Aging is a universal but poorly understood biological process. Free
radicals accumulate with age and have been proposed to be a major cause
of aging. We measured genome-wide changes in transcript levels as a
function of age in Drosophila melanogaster and compared
these changes with those caused by paraquat, a free-radical generator.
A number of genes exhibited changes in transcript levels with both age
and paraquat treatment. We also found genes whose transcript levels
changed with age but not with paraquat treatment. This study suggests
that free radicals play an important role in regulating transcript
levels in aging but that they are not the only factors. This
genome-wide survey also identifies candidates for molecular markers of
Bordetella pertussis, the causative agent of
whooping cough, has many well-studied virulence factors and a
characteristic clinical presentation. Despite this information, it is
not clear how B. pertussis interaction with host cells
leads to disease. In this study, we examined the interaction of
B. pertussis with a human bronchial epithelial cell line
(BEAS-2B) and measured host transcriptional profiles by using
high-density DNA microarrays. The early transcriptional response to
this pathogen is dominated by altered expression of cytokines,
DNA-binding proteins, and NFκB-regulated genes. This previously
unrecognized response to B. pertussis was modified in
similar but nonidentical fashions by the antiinflammatory agents
dexamethasone and sodium salicylate. Cytokine protein expression was
confirmed, as was neutrophil chemoattraction. We show that B.
pertussis induces mucin gene transcription by BEAS-2B cells
then counters this defense by using mucin as a binding substrate. A set
of genes is described for which the catalytic activity of pertussis
toxin is both necessary and sufficient to regulate transcription. Host
genomic transcriptional profiling, in combination with functional
assays to evaluate subsequent biological events...
The biological basis for developmental dyslexia remains unknown.
Research has suggested that a fundamental deficit in dyslexia is the
inability to process sensory input that enters the nervous system
rapidly and that deficits in processing rapid acoustic information are
associated with impaired reading. Functional magnetic resonance imaging
(fMRI) was used to identify the brain basis of rapid acoustic
processing in normal readers and to discover the status of that
response in dyslexic readers. Normal readers showed left prefrontal
activity in response to rapidly changing, relative to slowly changing,
nonlinguistic acoustic stimuli. Dyslexic readers showed no differential
left frontal response. Two dyslexic readers participated in a
remediation program and showed increased activity in left prefrontal
cortex after training. These fMRI results identify left prefrontal
regions as normally being sensitive to rapid relative to slow acoustic
stimulation, insensitive to the difference between such stimuli in
dyslexic readers, and plastic enough in adulthood to develop such
differential sensitivity after intensive training.
The ability of monkeys to follow the gaze of other individuals is a
matter of debate in many behavioral studies. Physiological studies have
shown that in monkeys, as in humans, there are neural correlates of eye
direction detection. There is little evidence at the behavioral level,
however, of the presence and development of such abilities in monkeys.
The aim of the present study was to assess in juveniles and adult
pig-tailed macaques (Macaca nemestrina) the capacity to
use eye cues only to follow the gaze of an experimenter. Biological
stimuli (head, eye, and trunk movements) were presented by an
experimenter to 2 adult monkeys with their heads restrained (Experiment
1) and to 11 monkeys of different ages, free to move in their home
cages (Experiment 2). A nonbiological stimulus served as a control.
