Showing papers by "Rockefeller University published in 1996"
TL;DR: A leptin receptor was recently cloned from choroid plexus and shown to map to the same 6-cM interval on mouse chromosome 4 as db8, suggesting that the weight-reducing effects of leptin may be mediated by signal transduction through a leptin receptor in the hypothalamus.
Abstract: MUTATIONS in the mouse diabetes(db) gene result in obesity and diabetes in a syndrome resembling morbid human obesity1. Previous data suggest that the db gene encodes the receptor for the obese(ob) gene product, leptin2–7. A leptin receptor was recently cloned from choroid plexus and shown to map to the same 6-cM interval on mouse chromosome 4 as db8. This receptor maps to the same 300-kilobase interval as db, and has at least six alternatively spliced forms. One of these splice variants is expressed at a high level in the hypothalamus, and is abnormally spliced in C57BL/Ks db/db mice. The mutant protein is missing the cytoplasmic region, and is likely to be defective in signal transduction. This suggests that the weight-reducing effects of leptin may be mediated by signal transduction through a leptin receptor in the hypothalamus.
2,381 citations
TL;DR: It is shown that subjects with the MODY3-form of NIDDM have mutations in the gene encoding hepatocyte nuclear factor-1α (HNF-1 α), which is encoded by the gene TCF1, which is a transcription factor that helps in the tissue-specific regulation of the expression of several liver genes.
Abstract: The disease non-insulin-dependent (type 2) diabetes mellitus (NIDDM) is characterized by abnormally high blood glucose resulting from a relative deficiency of insulin. It affects about 2% of the world's population and treatment of diabetes and its complications are an increasing health-care burden. Genetic factors are important in the aetiology of NIDDM, and linkage studies are starting to localize some of the genes that influence the development of this disorder. Maturity-onset diabetes of the young (MODY), a single-gene disorder responsible for 2-5% of NIDDM, is characterized by autosomal dominant inheritance and an age of onset of 25 years or younger. MODY genes have been localized to chromosomes 7, 12 and 20 (refs 5, 7, 8) and clinical studies indicate that mutations in these genes are associated with abnormal patterns of glucose-stimulated insulin secretion. The gene on chromosome 7 (MODY2) encodes the glycolytic enzyme glucokinases which plays a key role in generating the metabolic signal for insulin secretion and in integrating hepatic glucose uptake. Here we show that subjects with the MODY3-form of NIDDM have mutations in the gene encoding hepatocyte nuclear factor-1alpha (HNF-1alpha, which is encoded by the gene TCF1). HNF-1alpha is a transcription factor that helps in the tissue-specific regulation of the expression of several liver genes and also functions as a weak transactivator of the rat insulin-I gene.
1,584 citations
TL;DR: These studies provide compelling support for the view that one mechanism by which these mutant PS1 cause AD is by increasing the extracellular concentration of Abeta peptides terminating at 42(43), species that foster Abeta deposition.
1,552 citations
TL;DR: Electron microscopic analysis of serial sections showed that during chain migration, neural precursors moved associated with each other and were not guided by radial glial or axonal fibers.
Abstract: In the brain of adult mice, cells that divide in the subventricular zone of the lateral ventricle migrate up to 5 millimeters to the olfactory bulb where they differentiate into neurons. These migrating cells were found to move as chains through a well-defined pathway, the rostral migratory stream. Electron microscopic analysis of serial sections showed that these chains contained only closely apposed, elongated neuroblasts connected by membrane specializations. A second cell type, which contained glial fibrillary acidic protein, ensheathed the chains of migrating neuroblasts. Thus, during chain migration, neural precursors moved associated with each other and were not guided by radial glial or axonal fibers.
1,312 citations
TL;DR: X-ray crystallographic structures of the third PDZ domain from the synaptic protein PSD-95 in complex with and in the absence of its peptide ligand have been determined and reveal that specific side chain interactions and a prominent hydrophobic pocket explain the selective recognition of the C-terminal consensus sequence.
1,178 citations
TL;DR: The cloning of ob, and the demonstration that it encodes a secreted protein that binds specifically to a receptor (OB-R) in the brain, have validated critical aspects of this hypothesis.
