bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of Cosmic Ray Rejection Algorithms on Single-Shot Exposures
Authors: Farage Catherine L. , Pimbblet Kevin A.
Published: August 2005

To maximise data output from single-shot astronomical images, the rejection of cosmic rays is important. We present the results of a benchmark trial comparing various cosmic ray rejection algorithms. The procedures assess relative performances and characteristics of the processes in cosmic ray detection, rates of false detections of true objects, and the quality of image cleaning and reconstruction. The cosmic ray rejection algorithms developed by Rhoads (2000, PASP, 112, 703), van Dokkum (2001, PASP, 113, 1420), Pych (2004, PASP, 116, 148), and the IRAF task XZAP by Dickinson are tested using both simulated and real data. It is found that detection efficiency is independent of the density of cosmic rays in an image, being more strongly affected by the density of real objects in the field. As expected, spurious detections and alterations to real data in the cleaning process are also significantly increased by high object densities. We find the Rhoads' linear filtering method to produce the best performance in the detection of cosmic ray events; however, the popular van Dokkum algorithm exhibits the highest overall performance in terms of detection and cleaning.
Rates of inbreeding using DNA fingerprinting in aquaculture breeding programs at various broodstock fitness levels — a simulation study
Authors: Macbeth M.
Published: August 2005

A simulation study was used to examine the potential use of DNA fingerprinting (DNA tagging) as a tool to avoid excessive inbreeding by identifying suitable candidate breeders in genetic selection programs. ‘Broodstock fitness’ (the ability of broodstock to survive from harvest and reproduce) needs to be considered in designing breeding programs using DNA tagging. In this study, reduced broodstock fitness increased inbreeding exponentially. The level of inbreeding was also dependent on the intraclass correlation (t), selection intensity, number of individuals DNA tagged (NDNA), number of families maintained (Nf) and the number of candidate breeders retained per sex/family at harvest (C). With a broodstock fitness of 0.90, DNA tagging could theoretically achieve a selection intensity, in terms of the total phenotypic variance, of 2.90 standard deviations with 800 000 graded at harvest, while maintaining an inbreeding rate of 1.0% per generation (NDNA = 800, Nf = 30, C = 4, t = 0.3). In practice, the numbers required could be achieved by growing families in individual facilities (e.g. sea cages for barramundi or ponds for prawns). When mechanical grading is not possible, the selection pool may be limited to a level where physical tagging is feasible. In this case, there was no advantage in selection response using DNA tagging compared with physical tags. DNA tagging as a selection tool may be more feasible when broodstock fitness is above 0.6, and may fill a niche where industry infrastructure is not large enough to support separate rearing of families or where physical tagging is not economically viable or suitable. DNA tagging may also be useful as a means of recovering families in backup facilities where families have been pooled to reduce infrastructure costs. Due to the random nature of DNA sampling, not all families may be recovered and a reduction in selection pressure may facilitate family recovery.
Harnessing the bovine genome sequence for the Australian cattle and sheep industries
Authors: Dalrymple B. P.
Published: August 2005

Genomics is an emerging science and the release of the human and mouse genomes has significantly altered our picture of the information content of mammalian genomes. A smaller number of protein coding genes, and a larger number of genes that do not appear to encode protein products, the so-called non-coding RNAs (ncRNAs), have been identified. The first 2 drafts of the bovine genome sequence have been released, and work to utilise the framework of the bovine genome to facilitate ovine genomics is underway. In anticipation of the requirement for a detailed analysis of the ruminant genomes, their transcriptomes, interactomes, regulomes and similar, we have been developing the informatics platform for the analysis and integration of genome sequences and expression data for cattle and sheep. This resource will enable us to utilise the ruminant datasets and integrate them with equivalent data from other mammals for the advancement of animal scientific research for applications in the cattle and sheep industries in Australia.
Adaptability in tropical beef cattle: genetic parameters of growth, adaptive and temperament traits in a crossbred population
Authors: Prayaga K. C. , Henshall J. M.
Published: August 2005

Adaptability in tropical beef cattle can be assessed by measurable traits such as growth under the influence of environmental stressors, by parasite resistance as measured by indicator traits such as tick counts (TICK) and faecal egg counts of worms (EPG), by heat resistance as measured by indicator traits such as rectal temperatures (TEMP) and coat scores (COAT) and, to a certain extent, temperament of the animal as measured by flight time (FT). Data from a crossbreeding experiment involving various genotypes derived from tropically adapted British, Sanga-derived, Zebu cross, Zebu and Continental beef cattle breeds were analysed to estimate variance components and genetic parameters of growth, adaptive and temperament traits. Breed group differences were accounted for by including fractional coefficients of direct and maternal additive and dominance genetic effects as covariates. In the univariate analyses, 6 models were compared ranging from the simplest model with animal as the only random effect to the full model comprising direct and maternal additive genetic variance and their covariance and the permanent environment effect due to dam (growth traits) and animal (adaptive and temperament traits). The heritability estimates were 0.41, 0.21, 0.19, 0.28, 0.41 and 0.15 for birth weight (BWT), weaning weight (WWT), preweaning average daily gain (PREADG), yearling weight (YWT), final weight at about 18 months of age (FWT) and post-weaning average daily gain (POADG), respectively. The maternal component of additive genetic variance as a proportion of phenotypic variance in BWT, WWT and PREADG was 0.15, 0.10 and 0.10, respectively. The heritability estimates for TICK, EPG, TEMP, COAT and FT were 0.13, 0.24, 0.12, 0.26 and 0.20, respectively. High positive genetic and phenotypic correlations were observed among growth traits. Low (insignificant) genetic correlations were observed between TICK, EPG and growth traits. However, genetic correlations between growth traits and heat tolerance traits (TEMP and COAT) were moderately negative implying that as the ability of an animal to handle heat stress increases, growth also increases at the genetic level. Genetic correlations among TICK, EPG and TEMP were moderately positive, suggesting that closely-linked genes affect these adaptive traits. The significant negative genetic relationship between TEMP and FT suggests that cattle with high heat resistance have desirable temperament. With the increasing crossbred populations in the northern Australian beef cattle industry, the best breeding strategy should aim to exploit both crossbreeding and within population selection to make improvements in growth, adaptive and temperament traits to increase overall productivity of the enterprise.
Maternal productivity of Angus cows divergently selected for post-weaning residual feed intake
Authors: Arthur P. F. , Herd R. M. , Wilkins J. F. , Archer J. A.
Published: August 2005

