichia coli
periplasm, the formation of protein disulfide bonds is catalyzed by DsbA and DsbC. DsbA is a monomer that is maintained in a fully oxidized state by the membrane enzyme DsbB, whereas DsbC is a dimer that is kept reduced by a second membrane protein, DsbD. Although the catalytic regions of DsbA and DsbC are composed of structurally homologous thioredoxin motif domains, DsbA serves only as an oxidase
in vivo
, whereas DsbC catalyzes disulfide reduction and isomerization and also exhibits significant chaperone activity. To reconcile the distinct catalytic activities of DsbC and DsbA, we constructed a series of chimeras comprising of the dimerization domain of DsbC, with or without the adjacent α-helical linker region, fused either to the first, second, third, or fifth residue of intact DsbA or to thioredoxin. The chimeras fully substituted for DsbC in disulfide-bond rearrangement and also were able to restore protein oxidation in a
dsbA
background. Remarkably, the chimeras could serve as a single catalyst for both disulfide-bond formation and rearrangement, thus reconciling the kinetically competing DsbB–DsbA and DsbD–DsbC pathways. This property appeared to depend on the orientation of the DsbA active-site cysteines with respect to the DsbC dimerization domain.
In vitro
, the chimeras had high chaperone activity and significant reductase activity but only 15–22% of the disulfide-isomerization activity of DsbC, suggesting that rear-rangement of nonnative disulfides may be mediated primarily by cycles of random reduction and reoxidation.
The bioactive Taxol conformation on β-tubulin: Experimental evidence from highly active constrained analogs
Authors: Ganesh Thota , Guza Rebecca C. , Bane Susan , Ravindra Rudravajhala , Shanker Natasha , Lakdawala Ami S. , Snyder James P. , Kingston David G. I.
Published: June 2004

The important anticancer drug Taxol (paclitaxel) binds to tubulin in a stoichiometric ratio and promotes its assembly into microtubules. The conformation of microtubule-bound drug has been the subject of intense study, and various suggestions have been made. In this work we present experimental and theoretical evidence that Taxol adopts a T-shaped conformation when it is bound to tubulin.
NF-κB activation in human breast cancer specimens and its role in cell proliferation and apoptosis
Authors: Biswas Debajit K. , Shi Qian , Baily Shanon , Strickland Ian , Ghosh Sankar , Pardee Arthur B. , Iglehart J. Dirk
Published: June 2004

Lack of molecular targets in estrogen receptor-negative (ER-negative) breast cancer is a major therapeutic hurdle. We studied NF-κB activation in human breast tumors and in carcinoma cell lines. Activated NF-κB was detected predominantly in ER-negative vs. ER-positive breast tumors and mostly in ER-negative and ErbB2-positive tumors (86%). These
in vivo
results demonstrate association of activated NF-κB with a subgroup of human breast tumors and are consistent with previously reported
in vitro
observations using similar classes of human breast cancer cell lines. Finding such an association suggested functional and biological significance. Immunofluorescence demonstrated increased nuclear p65, a component of the active NF-κB complex, in cytokeratin 19 (CK19)-positive epithelial cells of ER-negative/ErbB2-positive tumor samples. In contrast, nuclear NF-κB was detected mostly in stroma of ER-negative and ErbB2-negative tumors, suggesting a role of activated NF-κB in intercellular signaling between epithelial and stromal cells in this type of breast cancers. To elucidate roles of activated NF-κB, we used an ER-negative and ErbB2-positive human breast tumor cell line (SKBr3). The polypeptide heregulin β1 stimulated, and herceptin, the anti-ErbB2 antibody, inhibited, NF-κB activation in SKBr3 cells. The NF-κB essential modulator (NEMO)-binding domain (NBD) peptide, an established selective inhibitor of IκB-kinase (IKK), blocked heregulin-mediated activation of NF-κB and cell proliferation, and simultaneously induced apoptosis only in proliferating and not resting cells. These results substantiate the hypothesis that certain breast cancer cells rely on NF-κB for aberrant cell proliferation and simultaneously avoid apoptosis, thus implicating activated NF-κB as a therapeutic target for distinctive subclasses of ER-negative breast cancers.
Yeast telomerase RNA: A flexible scaffold for protein subunits
Authors: Zappulla David C. , Cech Thomas R.
Published: June 2004