Abstract: Mice harboring mutations in the obese (ob) and diabetes (db) genes display similar phenotypes, and it has been proposed that these genes encode the ligand and receptor, respectively, for a physiologic pathway that regulates body weight. The cloning of ob, and the demonstration that it encodes a secreted protein (leptin) that binds specifically to a receptor (OB-R) in the brain, have validated critical aspects of this hypothesis. Here it is shown by genetic mapping and genomic analysis that mouse db, rat fatty (a homolog of db), and the gene encoding the OB-R are the same gene.
1,156 citations
TL;DR: Transcription initiation on protein-encoding genes represents a major control point for gene expression in eukaryotes, and is mediated by RNA polymerase II and a surprisingly complex array of general initiation factors that are highly conserved from yeast to man.
1,125 citations
TL;DR: CD4 binding, although not absolutely necessary for the gp120–CCR-5 interaction, greatly increases its efficiency, and interference with HIV-1 binding to one or both of its receptors (CD4 and CCR-5) may be an important mechanism of virus neutralization.
Abstract: The beta-chemokine receptor CCR-5 is an essential co-factor for fusion of HIV-1 strains of the non-syncytium-inducing (NSI) phenotype with CD4+ T-cells. The primary binding site for human immunodeficiency virus (HIV)-1 is the CD4 molecule, and the interaction is mediated by the viral surface glycoprotein gp120 (refs 6, 7). The mechanism of CCR-5 function during HIV-1 entry has not been defined, but we have shown previously that its beta-chemokine ligands prevent HIV-1 from fusing with the cell. We therefore investigated whether CCR-5 acts as a second binding site for HIV-1 simultaneously with or subsequent to the interaction between gp120 and CD4. We used a competition assay based on gp120 inhibition of the binding of the CCR-5 ligand, macrophage inflammatory protein (MIP)-1beta, to its receptor on activated CD4+ T cells or CCR-5-positive CD4- cells. We conclude that CD4 binding, although not absolutely necessary for the gp120-CCR-5 interaction, greatly increases its efficiency. Neutralizing monoclonal antibodies against several sites on gp120, including the V3 loop and CD4-induced epitopes, inhibited the interaction of gp120 with CCR-5, without affecting gp120-CD4 binding. Interference with HIV-1 binding to one or both of its receptors (CD4 and CCR-5) may be an important mechanism of virus neutralization.
1,113 citations
TL;DR: The data indicate the hypothalamus is a direct target of leptin action and that this activation is critically dependent on the gp-130-like leptin receptor isoform missing in C57BLKS/J db/db mice, the first in vivo demonstration of leptin signal transduction.
Abstract: Leptin, a hormone secreted by adipocytes, regulates the size of the adipose tissue mass through effects on satiety and energy metabolism. Leptin's precise sites of action are not known. The leptin receptor (Ob-R) is found in many tissues in several alternatively spliced forms raising the possibility that leptin exerts effects on many tissues including the hypothalamus. Ob-R is a member of the gp130 family of cytokine receptors which are known to stimulate gene transcription via activation of cytosolic STAT proteins. In order to identify the sites of leptin action in vivo, we assayed for activation of STAT proteins in mice treated with leptin. The STAT proteins bind to phosphotyrosine residues in the cytoplasmic domain of the ligand-activated receptor where they are phosphorylated. The activated STAT proteins dimerize and translocate to the nucleus where they bind DNA and activate transcription. The activation of STAT proteins in response to leptin was assayed in a variety of mouse tissues known to express Ob-R. Leptin injection activated Stat3 but no other STAT protein in the hypothalamus of ob/ob and wild-type mice but not db/db mice, mutants that lack an isoform of the leptin receptor. Leptin did not induce STAT activation in any of the other tissues tested. Activation of Stat3 by leptin was dose dependent and first observed after 15 minutes and maximal at 30 minutes. Our data indicate the hypothalamus is a direct target of leptin action and that this activation is critically dependent on the gp-130-like leptin receptor isoform missing in C57BLKS/J db/db mice. This is the first in vivo demonstration of leptin signal transduction.