Data on 185 Angus cows were used to study the effect of divergent selection for residual feed intake on maternal productivity across 3 mating seasons, starting from 2000. The cows were the result of 1 to 2.5 generations of selection (mean of 1.5), and differed in estimated breeding value for residual feed intake by 0.8 kg/day. In general, cows lost subcutaneous fat (measured 2 times a year) during the period when they were nursing calves, and gained fat thereafter. No significant selection line differences in fatness were observed except for those measured at the start of the 2000 (10.8 ± 0.4 v. 9.3 ± 0.4 mm), 2001 (11.3 ± 0.4 v. 9.8 ± 0.4 mm) and 2002 (7.0 ± 0.5 v. 5.7 ± 0.5 mm) mating seasons, where high residual feed intake cows had significantly (P<0.05) higher rib fat depths. No significant selection line differences in weight (measured 4 times a year) were observed. However, the cows either maintained or lost weight during the calf nursing period, and gained weight thereafter, with mean weights ranging from 450 to 658 kg. There were no significant selection line differences in pregnancy (mean 90.4%), calving (mean 88.7%) and weaning (mean of 80.8%) rates, milk yield (mean 7.7 kg/day) and weight of calf weaned per cow exposed to bull (mean 195 kg). The study indicates that after 1.5 generations of divergent selection for residual feed intake there are no significant selection line differences for maternal productivity traits.
The potential of new genetic technologies in selecting for stress resistance in pigs
Authors: Kerr C. A. , Hines B. M.
Published: August 2005

This paper examines the potential for breeding stress resistance in pigs through an understanding of the physiology of the stress response and its associated genetic basis. Pigs reared in commercial units can encounter numerous concurrent stressors that can have a negative impact on performance and welfare. Stress induces physiological and behavioural responses that are multidimensional, consisting of a complex neuroendocrine and immune signalling milieu. Some stress-related genetic parameters have been identified using conventional genetic approaches applied in experimental models. However, these traits do not capture the complexity of the stress response. As a result, the molecular mechanisms underlying the variation associated with stress resistance in pigs in a commercial environment is poorly understood. Gene expression profiling is a powerful tool that can be applied to systematically elucidate stress response pathways and networks. Consequently, gene expression technologies have been applied to identify some putative stress-regulated genes. Further application of these and more traditional technologies will aid in elucidating stress resistance using gene expression as a measure of phenotypic variation at a molecular level. It is envisaged that in the future, tools for selecting for stress resistance could eventually be applied on-farm to enhance production, health and welfare status.
Practical aspects of a genetic evaluation system using parentage assigned from genetic markers
Authors: Dodds K. G. , Sise J. A. , Tate M. L.
Published: August 2005

Animal breeding values can be calculated when genetic markers have been used to help determine the parentage of some of the animals, but their parentage has been incompletely determined. The pedigree sampling method is 1 computing strategy for calculating these breeding values. This paper describes and discusses methods for dealing with a number of practical issues that arise when implementing such a system for industry use. In particular, diagnostic systems for detecting inadequacies or possible errors in the genotyping systems and the recording of animal management are developed. Also, characteristics of the best assigned pedigrees are calculated according to mating group and used to check for sires missing from these groups. The correlation between breeding values estimated from a single sampled pedigree (using parentage probabilities) and those estimated as the mean from many sampled pedigrees gives a diagnostic to indicate which estimated breeding values are more influenced by uncertainties in relationships. For the analysis of survival traits, a method to enumerate and assign likely parentage to dead offspring which have not been DNA sampled and genotyped is described. When embryo transfer technology is used, the genetic dam needs to be included as a possible dam when considering parentage. If some fixed effects which depend on the parent are missing, these can be sampled similarly to parentage, and this may improve the evaluation if certain assumptions are met. A method to provide a likely list of parents, the ‘fitted pedigree’, which is based on the most likely parents, but modified to reduce the occurrence of unlikely family sets (e.g. very large litters) is also presented. The use of these methods will enhance the practical application of DNA parenting when used in conjunction with genetic evaluation.
The potential of microarrays to assist shrimp breeding and production: a review
Authors: Wilson K. J. , de la Vega E.
Published: August 2005