Results showed that macaques can follow the gaze of the experimenter by
using head/eye and eye cues alone. Trunk movements and nonbiological
stimuli did not significantly elicit similar reactions. Juvenile
monkeys were not able to orient their attention on the basis of eye
cues alone. In general, gaze following was more frequent in adults than
in juveniles. Like in humans, however, such abilities in macaques
dramatically improve with age suggesting that the transition to
adulthood is a crucial period in the development of gaze-following
Perou, Charles M.; Jeffrey, Stefanie S.; van de Rijn, Matt; Rees, Christian A.; Eisen, Michael B.; Ross, Douglas T.; Pergamenschikov, Alexander; Williams, Cheryl F.; Zhu, Shirley X.; Lee, Jeffrey C. F.; Lashkari, Deval; Shalon, Dari; Brown, Patrick O.; Bo
Fonte: The National Academy of SciencesPublicador: The National Academy of Sciences
cDNA microarrays and a clustering algorithm were used to identify patterns of gene expression in human mammary epithelial cells growing in culture and in primary human breast tumors. Clusters of coexpressed genes identified through manipulations of mammary epithelial cells in vitro also showed consistent patterns of variation in expression among breast tumor samples. By using immunohistochemistry with antibodies against proteins encoded by a particular gene in a cluster, the identity of the cell type within the tumor specimen that contributed the observed gene expression pattern could be determined. Clusters of genes with coherent expression patterns in cultured cells and in the breast tumors samples could be related to specific features of biological variation among the samples. Two such clusters were found to have patterns that correlated with variation in cell proliferation rates and with activation of the IFN-regulated signal transduction pathway, respectively. Clusters of genes expressed by stromal cells and lymphocytes in the breast tumors also were identified in this analysis. These results support the feasibility and usefulness of this systematic approach to studying variation in gene expression patterns in human cancers as a means to dissect and classify solid tumors.
As part of the European Sinorhizobium meliloti (strain 1021) chromosome sequencing project, four genomic bacterial artificial chromosome (BAC) libraries have been constructed, one of which was mainly used for chromosome mapping. This library consists of 1,824 clones with an average insert size of 80 kilobases and represents approximately 20-fold total genome coverage [6.8 megabases (Mbs)]. PCR screening of 384 BAC clones with 447 chromosomal markers (PCR primer pairs), consisting of 73 markers representing 118 genes (40 individual genes and 78 genes clustered in 23 operons), two markers from the rrn operon (three loci), four markers from insertion sequences (≈16 loci) and 368 sequence-tagged sites allowed the identification of 252 chromosomal BAC clones and the construction of a high-density physical map of the whole 3.7-Mb chromosome of S. meliloti. An average of 5.5 overlapping and colinear BAC clones per marker, correlated with a low rate of deleted or rearranged clones (0.8%) indicate a solid BAC contigation and a correct mapping. Systematic blastx analysis of sequence-tagged site marker sequences allowed prediction of a biological function for a number of putative ORFs. Results are available at http://www-recomgen.univ-rennes1.fr/meliloti. This map...
The Cercospora nicotianae SOR1 (singlet oxygen resistance) gene was identified previously as a gene involved in resistance of this fungus to singlet-oxygen-generating phototoxins. Although homologues to SOR1 occur in organisms in four kingdoms and encode one of the most highly conserved proteins yet identified, the precise function of this protein has, until now, remained unknown. We show that SOR1 is essential in pyridoxine (vitamin B6) synthesis in C. nicotianae and Aspergillus flavus, although it shows no homology to previously identified pyridoxine synthesis genes identified in Escherichia coli. Sequence database analysis demonstrated that organisms encode either SOR1 or E. coli pyridoxine biosynthesis genes, but not both, suggesting that there are two divergent pathways for de novo pyridoxine biosynthesis in nature. Pathway divergence appears to have occurred during the evolution of the eubacteria. We also present data showing that pyridoxine quenches singlet oxygen at a rate comparable to that of vitamins C and E, two of the most highly efficient biological antioxidants, suggesting a previously unknown role for pyridoxine in active oxygen resistance.
Receptor proteins in both eukaryotic and prokaryotic cells have been found to form two-dimensional clusters in the plasma membrane. In this study, we examine the proposition that such clusters might show coordinated responses because of the spread of conformational states from one receptor to its neighbors. A Monte Carlo simulation was developed in which receptors flipped in probabilistic fashion between an active and an inactive state. Conformational energies depended on (i) ligand binding, (ii) a chemical modification of the receptor conferring adaptation, and (iii) the activity of neighboring receptors. Rate constants were based on data from known biological receptors, especially the bacterial Tar receptor, and on theoretical constraints derived from an analogous Ising model. The simulated system showed a greatly enhanced sensitivity to external signals compared with a corresponding set of uncoupled receptors and was operational over a much wider range of ambient concentrations. These and other properties should make a lattice of conformationally coupled receptors ideally suited to act as a “nose” by which a cell can detect and respond to extracellular stimuli.