In the yeast
Saccharomyces cerevisiae
, distinct regions of the 1.2-kb telomerase RNA (TLC1) bind to the catalytic subunit Est2p and to accessory proteins. In particular, a bulged stem structure binds the essential regulatory subunit Est1p. We now show that the Est1p-binding domain of the RNA can be moved to three distant locations with retention of telomerase function
in vivo
. We present the Est1p relocation experiment in the context of a working model for the secondary structure of the entire TLC1 RNA, based on thermodynamic considerations and comparative analysis of sequences from four species. The model for TLC1 has three long quasihelical arms that bind the Ku, Est1p, and Sm proteins. These arms emanate from a central catalytic core that contains the template and Est2p-binding region. Deletion mutagenesis provides evidence that the Sm arm exists
in vivo
and can be shortened by 42 predicted base pairs with retention of function; therefore, precise positioning of Sm proteins, like Est1p, is not required within telomerase. In the best-studied ribonucleoprotein enzyme, the ribosome, the RNAs have specific three-dimensional structures that orient the functional elements. In the case of yeast telomerase, we propose that the RNA serves a very different function, providing a flexible tether for the protein subunits.
A probable Na
+
(K
+
)/H
+
exchanger on the chloroplast envelope functions in pH homeostasis and chloroplast development in
Arabidopsis thaliana
Authors: Song Chun-Peng , Guo Yan , Qiu Quansheng , Lambert Georgina , Galbraith David W. , Jagendorf André , Zhu Jian-Kang
Published: June 2004

Electroneutral monovalent cation/proton antiport across the chloroplast envelope has been shown previously to have an important regulatory effect on stromal pH and thereby on photosynthetic carbon reduction. Here we report that an
Arabidopsis
nuclear gene,
AtCHX23
, encodes a putative Na
+
(K
+
)/H
+
exchanger and functions in the adjustment of pH in the cytosol and possibly in maintaining a high pH level in the chloroplast stroma. The AtCHX23 protein is localized in the chloroplast envelope. Plastids from
chx23
mutants had straight thylakoids but lacked grana lamellae.
chx23
mutant leaves were pale yellow and had a much reduced chlorophyll content. The chlorophyll content of
chx23
was increased by growing in medium at low (4.0) pH and decreased by growing at high (7.0) pH. The cytosolic pH in the leaves of the mutant was significantly higher than that in the wild type.
chx23
mutants displayed a high sensitivity to NaCl. Together, these data indicate that CHX23 is a probable chloroplast Na
+
(K
+
)/H
+
exchanger important for pH homeostasis and chloroplast development and function.
Rice yields decline with higher night temperature from global warming
Authors: Peng Shaobing , Huang Jianliang , Sheehy John E. , Laza Rebecca C. , Visperas Romeo M. , Zhong Xuhua , Centeno Grace S. , Khush Gurdev S. , Cassman Kenneth G.
Published: June 2004

The impact of projected global warming on crop yields has been evaluated by indirect methods using simulation models. Direct studies on the effects of observed climate change on crop growth and yield could provide more accurate information for assessing the impact of climate change on crop production. We analyzed weather data at the International Rice Research Institute Farm from 1979 to 2003 to examine temperature trends and the relationship between rice yield and temperature by using data from irrigated field experiments conducted at the International Rice Research Institute Farm from 1992 to 2003. Here we report that annual mean maximum and minimum temperatures have increased by 0.35°C and 1.13°C, respectively, for the period 1979–2003 and a close linkage between rice grain yield and mean minimum temperature during the dry cropping season (January to April). Grain yield declined by 10% for each 1°C increase in growing-season minimum temperature in the dry season, whereas the effect of maximum temperature on crop yield was insignificant. This report provides a direct evidence of decreased rice yields from increased nighttime temperature associated with global warming.
Treating cocaine addiction with viruses
Authors: Carrera M. Rocio A. , Kaufmann Gunnar F. , Mee Jenny M. , Meijler Michael M. , Koob George F. , Janda Kim D.
Published: June 2004

Cocaine addiction continues to be a major health and social problem in the United States and other countries. Currently used pharmacological agents for treating cocaine abuse have proved inadequate, leaving few treatment options. An alternative is to use protein-based therapeutics that can eliminate the load of cocaine, thereby attenuating its effects. This approach is especially attractive because the therapeutic agents exert no pharmacodynamic action of their own and therefore have little potential for side effects. The effectiveness of these agents, however, is limited by their inability to act directly within the CNS. Bacteriophage have the capacity to penetrate the CNS when administered intranasally. Here, a method is presented for engineering filamentous bacteriophage to display cocaine-binding proteins on its surface that sequester cocaine in the brain. These antibody-displaying constructs were examined by using a locomotor activity rodent model to assess the ability of the phage-displayed proteins to block the psychoactive effects of cocaine. Results presented demonstrate a strategy in the continuing efforts to find effective treatments for cocaine addiction and suggest the application of this protein-based treatment for other drug abuse syndromes.
Food-frequency questionnaires: a review of their design, validation and utilisation
Authors: Cade J. E. , Burley V. J. , Warm D. L. , Thompson R. L. , Margetts B. M.
Published: June 2004