1,074 citations
National Institutes of Health1, Stanford University2, Massachusetts Institute of Technology3, Wellcome Trust Sanger Institute4, University of Oxford5, Wellcome Trust6, University of Michigan7, Rockefeller University8, Laval University9, J. Craig Venter Institute10, Centre national de la recherche scientifique11, Anschutz Medical Campus12, University of Cambridge13, McGill University14
TL;DR: The gene map unifies the existing genetic and physical maps with the nucleotide and protein sequence databases in a fashion that should speed the discovery of genes underlying inherited human disease.
Abstract: The human genome is thought to harbor 50,000 to 100,000 genes, of which about half have been sampled to date in the form of expressed sequence tags. An international consortium was organized to develop and map gene-based sequence tagged site markers on a set of two radiation hybrid panels and a yeast artificial chromosome library. More than 16,000 human genes have been mapped relative to a framework map that contains about 1000 polymorphic genetic markers. The gene map unifies the existing genetic and physical maps with the nucleotide and protein sequence databases in a fashion that should speed the discovery of genes underlying inherited human disease. The integrated resource is available through a site on the World Wide Web at http://www.ncbi.nlm.nih.gov/SCIENCE96/.
1,072 citations
TL;DR: The apolipoprotein E (APOE) E4 allele is associated with Alzheimer's disease, cardiovascular disease, and decreased longevity, and cell lines were created which secrete each apoE isoform to probe the mechanism of these associations.
Abstract: The apolipoprotein E (APOE) E4 allele is associated with Alzheimer's disease, cardiovascular disease, and decreased longevity To probe the mechanism of these associations, cell lines were created which secrete each apoE isoform ApoE conditioned media, purified apoE, and commercially obtained apoE protected B12 cells from hydrogen peroxide cytotoxicity with E2 > E3 > E4 Physiological levels of apoE protected cells from beta-amyloid peptides, while higher doses of apoE led to increased cytotoxicity E2 > E3 > E4 possessed antioxidant activity, and apoE bound certain metal ions The decreased antioxidant activity of E4 could contribute to its association with Alzheimer's disease, cardiovascular disease and decreased longevity
TL;DR: DPC4 is essential for the function of Smadl and Smad2 in pathways that signal mesoderm induction and patterning in Xenopus embryos, as well as antimitogenic and transcriptional responses in breast epithelial cells.
Abstract: The TGF-beta/activin/BMP superfamily of growth factors signals through heteromeric receptor complexes of type I and type II serine/threonine kinase receptors. The signal originated by TGF-beta-like molecules appears to be transduced by a set of evolutionarily conserved proteins known as SMADs, which upon activation directly translocate to the nucleus where they may activate transcription. Five SMAD proteins have so far been characterized in vertebrates. These factors are related to the mediator of decapentaplegic (dpp) signalling, mothers against dpp (Mad), in Drosophila and to the Sma genes from Caenorhabditis elegans. Smad1 and Smad2 have been shown to mimic the effects of BMP and activin, respectively, both in Xenopus and in mammalian cells, whereas Smad3 (a close homologue of Smad2) and the related protein DPC4, a tumour-suppressor gene product, mediate TGF-beta actions. We report here that DPC4 is essential for the function of Smad1 and Smad2 in pathways that signal mesoderm induction and patterning in Xenopus embryos, as well as antimitogenic and transcriptional responses in breast epithelial cells. DPC4 associates with Smad1 in response to BMP and with Smad2 in response to activin or TGF-beta. DPC4 is therefore a regulated partner of SMADs that function in different signalling pathways of the TGF-beta family.
TL;DR: The results suggest that the naturalistic stressorpsychosocial stress induces specific structural changes in hippocampal neurons of subordinate male tree shrews, probably are mediated by activation of the hypothalamo–pituitary–adrenal-axis acting in concert with endogenous EAAs from mossy fiber input.