The interaction of a ligand with a protein occurs at a local site
(the binding site) and involves only a few residues; however, the
effects of that interaction are often propagated to remote locations.
The chain of events initiated by binding provides the basis for
fundamental biological phenomena such as allosterism, signal
transduction, and structural-stability modification. In this paper, a
structure-based statistical thermodynamic approach is presented and
used to predict the propagation of the stabilization effects triggered
by the binding of the monoclonal antibody D1.3 to hen egg white
lysozyme. Previously, Williams et al. [Williams,
D. C., Benjamin, D. C., Poljak, R. J. & Rule, G. S.
(1996) J. Mol. Biol. 257, 866–876] showed that the
binding of this antibody affects the stability of hen egg white
lysozyme and that the binding effects propagate to a selected number of
residues at remote locations from the binding epitope. In this paper,
we show that this phenomenon can be predicted from structure. The
formalism presented here permits the identification of the structural
path followed by cooperative interactions that originate at the binding
site. It is shown that an important condition for the propagation of
binding effects to distal regions is the presence of a significant
fraction of residues with low structural stability in the uncomplexed
binding site. A survey of protein structures indicates that many
binding sites have a dual character and are defined by regions of high
and low structural stabilities. The low-stability regions might be
involved in the transmission of binding information to other regions in
Hepatocyte nuclear factors 3 (HNF-3) belong to an evolutionarily conserved family of transcription factors that are critical for diverse biological processes such as development, differentiation, and metabolism. To study the physiological role of HNF-3α, we generated mice that lack HNF-3α by homologous recombination in embryonic stem cells. Mice homozygous for a null mutation in the HNF-3α gene develop a complex phenotype that is characterized by abnormal feeding behavior, progressive starvation, persistent hypoglycemia, hypotriglyceridemia, wasting, and neonatal mortality between days 2 and 14. Hypoglycemia in HNF-3α-null mice leads to physiological counter-regulatory responses in glucocorticoid and growth hormone production and an inhibition of insulin secretion but fails to stimulate glucagon secretion. Glucagon-producing pancreatic alpha cells develop normally in HNF-3α−/− mice, but proglucagon mRNA levels are reduced 50%. Furthermore, the transcriptional levels of neuropeptide Y are also significantly reduced shortly after birth, implying a direct role of HNF-3α in the expression of these genes. In contrast, mRNA levels were increased in HNF-3 target genes phosphofructo-2-kinase/fructose-2,6-bisphophatase, insulin growth factor binding protein-1...
Biological information-processing systems, such as populations of sensory and motor neurons, may use correlations between the firings of individual elements to obtain lower noise levels and a systemwide performance improvement in the dynamic range or the signal-to-noise ratio. Here, we implement such correlations in networks of coupled integrate-and-fire neurons using inhibitory coupling and demonstrate that this can improve the system dynamic range and the signal-to-noise ratio in a population rate code. The improvement can surpass that expected for simple averaging of uncorrelated elements. A theory that predicts the resulting power spectrum is developed in terms of a stochastic point-process model in which the instantaneous population firing rate is modulated by the coupling between elements.
Supramolecular self-assembly is an integral step in the formation of many biological structures. Here we report a DNA pentaplex that derives from a metal-assisted, hydrogen bond-mediated self-assembly process. In particular, cesium ions are found to induce pentameric assembly of DNA bearing the nonstandard nucleobase iso-guanine. The pentaplex was designed by using a simple algorithm to predict nucleobase structural requirements within a quintet motif. The design principles are general and should extend to complexes beyond pentaplex. Structures exhibiting molecularities of five or more were previously accessible to peptides, but not nucleic acids.