A review of the literature concerning the design, utilisation and validation of food-frequency questionnaires (FFQ) has been carried out using a semi-systematic approach to obtaining, reviewing and extracting data from articles. Databases were searched from 1980 to 1999. The present review identified 227 validation (from 1980 to September 1999) and 164 utilisation (for 1998 only) studies. A number of design issues have been evaluated through the present review. These include: the need to consider how portion sizes have been described, self-defined giving higher mean correlations; how an FFQ was administered, interviewer-administered giving higher mean correlations for some nutrients; how many items to include on an FFQ, those with the largest number of items having higher correlations. Validation techniques were described. Most validation studies involved comparing an FFQ against another dietary assessment method; only 19 % compared an FFQ to a biomarker. Measurement differences were most commonly assessed by correlation coefficients as opposed to other more appropriate methods. Mean correlation coefficients were highest for Ca and fat, and lowest for vitamin A and vegetables. The utilisation studies showed that FFQ were most commonly used in cross-sectional surveys, with ninety-three of the FFQ being designed to be disease-specific. The present review results were presented to a group of experts and a consensus arrived at concerning the development, validation and use of FFQ. Recommendations derived from the consensus arising from the literature review are presented as an appendix to the present paper.
The potential of genetically enhanced plants to address food insecurity
Authors: Christou Paul , Twyman Richard M.
Published: June 2004

Food insecurity is one of the most important social issues faced today, with 840 million individuals enduring chronic hunger and three billion individuals suffering from nutrient deficiencies. Most of these individuals are poverty stricken and live in developing countries. Strategies to address food insecurity must aim to increase agricultural productivity in the developing world in order to tackle poverty, and must provide long-term improvements in crop yields to keep up with demand as the world's population grows. Genetically enhanced plants provide one route to sustainable higher yields, either by increasing the intrinsic yield capability of crop plants or by protecting them from biotic and abiotic constraints. The present paper discusses a range of transgenic approaches that could increase agricultural productivity if applied on a large scale, including the introduction of genes that confer resistance to pests and diseases, or tolerance of harsh environments, and genes that help to lift the intrinsic yield capacity by increasing metabolic flux towards storage carbohydrates, proteins and oils. The paper also explores how the nutritional value of plants can be improved by genetic engineering. Transgenic plants, as a component of integrated strategies to relieve poverty and deliver sustainable agriculture to subsistence farmers in developing countries, could have a significant impact on food security now and in the future.
Influence of age on mastication: effects on eating behaviour
Authors: Mioche Laurence , Bourdiol Pierre , Peyron Marie-Agnès
Published: June 2004

The present review covers current knowledge about the ageing of oral physiology related to mastication and its effects on eating behaviour. Mastication is the first process undergone by a food during feeding. It has a key role in the maintenance of nutritional status in two respects. First, the perceptions of food's sensory properties elicited during chewing and swallowing are one of the major determinants of the pleasure which drives us to eat; second, the properties of the swallowed bolus are affected by oral conditions and this may modulate the subsequent phases of digestion. Ageing in healthy dentate subjects induces moderate changes in oral physiology. Changes in neuromuscular activity are partly compensated by changes in chewing behaviour. No clear age effect is seen in texture perception, although this does impact on food bolus properties. In contrast, great alterations in both chewing behaviour and food bolus properties are observed when ageing is associated with a compromised dentition, general health alterations and drug intake. Eating behaviour is far more complex than just chewing behaviour and the concerns of the elderly about food cannot be explained solely by oral physiology. Discrepancies are often noticed with older subjects between various objective measurements of oral performance and corresponding measures of self-perception. In addition, although more foods are recognised as hard to chew with increasing age, there is no clear shift in preference towards food that is easy to chew. Food choices and food consumption are also driven by memory, psychology and economic factors. Advances in the understanding of food choice in the elderly need a sustained collaborative research effort between sensory physiologists, nutritionists, and food scientists.
Biochemical approaches for nutritional support of skeletal muscle protein metabolism during sepsis
Authors: Vary Thomas C. , Lynch Christopher J.
Published: June 2004

Sepsis initiates a unique series of modifications in the homeostasis of N metabolism and profoundly alters the integration of inter-organ cooperatively in the overall N and energy economy of the host. The net effect of these alterations is an overall N catabolic state, which seriously compromises recovery and is semi-refractory to treatment with current therapies. These alterations lead to a functional redistribution of N (amino acids and proteins) and substrate metabolism among injured tissues and major body organs. The redistribution of amino acids and proteins results in a quantitative reordering of the usual pathways of C and N flow within and among regions of the body with a resultant depletion of the required substrates and cofactors in important organs. The metabolic response to sepsis is a highly integrated, complex series of reactions. To understand the regulation of the response to sepsis, a comprehensive, integrated analysis of the fundamental physiological relationships of key metabolic pathways and mechanisms in sepsis is essential. The catabolism of skeletal muscles, which is manifested by an increase in protein degradation and a decrease in synthesis, persists despite state-of-the-art nutritional care. Much effort has focused on the modulation of the overall amount of nutrients given to septic patients in a hope to improve efficiencies in utilisation and N economies, rather than the support of specific end-organ targets. The present review examines current understanding of the processes affected by sepsis and testable means to circumvent the sepsis-induced defects in protein synthesis in skeletal muscle through increasing provision of amino acids (leucine, glutamine, or arginine) that in turn act as nutrient signals to regulate a number of cellular processes.