The shrimp aquaculture industry is a relatively new livestock industry, having developed over the past 30 years. Thus, it is poised to take advantage of new technologies from the outset of selective breeding programs. This contrasts with long established livestock industries, where there are already highly specialised breeds. This review focuses specifically on the potential application of microarrays to shrimp breeding. Potential applications of microarrays in selective breeding programs are summarised. Microarrays can be used as a rapid means to generate molecular markers for genetic linkage mapping, and genetic maps have been constructed for yeast, Arabidopsis and barley using microarray technology. Microarrays can also be used in the hunt for candidate genes affecting particular traits, leading to development of perfect markers for these traits (i.e. causative mutations). However, this requires that microarray analysis be combined with genetic linkage mapping, and that substantial genomic information is available for the species in question. A novel application of microarrays is to treat gene expression as a quantitative trait in itself and to combine this with linkage mapping to identify quantitative trait loci controlling the levels of gene expression; this approach may identify higher level regulatory genes in specific pathways. Finally, patterns of gene expression observed using microarrays may themselves be treated as phenotypic traits in selection programs (e.g. a particular pattern of gene expression might be indicative of a disease tolerant individual). Microarrays are now being developed for a number of shrimp species in laboratories around the world, primarily with a focus on identifying genes involved in the immune response. However, at present, there is no central repository of shrimp genomic information, which limits the rate at which shrimp genomic research can be progressed. The application of microarrays to shrimp breeding will be extremely limited until there is a shared repository of genomic information for shrimp, and the collective will and resources to develop comprehensive genomic tools for shrimp.
Genetics research in the Cooperative Research Centre for Cattle and Beef Quality
Authors: Burrow H. M. , Bindon B. M.
Published: August 2005

In its first 7-year term, the Cooperative Research Centre (CRC) for the Cattle and Beef Industry (Meat Quality) identified the genetic and non-genetic factors that impacted on beef eating quality. Following this, the CRC for Cattle and Beef Quality was established in 1999 to identify the consequences of improving beef eating quality and feed efficiency by genetic and non-genetic means on traits other than carcass and beef quality. The new CRC also had the responsibility to incorporate results from the first Beef CRC in national schemes such as BREEDPLAN (Australia’s beef genetic evaluation scheme) and Meat Standards Australia (Australia’s unique meat grading scheme that guarantees the eating quality of beef). This paper describes the integrated research programs and their results involving molecular and quantitative genetics, meat science, growth and nutrition and industry economics in the Beef CRC’s second phase (1999–2006) and the rationale for the individual genetics programs established. It summarises the planned scientific and beef industry outcomes from each of these programs and also describes the development and/or refinement by CRC scientists of novel technologies targeting increased genetic gains through enhanced measurement and recording in beef industry herds, thereby ensuring industry use of CRC results.
Biotechnologies and their potential impact on animal breeding and production: a review
Authors: Raadsma H. W. , Tammen I.
Published: August 2005

Recent developments in mammalian biotechnologies that have been driven largely by medical bioscience, offer new opportunities for livestock industries. Major impacts may be expected in the area of reproductive, genomic and cell technologies that could lead to improved animal breeding strategies or animal production and health applications. In particular, the use of advanced reproductive technologies to select animals at very early stages of life, possibly as early as a 4-day embryo, combined with genomic technologies to predict genetic merit, could lead to significantly increased rates of genetic gain. Such advanced animal breeding technologies will depend strongly on conventional quantitative genetic evaluation systems. Genetic modification in the near future will offer targeted animal improvement options for control of health and production. Long-term impact of genetic modification on animal production systems will depend on consumer acceptance, and its perception by social, environmental and animal welfare groups. However, the opportunity to develop animal products beyond conventional boundaries may prove too attractive with genetic modification eventually being accepted as the norm. The naturally synergistic effect of ex vivo transgenic modification of embryo stem cell or somatic cell lines, combined with nuclear transfer present potentially high value propositions for development of novel and high value products. Opportunities for the mass production of elite males for use in extensive animal production systems will be possible.
Breeding by numbers: an ancient endeavour that still resonates in the exciting era of functional genomics
Authors: Nicholas F. W.
Published: August 2005

‘Breeding by numbers’ is the term commonly used to deride the practical application of quantitative genetics. However, even the most subjective methods of assessment of breeding stock actually involve breeding by numbers: whenever a judge has to choose a winner and a runner-up from a line of animals, he/she evaluates each animal for each of several traits, then assesses the relative importance of each trait, and then ranks the animals on a weighted sum of the traits, where the weights reflect relative importance. And if the aim is to rank animals in terms of how good their offspring will be, then the mental gymnastics also involves unwittingly taking account of all the relevant genetic and phenotypic parameters (heritabilities and correlations) of each of the traits and combinations of traits. This has been the situation for as long as animals have been domesticated. The only thing that is different with the quantitative-genetics approach is that the guessing and mental gymnastics inherent in the traditional method of assessment have been replaced by use of a transparent set of numbers that reflect actual performance (or subjective assessment of certain traits) that is directly relevant to profitability, followed by a set of calculations using the best-available estimates of genetic and phenotypic parameters, ending up with the best-available prediction of how the offspring of each animal will perform. Having entered the genomics age, humans will still be breeding animals by numbers in the decades to come, but the numbers will be informed by knowledge of the action of thousands of individual genes.
Australian Spermacoce (Rubiaceae: Spermacoceae). I. Northern Territory
Authors: Harwood R. , Dessein S.
Published: August 2005