Technological advances over the past 10 years have generated powerful tools for parallel analysis of complex biological problems. Among these new technologies, DNA arrays have provided an important experimental approach for identifying changes in the levels of individual mRNA molecules during important cellular transitions. However, cellular behavior is dictated not by mRNA levels, but by the proteins translated from the individual mRNA species. We report a high-throughput method for simultaneously monitoring the translation state and level of individual mRNA species. Messenger RNAs from resting and mitogenically activated fibroblasts were separated, according to degree of ribosome loading, into well-translated and under-translated pools. cDNA probes generated from these fractions were used to interrogate cDNA arrays. Among approximately 1,200 genes analyzed, less than 1% were found to be translationally regulated in response to mitogenic activation, demonstrating the strong selectivity of this regulatory mechanism. This high-throughput approach is shown to be an effective tool for superimposing translation profile on mRNA level for large numbers of genes, as well as for identifying translationally regulated genes for further study.
Homeostasis, the creation of a stabilized internal milieu, is ubiquitous in biological evolution, despite the entropic cost of excluding noise information from a region. The advantages of stability seem self evident, but the alternatives are not so clear. This issue was studied by means of numerical experiments on a simple evolution model: a population of Boolean network “organisms” selected for performance of a curve-fitting task while subjected to noise. During evolution, noise sensitivity increased with fitness. Noise exclusion evolved spontaneously, but only if the noise was sufficiently unpredictable. Noise that was limited to one or a few stereotyped patterns caused symmetry breaking that prevented noise exclusion. Instead, the organisms incorporated the noise into their function at little cost in ultimate fitness and became totally noise dependent. This “noise imprinting” suggests caution when interpreting apparent adaptations seen in nature. If the noise was totally random from generation to generation, noise exclusion evolved reliably and irreversibly, but if the noise was correlated over several generations, maladaptive selection of noise-dependent traits could reverse noise exclusion, with catastrophic effect on population fitness. Noise entering the selection process rather than the organism had a different effect: adaptive evolution was totally abolished above a critical noise amplitude...
Drug activity is often studied in well controlled and characterized cellular environments in vitro. However, the biology of cells in culture is only a part of the tissue behavior in vivo. Quantitative studies of the dose response to drugs in vivo have been limited by the inability to reliably determine or predict the concentrations achieved in tissues. We developed a method to study the dose response of injured arteries to exogenous heparin in vivo by providing steady and predictable arterial levels of drug. Controlled-release devices were fabricated to direct heparin uniformly and at a steady rate to the adventitial surface of balloon-injured rat carotid arteries. We predicted the distribution of heparin throughout the arterial wall by using computational simulations of intravascular drug binding and transport, and we correlated these concentrations with the biologic response of the tissues. This allowed the estimation of the arterial concentration of heparin required to maximally inhibit intimal hyperplasia after injury in vivo, 0.3 mg/ml. This estimation of the required concentration of drug seen by a specific tissue is independent of the route of administration and holds for all forms of drug release. In this way we may now be able to evaluate the potential of widely disparate forms of drug release and to finally create some rigorous criteria by which to guide the development of particular delivery strategies for local diseases.
The interaction between two regulatory proteins plays a crucial role in the control of several biological events, including gene transcription. In this report, we demonstrate that the interaction between the cellular sequence-specific single-stranded DNA binding protein Purα and the HIV type 1 (HIV-1) Tat protein is mediated by specific ribonucleic acids. The region of Tat that is important for its interaction with Purα includes the region demonstrated to bind Tat’s viral RNA target, TAR. A 10-nucleotide GC-rich consensus sequence identified in RNAs associated with Purα derived from human U-87MG cells plays an important role in the Purα:Tat interaction as examined by an in vitro reconstitution assay. Furthermore, expression of the Purα-associated RNA in these cells enhances transcriptional activation of the HIV-1 promoter by Tat and Purα.