Abstract: We have shown previously that repeated laboratory restraint stress or daily corticosterone administration affects the structure of CA3 hippocampal neurons in rats. In the present study, we investigated the effect of repeated daily psychosocial stress on the structure of hippocampal CA3 pyramidal neurons in male tree shrews (Tupaia belangeri). Male tree shrews develop social hierarchies in which subordinates show characteristic changes in physiological and behavioral parameters when confronted with a dominant. In the present experiments, subordinate animals lost body weight soon after starting the daily social conflict, and urinary excretion of cortisol was elevated throughout the experiment as compared with the control period. Golgi-impregnated brain tissue from subordinates exposed to 28 d (1 hr/d) of social confrontations was compared with that fromcontrol nonstressed animals. The apical dendrites of the CA3 pyramidal cells from subordinates had a decreased number of branch points and total dendritic length as compared with controls. No differences were observed in apical dendritic spine density or in the basal dendritic tree morphology. The stress-induced CA3 apical dendritic atrophy in subordinates was prevented by administering daily oral doses of the antiepileptic drug phenytoin (Dilantin, Sigma, St. Louis, MO) (200 mg/kg), which interferes with excitatory amino acid (EAA) action. These results suggest that the naturalistic stressorpsychosocial stress induces specific structural changes in hippocampal neurons of subordinate male tree shrews. These changes, like those in the rat after glucocorticoid treatment or restraint stress, probably are mediated by activation of the hypothalamo–pituitary–adrenal-axis acting in concert with endogenous EAAs from mossy fiber input.
TL;DR: While plasma [leptin], corrected for FM, declines significantly in women post-menopause, this decline is not sufficient to account for the striking sexual dimorphism in the relationship of leptin to fat mass.
Abstract: Circulating concentrations of leptin ([leptin]) vary directly with body mass index and percentage body fat, and may thus constitute an afferent limb of a system regulating body fatness. We tested the hypotheses that: 1) Plasma [leptin] vary more directly with absolute fat mass than with fractional body fatness per se: and 2). The relationship between fat mass and [leptin] is significantly affected by gender and by menopausal status. [Leptin] in the post-absorptive state was examined in 67 subjects (26 male, 20 premenopausal female, 21 postmenopausal females; 43 never-obese, 24 obese) at usual body weight. Body composition was determined by hydrodensitometry, and [leptin] was determined by a double antibody ELISA assay. In male and pre-menopausal female subjects, subcutaneous adipose tissue aspirations were performed for determination of adipocyte volume by the osmium fixation method, and a 3 hour oral glucose tolerance tests was performed. At usual body weight, ([leptin]) was better correlated with absolu...
TL;DR: The crystal structure of the human DNA polymerase delta processivity factor PCNA (proliferating cell nuclear antigen) complexed with a 22 residue peptide derived from the C-terminus of the cell-cycle checkpoint protein p21(WAF1/CIP1) has been determined at 2.6 angstrom resolution.
TL;DR: It is shown that inhibitory signalling by FcγRIIB does not require the SH2-domain-containing protein tyrosine phosphatase, SHP-1, in mast cells and results in the recruitment of the SH1- domain-containing inositol polyphosphate 5-phosphatase to the tyrosinesine- phosphorylated 13-amino-acid inhibitory motif of FcβB in both B cells and mast cells.
Abstract: Immune complexes are potent activators of inflammatory cells, triggering effector responses through the crosslinking of Fc receptors (FcRs) such as Fc(epsilon)RI or Fc(gamma)RIII. On B cells and mast cells, immune complexes are also negative regulators of activation triggered by antigen and Fc receptors, a consequence of coligation of the B-cell antigen receptor or Fc(epsilon)RI, respectively, and the inhibitory receptor Fc(gamma)RIIB. Here we show that inhibitory signalling by Fc(gamma)RIIB does not require the SH2-domain-containing protein tyrosine phosphatase, SHP-1, in mast cells and results in the recruitment of the SH2-domain-containing inositol polyphosphate 5-phosphatase, SHIP, to the tyrosine-phosphorylated 13-amino-acid inhibitory motif of Fc(gamma)RIIB in both B cells and mast cells. SHIP, by hydrolysing the 5-phosphate of phosphatidylinositol(3,4,5)P3 and inositol(1,3,4,5)P4, suggests a mechanism by which Fc(gamma)RIIB can inhibit calcium influx and downstream responses triggered by immune receptors.