Northern Territory Spermacoce are revised, with 53 native species recognised, as well as four introduced species. Thirty-nine new species are described, and S. laevigata var. hispida Benth. is raised to species level. Lectotypes are chosen for all Australian Spermacoce names that had any ambiguity regarding types. Key features used to distinguish species, such as the appendages that occur on the inside of the corolla lobes and various seed characteristics, are illustrated and discussed. A key to all the Spermacoce species occurring in the Northern Territory is provided, along with distribution maps.
Inhibition of Single Minded 2 gene expression mediates tumor-selective apoptosis and differentiation in human colon cancer cells
Authors: Aleman Mireille J. , DeYoung Maurice Phil , Tress Matthew , Keating Patricia , Perry Gary W. , Narayanan Ramaswamy
Published: August 2005

A Down's syndrome associated gene, Single Minded 2 gene short form (
SIM2-s
), is specifically expressed in colon tumors but not in the normal colon. Antisense inhibition of
SIM2-s
in a RKO-derived colon carcinoma cell line causes growth inhibition, apoptosis, and inhibition of tumor growth in a nude mouse tumoriginicity model. The mechanism of cell death in tumor cells is unclear. In the present study, we investigated the pathways underlying apoptosis. Apoptosis was seen in a tumor cell-specific manner in RKO cells but not in normal renal epithelial cells, despite inhibition of SIM2-s expression in both of these cells by the antisense. Apoptosis was depended on WT p53 status and was caspase-dependent; it was inhibited by a pharmacological inhibitor of mitogen-activated protein kinase activity. Expression of a key stress response gene, growth arrest and DNA damage gene (
GADD
)
45
α, was up-regulated in antisense-treated tumor cells but not in normal cells. In an isogenic RKO cell line expressing stable antisense RNA to
GADD45
α, a significant protection of the antisense-induced apoptosis was seen. Whereas antisense-treated RKO cells did not undergo cell cycle arrest, several markers of differentiation were deregulated, including alkaline phosphatase activity, a marker of terminal differentiation. Protection of apoptosis and block of differentiation showed a correlation in the RKO model. Our results support the tumor cell-selective nature of
SIM2-s
gene function, provide a direct link between SIM2-s and differentiation, and may provide a model to identify SIM2-s targets.
Thyroid hormone controls multiple independent programs required for limb development in
Xenopus laevis
metamorphosis
Authors: Brown Donald D. , Cai Liquan , Das Biswajit , Marsh-Armstrong Nicholas , Schreiber Alexander M. , Juste Rejeanne
Published: August 2005

Thyroid hormone (TH) is required for limb development in
Xenopus laevis
. Specific cell types in the growing limb were targeted for expression of a dominant negative form of the TH receptor by sperm-mediated transgenesis. Limb muscle development, the innervation of muscle from the spinal cord, and cartilage growth can be inhibited without affecting patterning of the limb or differentiation of other cell types. Remodeling of the skin occurs late in metamorphosis after the limb has formed. The coordination of these independent programs is affected in part by the control that TH exerts over DNA replication in all cell types of the limb.
Inhibiting farnesylation of progerin prevents the characteristic nuclear blebbing of Hutchinson-Gilford progeria syndrome
Authors: Capell Brian C. , Erdos Michael R. , Madigan James P. , Fiordalisi James J. , Varga Renee , Conneely Karen N. , Gordon Leslie B. , Der Channing J. , Cox Adrienne D. , Collins Francis S.
Published: August 2005

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder that is characterized by dramatic premature aging and accelerated cardiovascular disease. HGPS is almost always caused by a
de novo
point mutation in the lamin A gene (
LMNA
) that activates a cryptic splice donor site, producing a truncated mutant protein termed “progerin.” WT prelamin A is anchored to the nuclear envelope by a farnesyl isoprenoid lipid. Cleavage of the terminal 15 aa and the farnesyl group releases mature lamin A from this tether. In contrast, this cleavage site is deleted in progerin. We hypothesized that retention of the farnesyl group causes progerin to become permanently anchored in the nuclear membrane, disrupting proper nuclear scaffolding and causing the characteristic nuclear blebbing seen in HGPS cells. Also, we hypothesized that blocking farnesylation would decrease progerin toxicity. To test this hypothesis, the terminal CSIM sequence in progerin was mutated to SSIM, a sequence that cannot be farnesylated. SSIM progerin relocalized from the nuclear periphery into nucleoplasmic aggregates and produced no nuclear blebbing. Also, blocking farnesylation of authentic progerin in transiently transfected HeLa, HEK 293, and NIH 3T3 cells with farnesyltransferase inhibitors (FTIs) restored normal nuclear architecture. Last, treatment of both early- and late-passage human HGPS fibroblasts with FTIs resulted in significant reductions in nuclear blebbing. Our results suggest that treatment with FTIs represents a potential therapy for patients with HGPS.
RNA–protein recognition: Single-residue ultrafast dynamical control of structural specificity and function
Authors: Xia Tianbing , Wan Chaozhi , Roberts Richard W. , Zewail Ahmed H.
Published: August 2005