Corticotropin-releasing hormone (CRH) is widely recognized as the primary mediator of the neuroendocrine and behavioral responses to stress, including stress-induced anxiety. The biological activity of CRH and other mammalian CRH-like peptides, such as urocortin, may be modulated by CRH-binding protein (CRH-BP). To assess directly the CRH-BP function, we created a mouse model of CRH-BP deficiency by gene targeting. Basal adrenocorticotropic hormone and corticosterone levels are unchanged in the CRH-BP-deficient mice, and the animals demonstrate a normal increase in adrenocorticotropic hormone and corticosterone after restraint stress. In contrast, adult male CRH-BP-deficient mice show significantly reduced body weight when compared with wild-type controls. CRH-BP-deficient mice also exhibit a significant increase in anxiogenic-like behavior as assessed by the elevated plus maze and defensive withdrawal tests. The increased anorectic and anxiogenic-like behavior most likely is caused by increased “free” CRH and/or urocortin levels in the brain of CRH-BP-deficient animals, suggesting an important role for CRH-BP in maintaining appropriate levels of these peptides in the central nervous system.
Intestinal trefoil factor (ITF), a small, compact protease-resistant peptide, is abundantly expressed in goblet cells of large and small intestine. Although several biological activities of ITF have been identified, including promotion of wound healing, stimulation of epithelial cell migration, and protection of intestinal epithelial barrier, little is known about signaling events through which ITF mediates its physiological function. In this study, the effects of exogenous ITF on mitogen-activated protein kinase (MAPK) signaling cascades were examined in IEC-6 cells, a nontransformed intestinal epithelial cell line that does not express endogenous trefoil peptides. Stimulation with ITF resulted in rapid decrease in extracellular signal-related protein kinase (ERK) activity and concomitant reduced ERK tyrosine phosphorylation. ITF also decreased activation of ERK activity induced by either transforming growth factor-α, which links extracellular stimuli to the Ras/Raf/MEK/ERK pathway via the epidermal growth factor receptor, or phorbol 12-myristate 13-acetate, which activates Raf through protein kinase C. ITF-induced inhibition of ERK activity was blocked by an inhibitor of tyrosine and dual-specific phosphatases, sodium orthovanadate. In summary...
The secretion and the blood levels of the adrenal steroid dehydroepiandrosterone (DHEA) and its sulfate ester (DHEAS) decrease profoundly with age, and the question is posed whether administration of the steroid to compensate for the decline counteracts defects associated with aging. The commercial availability of DHEA outside the regular pharmaceutical–medical network in the United States creates a real public health problem that may be resolved only by appropriate long-term clinical trials in elderly men and women. Two hundred and eighty healthy individuals (women and men 60–79 years old) were given DHEA, 50 mg, or placebo, orally, daily for a year in a double-blind, placebo-controlled study. No potentially harmful accumulation of DHEAS and active steroids was recorded. Besides the reestablishment of a “young” concentration of DHEAS, a small increase of testosterone and estradiol was noted, particularly in women, and may be involved in the significantly demonstrated physiological–clinical manifestations here reported. Bone turnover improved selectively in women >70 years old, as assessed by the dual-energy x-ray absorptiometry (DEXA) technique and the decrease of osteoclastic activity. A significant increase in most libido parameters was also found in these older women. Improvement of the skin status was observed...
Aspergillus flavus, like approximately one–third of ascomycete fungi, is thought to be cosmopolitan and clonal because it has uniform asexual morphology. A. flavus produces aflatoxin on nuts, grains, and cotton, and assumptions about its life history are being used to develop strategies for its biological control. We tested the assumptions of clonality and conspecificity in a sample of 31 Australian isolates by assaying restriction site polymorphisms from 11 protein encoding genes and DNA sequences from five of those genes. A. flavus isolates fell into two reproductively isolated clades (groups I and II). The lack of concordance among gene genealogies among isolates in one of the clades (group I) was consistent with a history of recombination. Our analysis included five strains of the closely related industrial fungus A. oryzae, all of which proved to be clonally related to group I.