TL;DR: Results from Experiment 4 showed that chronic restraint stress impaired spatial memory performance on the Y maze when rats were tested the day after the last stress session and that tianeptine prevented the stress-induced spatial memory impairment.
Abstract: Chronic restraint stress causes significant dendritic atrophy of CA3 pyramidal neurons that reverts to baseline within a week. Therefore, the authors assessed the functional consequences of this atrophy quickly (within hours) using the Y maze. Experiments 1-3 demonstrated that rats relied on extrinsic, spatial cues located outside of the Y maze to determine arm location and that rats with hippocampal damage (through kainic acid, colchicine, or trimethyltin) had spatial memory impairments. After the Y maze was validated as a hippocampally relevant spatial task, Experiment 4 showed that chronic restraint stress impaired spatial memory performance on the Y maze when rats were tested the day after the last stress session and that tianeptine prevented the stress-induced spatial memory impairment. These data are consistent with the previously demonstrated ability of tianeptine to prevent chronic stress-induced atrophy of the CA3 dendrites.
TL;DR: It is demonstrated that cells originating at different rostrocaudal levels of the SVZ migrate rostrally and reach the olfactory bulb where they differentiate into neurons.
Abstract: Cells in the brains of adult mammals continue to proliferate in the subventricular zone (SVZ) throughout the lateral wall of the lateral ventricle. Here we show, using whole mount dissections of this wall from adult mice, that the SVZ is organized as an extensive network of chains of neuronal precursors. These chains are immunopositive to the polysialylated form of NCAM, a molecule present at sites of plasticity, and TuJ1, an early neuronal marker. The majority of the chains are oriented along the rostrocaudal axis and many join the rostral migratory stream that terminates in the olfactory bulb. Using focal microinjections of DII and transplantation of SVZ cells carrying a neuron-specific reporter gene, we demonstrate that cells originating at different rostrocaudal levels of the SVZ migrate rostrally and reach the olfactory bulb where they differentiate into neurons. Our results reveal an extensive network of pathways for the tangential chain migration of neuronal precursors throughout the lateral wall of the lateral ventricle in the adult mammalian brain.
TL;DR: Improved method for generating sizable numbers of mature dendritic cells from nonproliferating progenitors in human blood is investigated, suggesting that these cells will be effective in vivo as adjuvants for active immunotherapy.
TL;DR: This work has shown that TFIID, a 700-kD complex composed of the TATA box binding protein (TBP) and a set of phylogenetically conserved, polymerase-specific TBP-associated factors or TAFIIS, play critical regulatory roles in eukaryotic gene expression.
Abstract: Eukaryotes have three distinct RNA polymerases that catalyze transcription of nuclear genes. RNA polymerase II is responsible for transcribing nuclear genes encoding the messenger RNAs and several small nuclear RNAs. Like RNA polymerases I and III, pol II cannot recognize its target promoter directly and initiate transcription without accessory factors. Instead, this large multisubunit enzyme relies on both general transcription factors and transcriptional activators and coactivators to regulate transcription from class II promoters. At the center of this process is TFIID, a 700-kD complex composed of the TATA box binding protein (TBP) and a set of phylogenetically conserved, polymerase-specific TBP-associated factors or TAFns. Together, TBP and the TAFIIs direct assembly of the transcription machinery and play critical regulatory roles in eukaryotic gene expression.
TL;DR: Low-dimensional representations of sensory signals are key to solving many of the computational problems encountered in high-level vision.
Abstract: Low-dimensional representations of sensory signals are key to solving many of the computational problems encountered in high-level vision. Principal component analysis (PCA) has been used in the pa...
TL;DR: DC exhibited features that had not been seen with other antigen‐presenting cells: they produced bioactive IL‐12 upon antigen‐specific interaction with T cells without any other stimuli; in an allogeneic mixed leukocyte reaction model, neutralizing anti‐IL‐12 antibodies showed that DC‐derived IL‐ 12 was critical for optimal proliferation and IFN‐γ production by activated Th1 blasts.