The transcription antiterminator N protein from bacteriophage λ uses its arginine-rich motif to specifically bind a stem-loop RNA hairpin (boxB) as a bent α-helix. A single stacking interaction between a tryptophan (Trp-18) and an adenosine (A7) in the RNA loop is robust and necessary for antitermination activityin vivo. Previously, femtosecond fluorescence up-conversion experiments from this laboratory indicated that the N/boxBcomplex exists in a dynamical two-state equilibrium between stacked and unstacked conformations and that the extent of stacking depends on the identity of peptide residues 14 and 15. In the present work, we have combined transient absorption and fluorescence up-conversion to determine the nature of interactions responsible for this sequence-dependent behavior. Analysis of mutant complexes supports the idea that the β-carbon of residue 14 enforces the stacked geometry by hydrophobic interaction with the ribose of A7, whereas a positive charge at this residue plays only a secondary role. A positive charge at position 15 substantially disfavors the stacked state but retains much of the binding energy. Remarkably,in vivoantitermination experiments show strong correlation with our femtosecond dynamics, demonstrating how conformational interplay can control the activity of a macromolecular machine.
A
Drosophila
DEG/ENaC channel subunit is required for male response to female pheromones
Authors: Lin Heping , Mann Kevin J. , Starostina Elena , Kinser Ronald D. , Pikielny Claudio W.
Published: August 2005

Odorants and pheromones as well as sweet- and bitter-tasting small molecules are perceived through activation of G protein-coupled chemosensory receptors. In contrast, gustatory detection of salty and sour tastes may involve direct gating of sodium channels of the DEG/ENaC family by sodium and hydrogen ions, respectively. We have found that
ppk25
, a
Drosophila melanogaster
gene encoding a DEG/ENaC channel subunit, is expressed at highest levels in the male appendages responsible for gustatory and olfactory detection of female pheromones: the legs, wings, and antennae. Mutations in the
ppk25
gene reduce or even abolish male courtship response to females in the dark, conditions under which detection of female pheromones is an essential courtship-activating sensory input. In contrast, the same mutations have no effect on other behaviors tested. Importantly,
ppk25
mutant males that show no response to females in the dark execute all of the normal steps of courtship behavior in the presence of visible light, suggesting that
ppk25
is required for activation of courtship behavior by chemosensory perception of female pheromones. Finally, a
ppk25
mutant allele predicted to encode a truncated protein has dominant-negative properties, suggesting that the normal Ppk25 protein acts as part of a multiprotein complex. Together, these results indicate that
ppk25
is necessary for response to female pheromones by
D. melanogaster
males, and suggest that members of the DEG/ENaC family of genes play a wider role in chemical senses than previously suspected.
Responsiveness of certain agronomic weed species to arbuscular mycorrhizal fungi
Authors: Vatovec C. , Jordan N. , Huerd S.
Published: August 2005

Arbuscular mycorrhizal fungi (AMF) are plant root symbionts that provide many benefits to crop production and agro-ecosystem function; therefore, management of AMF is increasingly seen as important to ecological farming. Agronomic weeds that form a symbiotic relationship with AMF can increase diversity and abundance of agronomically beneficial AMF taxa. Also, AMF can strongly affect plant community composition, and may thus provide some degree of biological control for weeds. Therefore, relationships between weeds and AMF have a dual significance in ecological farming, but are relatively unexamined. In glasshouse experiments, seedlings of 14 agronomic weed species were grown in the presence or absence of AMF inocula sampled from each of three types of cropping systems: organic, transitional-organic or high-input/conventional. For each weed species, AMF root colonization rates and growth responses to AMF were assessed. On the basis of observed colonization levels, the species were classified as strong hosts (five species), weak hosts (three) and non-host species (six). Among species, biomass responses to AMF were highly variable. Strong hosts showed more positive responses to AMF than weak hosts, although the range of responses was great. Non-hosts did not suffer consistent negative biomass responses to AMF, although strong biomass reductions were noted for certain species–inoculum combinations. Biomass responses to inocula from different cropping systems varied significantly among weed species in one of two experiments. Results suggest that weed–AMF interactions can affect weed community dynamics. We recommend investigation of these interactions in agro-ecosystems that use management methods likely to intensify weed–AMF interactions, such as conservation tillage and cover cropping.
A Rac switch regulates random versus directionally persistent cell migration
Authors: Pankov Roumen , Endo Yukinori , Even-Ram Sharona , Araki Masaru , Clark Katherine , Cukierman Edna , Matsumoto Kazue , Yamada Kenneth M.
Published: August 2005

Directional migration moves cells rapidly between points, whereas random migration allows cells to explore their local environments. We describe a Rac1 mechanism for determining whether cell patterns of migration are intrinsically random or directionally persistent. Rac activity promoted the formation of peripheral lamellae that mediated random migration. Decreasing Rac activity suppressed peripheral lamellae and switched the cell migration patterns of fibroblasts and epithelial cells from random to directionally persistent. In three-dimensional rather than traditional two-dimensional cell culture, cells had a lower level of Rac activity that was associated with rapid, directional migration. In contrast to the directed migration of chemotaxis, this intrinsic directional persistence of migration was not mediated by phosphatidylinositol 3′-kinase lipid signaling. Total Rac1 activity can therefore provide a regulatory switch between patterns of cell migration by a mechanism distinct from chemotaxis.
Elevated expression of MITF counteracts B-RAF–stimulated melanocyte and melanoma cell proliferation
Authors: Wellbrock Claudia , Marais Richard
Published: August 2005

The protein kinase B-RAF is a human oncogene that is mutated in ∼70% of human melanomas and transforms mouse melanocytes. Microphthalmia-associated transcription factor (MITF) is an important melanocyte differentiation and survival factor, but its role in melanoma is unclear. In this study, we show that MITF expression is suppressed by oncogenic B-RAF in immortalized mouse and primary human melanocytes. However, low levels of MITF persist in human melanoma cells harboring oncogenic B-RAF, suggesting that additional mechanisms regulate its expression. MITF reexpression in B-RAF–transformed melanocytes inhibits their proliferation. Furthermore, differentiation-inducing factors that elevate MITF expression in melanoma cells inhibit their proliferation, but when MITF up-regulation is prevented by RNA interference, proliferation is not inhibited. These data suggest that MITF is an antiproliferation factor that is down-regulated by B-RAF signaling and that this is a crucial event for the progression of melanomas that harbor oncogenic B-RAF.
The positioning and segregation of apical cues during epithelial polarity establishment in Drosophila
Authors: Harris Tony J.C. , Peifer Mark
Published: August 2005

Cell polarity is critical for epithelial structure and function. Adherens junctions (AJs) often direct this polarity, but we previously found that Bazooka (Baz) acts upstream of AJs as epithelial polarity is first established in Drosophila. This prompted us to ask how Baz is positioned and how downstream polarity is elaborated. Surprisingly, we found that Baz localizes to an apical domain below its typical binding partners atypical protein kinase C (aPKC) and partitioning defective (PAR)-6 as the Drosophila epithelium first forms. In fact, Baz positioning is independent of aPKC and PAR-6 relying instead on cytoskeletal cues, including an apical scaffold and dynein-mediated basal-to-apical transport. AJ assembly is closely coupled to Baz positioning, whereas aPKC and PAR-6 are positioned separately. This forms a stratified apical domain with Baz and AJs localizing basal to aPKC and PAR-6, and we identify specific mechanisms that keep these proteins apart. These results reveal key steps in the assembly of the apical domain in Drosophila.
Assembly and trafficking of caveolar domains in the cell
Authors: Tagawa Akiko , Mezzacasa Anna , Hayer Arnold , Longatti Andrea , Pelkmans Lucas , Helenius Ari
Published: August 2005

Using total internal reflection fluorescence microscopy (TIR-FM), fluorescence recovery after photobleaching (FRAP), and other light microscopy techniques, we analyzed the dynamics, the activation, and the assembly of caveolae labeled with fluorescently tagged caveolin-1 (Cav1). We found that when activated by simian virus 40 (SV40), a nonenveloped DNA virus that uses caveolae for cell entry, the fraction of mobile caveolae was dramatically enhanced both in the plasma membrane (PM) and in the caveosome, an intracellular organelle that functions as an intermediate station in caveolar endocytosis. Activation also resulted in increased microtubule (MT)-dependent, long-range movement of caveolar vesicles. We generated heterokaryons that contained GFP- and RFP-tagged caveolae by fusing cells expressing Cav1-GFP and -RFP, respectively, and showed that even when activated, individual caveolar domains underwent little exchange of Cav1. Only when the cells were subjected to transient cholesterol depletion, did the caveolae domain exchange Cav1. Thus, in contrast to clathrin-, or other types of coated transport vesicles, caveolae constitute stable, cholesterol-dependent membrane domains that can serve as fixed containers through vesicle traffic. Finally, we identified the Golgi complex as the site where newly assembled caveolar domains appeared first.
Regulatory mechanisms required for DE-cadherin function in cell migration and other types of adhesion
Authors: Pacquelet Anne , Rørth Pernille
Published: August 2005

Cadherin-mediated adhesion can be regulated at many levels, as demonstrated by detailed analysis in cell lines. We have investigated the requirements for Drosophila melanogaster epithelial (DE) cadherin regulation in vivo. Investigating D. melanogaster oogenesis as a model system allowed the dissection of DE-cadherin function in several types of adhesion: cell sorting, cell positioning, epithelial integrity, and the cadherin-dependent process of border cell migration. We generated multiple fusions between DE-cadherin and α-catenin as well as point-mutated β-catenin and analyzed their ability to support these types of adhesion. We found that (1) although linking DE-cadherin to α-catenin is essential, regulation of the link is not required in any of these types of adhesion; (2) β-catenin is required only to link DE-cadherin to α-catenin; and (3) the cytoplasmic domain of DE-cadherin has an additional specific function for the invasive migration of border cells, which is conserved to other cadherins. The nature of this additional function is discussed.
IL-18 with IL-2 protects against Strongyloides venezuelensis infection by activating mucosal mast cell–dependent type 2 innate immunity
Authors: Sasaki Yuki , Yoshimoto Tomohiro , Maruyama Haruhiko , Tegoshi Tatsuya , Ohta Nobuo , Arizono Naoki , Nakanishi Kenji
Published: August 2005