Abstract: Interleukin-12 (IL-12), a 70-kDa heterodimeric cytokine composed of covalently linked p35 and p40 chains, is to date the most critical factor for skewing the immune response towards a T helper 1 (Th1) of cytokine profile [high interferon-gamma (IFN-gamma), low IL-4]. Established sources of IL-12 are stimulated macrophages, neutrophils and B cells. As dendritic cells (DC) process antigen in the periphery and then migrate to lymphoid organs to sensitize T cells and induce cell mediated immunity, we reasoned that DC should constitute a critical source of IL-12. The criteria used to detect IL-12 in DC were the demonstration of p40 and p35 mRNA (semiquantitative polymerase chain reaction, northern blotting, and in situ hybridization) as well as IL-12 protein (p70 enzyme-linked immunosorbent assay, p70 antigen capture followed by IFN-gamma bioassay, free p40 chain radioimmunoassay or immunoprecipitation). We found that conventional stimuli such as Staphylococcus aureus induced production of IL-12 by murine as well as human DC in amounts comparable to spleen cells, peritoneal macrophages or peripheral mononuclear cells. DC exhibited, however, features that had not been seen with other antigen-presenting cells: they produced bioactive IL-12 upon antigen-specific interaction with T cells without any other stimuli; in an allogeneic mixed leukocyte reaction model, neutralizing anti-IL-12 antibodies showed that DC-derived IL-12 was critical for optimal proliferation and IFN-gamma production by activated Th1 blasts; and finally, the priming of resting, naive allogeneic T cells by DC, followed by restimulation of primed T blasts by DC, skewed the response to Th1 without the need for any exogenous cytokines or stimuli such as microorganisms. This skewing to Th1 cytokine production, which depended on DC-derived IL-12, but did not require anti-IL-4, exogenous IL-12, or microbes, might be a major function of DC.
TL;DR: Induced mutations in mice that are deficient in apolipoprotein E, a ligand important in lipoprotein clearance, develop atherosclerotic lesions resembling those observed in humans, and these models are now being used to study the pathogenesis of atherosclerosis.
Abstract: As a species the mouse is highly resistant to atherosclerosis. However, through induced mutations it has been possible to develop lines of mice that are susceptible to this disease. For example, mice that are deficient in apolipoprotein E, a ligand important in lipoprotein clearance, develop atherosclerotic lesions resembling those observed in humans. These lesions are exacerbated when the mice are fed a high-cholesterol, high-fat, Western-type diet. Other promising models are mice that are deficient in the low density lipoprotein receptor and transgenic mice that express human apolipoprotein B and transdominant mutant forms of apolipoprotein E. These models are now being used to study the pathogenesis of atherosclerotic lesions, as well as the influence of genetics, environment, hormones, and drugs on lesion development.
TL;DR: The unexpected growth phenotype suggests a new and important link between Ku80 and growth control and the role of the DNA-binding subunit of DNA-PK in vivo is targeted.
Abstract: The DNA-dependent protein kinase (DNA-PK) is a mammalian serine/threonine kinase that is implicated in the repair of DNA double-strand breaks, DNA replication, transcription, and V(D)J recombination. To determine the role of the DNA-binding subunit of DNA-PK in vivo, we targeted Ku80 in mice. In mutant mice, T and B lymphocyte development is arrested at early progenitor stages and there is a profound deficiency in V(D)J rearrangement. Although Ku80-/- mice are viable and reproduce, they are 40-60% of the size of littermate controls. Consistent with this growth defect, fibroblasts derived from Ku80-/- embryos showed an early loss of proliferating cells, a prolonged doubling time, and intact cell-cycle checkpoints that prevented cells with damaged DNA from entering the cell-cycle. The unexpected growth phenotype suggests a new and important link between Ku80 and growth control.
TL;DR: It is concluded that NSAIDs inhibit the proliferation rate of the two colon cancer cell lines independent of their ability to inhibit PG synthesis, meaning that alternative mechanisms for their activity on tumor cell growth must be entertained.