C57BL/6 (B6) and B6 background STAT6−/− mice pretreated with IL-18 plus IL-2 showed prominent intestinal mastocytosis and rapidly expelled implanted adult worms of the gastrointestinal nematode Strongyloides venezuelensis. In contrast, identically pretreated mast cell–deficient W/Wv mice failed to do so. Thus, activated mucosal mast cells (MMC) are crucial for parasite expulsion. B6 mice infected with S. venezuelensis third-stage larvae (L3) completed parasite expulsion by day 12 after infection, whereas IL-18−/− or IL-18Rα−/− B6 mice exhibited marked impairment in parasite expulsion, suggesting a substantial contribution of IL-18–dependent MMC activation to parasite expulsion. Compared with IL-18−/− or IL-18Rα−/− mice, S. venezuelensis L3–infected STAT6−/− mice have poorly activated MMC and sustained infection; although their IL-18 production is normal. Neutralization of IL-18 and IL-2 further reduces expulsion in infected STAT6−/− mice. These results suggest that collaboration between IL-18–dependent and Th2 cell–dependent mastocytosis is important for prompt parasite expulsion.
Protein phosphatase subunit G5PR is needed for inhibition of B cell receptor–induced apoptosis
Authors: Xing Yan , Igarashi Hideya , Wang Xiaodan , Sakaguchi Nobuo
Published: August 2005

B cell receptor (BCR) cross-linking induces B cell proliferation and sustains survival through the phosphorylation-dependent signals. We report that a loss of the protein phosphatase component G5PR increased the activation-induced cell death (AICD) and thus impaired B cell survival. G5PR associates with GANP, whose expression is up-regulated in mature B cells of the peripheral lymphoid organs. To study G5PR function, the G5pr gene was conditionally targeted with the CD19-Cre combination (G5pr−/− mice). The G5pr−/− mice had a decreased number of splenic B cells (60% of the controls). G5pr−/− B cells showed a normal proliferative response to lipopolysaccharide or anti-CD40 antibody stimulation but not to BCR cross-linking with or without IL-4 in vitro. G5pr−/− B cells did not show abnormalities in the BCR-mediated activation of Erks and NF-κB, cyclin D2 induction, or Akt activation. However, G5pr−/− B cells were sensitive to AICD caused by BCR cross-linking. This was associated with an increased depolarization of the mitochondrial membrane and the enhanced activation of c-Jun NH2-terminal protein kinase and Bim. These results suggest that G5PR is required for the BCR-mediated proliferation associated with the prevention of AICD in mature B cells.
Ligand-dependent Linkage of the ATP Site to Inhibition Gate Closure in the KATP Channel
Authors: Li Lehong , Geng Xuehui , Yonkunas Michael , Su Anjey , Densmore Erik , Tang Pei , Drain Peter
Published: August 2005

Major advances have been made on the inhibition gate and ATP site of the Kir6.2 subunit of the KATP channel, but little is known about conformational coupling between the two. ATP site mutations dramatically disrupt ATP-dependent gating without effect on ligand-independent gating, observed as interconversions between active burst and inactive interburst conformations in the absence of ATP. This suggests that linkage between site and gate is conditionally dependent on ATP occupancy. We studied all substitutions at position 334 of the ATP site in Kir6.2ΔC26 that express in Xenopus oocytes. All substitutions disrupted ATP-dependent gating by 10-fold or more. Only positive-charged arginine or lysine at 334, however, slowed ligand-independent gating from the burst, and this was in some but not all patches. Moreover, the polycationic peptide protamine reversed the slowed gating from the burst of 334R mutant channels, and speeded the slow gating from the burst of wild-type SUR1/Kir6.2 in the absence of ATP. Our results support a two-step ligand-dependent linkage mechanism for Kir6.2 channels in which ATP-occupied sites function to electrostatically dissociate COOH-terminal domains from the membrane, then as in all Kir channels, free COOH-terminal domains and inner M2 helices transit to a lower energy state for gate closure.
Phospholipase C in Living Cells
Authors: Horowitz Lisa F. , Hirdes Wiebke , Suh Byung-Chang , Hilgemann Donald W. , Mackie Ken , Hille Bertil
Published: August 2005

We have further tested the hypothesis that receptor-mediated modulation of KCNQ channels involves depletion of phosphatidylinositol 4,5-bisphosphate (PIP2) by phosphoinositide-specific phospholipase C (PLC). We used four parallel assays to characterize the agonist-induced PLC response of cells (tsA or CHO cells) expressing M1 muscarinic receptors: translocation of two fluorescent probes for membrane lipids, release of calcium from intracellular stores, and chemical measurement of acidic lipids. Occupation of M1 receptors activates PLC and consumes cellular PIP2 in less than a minute and also partially depletes mono- and unphosphorylated phosphoinositides. KCNQ current is simultaneously suppressed. Two inhibitors of PLC, U73122 and edelfosine (ET-18-OCH3), can block the muscarinic actions completely, including suppression of KCNQ current. However, U73122 also had many side effects that were attributable to alkylation of various proteins. These were mimicked or occluded by prior reaction with the alkylating agent N-ethylmaleimide and included block of pertussis toxin–sensitive G proteins and effects that resembled a weak activation of PLC or an inhibition of lipid kinases. By our functional criteria, the putative PLC activator m-3M3FBS did stimulate PLC, but with a delay and an irregular time course. It also suppressed KCNQ current. The M1 receptor–mediated activation of PLC and suppression of KCNQ current were stopped by lowering intracellular calcium well below resting levels and were slowed by not allowing intracellular calcium to rise in response to PLC activation. Thus calcium release induced by PLC activation feeds back immediately on PLC, accelerating it during muscarinic stimulation in strong positive feedback. These experiments clarify important properties of receptor-coupled PLC responses and their inhibition in the context of the living cell. In each test, the suppression of KCNQ current closely paralleled the expected fall of PIP2. The results are described by a kinetic model.
Rod Sensitivity of Neonatal Mouse and Rat
Authors: Luo Dong-Gen , Yau King-Wai
Published: August 2005

We have measured the sensitivity of rod photoreceptors isolated from overnight dark–adapted mice of age P12 (neonate) through P45 (adult) with suction-pipette recording. During this age period, the dark current increased roughly in direct proportion to the length of the rod outer segment. In the same period, the flash sensitivity of rods (reciprocal of the half-saturating flash intensity) increased by ∼1.5-fold. This slight developmental change in sensitivity was not accentuated by dark adapting the animal for just 1 h or by increasing the ambient luminance by sixfold during the prior light exposure. The same small, age-dependent change in rod sensitivity was found with rat. After preincubation of the isolated retina with 9-cis-retinal, neonatal mouse rods showed the same sensitivity as adult rods, suggesting the presence of a small amount of free opsin being responsible for their lower sensitivity. The sensitivity of neonate rods could also be increased to the adult level by dark adapting the animal continuously for several days. By comparing the sensitivity of neonate rods in darkness to that of adult rods after light bleaches, we estimated that ∼1% of rod opsin in neonatal mouse was devoid of chromophore even after overnight dark adaptation. Overall, we were unable to confirm a previous report that a 50-fold difference in rod sensitivity existed between neonatal and adult rats.
Inhibition of IgE‐dependent Mouse Triphasic Cutaneous Reaction by a Boiling Water Fraction Separated from Mycelium of Phellinus linteus
Authors: Inagaki Naoki , Shibata Tomonori , Itoh Tomokazu , Suzuki Tomohiro , Tanaka Hiroyuki , Nakamura Tomoyuki , Akiyama Yukihito , Kawagishi Hirokazu , Nagai Hiroichi
Published: August 2005

Phellinus linteus, a mushroom, contains constituents that exhibit potent antitumor effects through activating immune cells. Recently, anti‐inflammatory and anti‐allergic properties of P. linteus extracts have also been implicated. In the present study, therefore, we separated the constituents of mycelium of P. linteus into five fractions—chloroform‐soluble (CF), ethyl acetate‐soluble (EA), methanol‐soluble (AE), water‐soluble (WA) and boiling water‐soluble (BW) fractions—and examined their suppressive effects on the IgE‐dependent mouse triphasic cutaneous reaction. The triphasic reaction was induced in the ear of BALB/c mice passively sensitized with anti‐dinitrophenol IgE by painting with 2,4‐dinitrofluorobenzene 24 h later. Ear swelling appeared triphasically with peak responses at 1 h, 24 h and 8 days after the challenge. ME, WA and BW given orally at a dose of 100 mg kg−1 significantly inhibited the first and second phase ear swelling, and BW also inhibited the third phase response. CF only inhibited the second phase. The inhibition by BW was the most potent and almost dose‐dependent at doses of 30–300 mg kg−1. BW also inhibited vascular permeability increase caused by passive cutaneous anaphylaxis and histamine, and ear swelling caused by tumor necrosis factor‐α. In contrast, BW apparently potentiated the production of interleukin‐4 and interferon‐γ from anti‐CD3‐stimulated mouse splenocytes. These results indicate that BW derived from mycelium of P. linteus contains some constituents with anti‐allergic as well as immunopotentiating properties.
Echinacea: a Miracle Herb against Aging and Cancer? Evidence In vivo in Mice
Authors: Miller Sandra C.
Published: August 2005

Echinacea has been viewed as an immunoenhancing herb since it became commercially available several years ago. Indeed, its medicinal significance is responsible for billions of dollars in worldwide sales annually. Unfortunately, most of the ‘evidence’ for the purported medicinal efficacy of Echinacea has been anecdotal and, moreover, to this day, there is no formal proof on how to achieve the best results—whether it should be consumed daily throughout life as a prophylactic; consumed by either young or old; or consumed after diseases, such as cancer, have taken hold. Our work over the past 5 years has led to conclusive answers to some of these questions, at least in mice. Our results have shown that daily consumption of Echinacea is indeed prophylactic, extends the life span of aging mice, significantly abates leukemia and extends the life span of leukemic mice. Given that humans are 97% genetically common with mice and that virtually all our basic physiology is identical, it is neither unjustified to extrapolate these observations to humans nor would it be an arduous task to perform many of these studies in humans, thus establishing viable scientific evidence replacing the anecdotal.