TL;DR: These findings demonstrate the ligand-dependent in vivo formation of a transcriptionally active TR-multisubunit protein complex and suggest a role for TRAPs as positive coactivators for gene-specific transcriptional activation.
Abstract: Transcriptional regulation by nuclear hormone receptors is thought to involve interactions with putative cofactors that may potentiate receptor function. Here we show that human thyroid hormone receptor alpha purified from HeLa cells grown in the presence of thyroid hormone (T3) is associated with a group of distinct nuclear proteins termed thyroid hormone receptor-associated proteins (TRAPs). In an in vitro system reconstituted with general initiation factors and cofactors (and in the absence of added T3), the "liganded" thyroid hormone receptor (TR)/TRAP complex markedly activates transcription from a promoter template containing T3-response elements. Moreover, whereas the retinoid X receptor is not detected in the TR/TRAP complex, its presence is required for the function of the complex. In contrast, human thyroid hormone receptor alpha purified from cells grown in the absence of T3 lacks the TRAPs and effects only a low level of activation that is dependent on added ligand. These findings demonstrate the ligand-dependent in vivo formation of a transcriptionally active TR-multisubunit protein complex and suggest a role for TRAPs as positive coactivators for gene-specific transcriptional activation.
TL;DR: Results indicate that there is a reverse-phase chromatographic-like dimension in the sample-matrix preparation procedures that can be exploited to optimize the analysis of MALDI mass spectra.
Abstract: Sample−matrix preparation procedures are shown to greatly influence the quality of the matrix-assisted laser desorption/ionization (MALDI) mass spectra of peptides and proteins. In particular, dramatic mass discrimination effects are observed when the matrix 4-hydroxy-α-cyanocinnamic acid is used for analyzing complex mixtures of peptides and proteins. The discrimination effects are found to be strongly dependent on the sample−matrix solution composition, pH, and the rates at which the sample−matrix cocrystals are grown. These findings demonstrate the need to exercise great care in performing and interpreting the MALDI analysis of biological samples. The results also indicate that there is a reverse-phase chromatographic-like dimension in the sample−matrix preparation procedures that can be exploited to optimize the analysis. The present work describes the conditions under which the majority of components of a complex mixture of peptides and proteins can be successfully measured.
TL;DR: Injections of testosterone into the yolk of unincubated eggs enhanced the growth after hatching compared to nestlings that had hatched simultaneously from control eggs, but more testosterone did not compensate for reduced growth that was caused by later hatching due to asynchronous incubation of clutches.
TL;DR: It is found that wild-type primary mouse embryonic fibroblasts were inhibited by IFNs, but fibro Blasts from Stat1-deficient mouse embryos were not inhibited byIFNs.
Abstract: Type I (alpha, beta) and type II (gamma) interferons (IFNs) can restrict the growth of many cell types. INF-stimulated gene transcription, a key early event in IFN response, acts through the Janus kinase-signal transducers and activators of transcription pathway, in which both IFN-alpha and IFN-gamma activate the transcription factor Stat1. A cell line lacking Stat1 (U3A) was not growth-arrested by IFN-alpha or IFN-gamma, and experiments were carried out with U3A cells permanently expressing normal or various mutant forms of Stat1 protein. Only cells in which complete Stat1 activity was available (Stat1alpha) were growth-inhibited by IFN-gamma. A mutant that supports 20-30% normal transcription did not cause growth restraint. In contrast, IFN-alpha growth restraint was imposed by cells producing Stat1beta, which lacks transcriptional activation potential. This parallels earlier results showing the truncated Stat1 can function in IFN-alpha gene activation. In addition to experiments on long-term cultured cells, we also found that wild-type primary mouse embryonic fibroblasts were inhibited by IFNs, but fibroblasts from Stat1-deficient mouse embryos were not inhibited by IFNs.
TL;DR: These data represent the most complete survey to date of auditory pathways in the adult male zebra finch brain, and of their projections to motor stations of the song system.
Abstract: Auditory information is critical for vocal imitation and other elements of social life in song birds. In zebra finches, neural centers that are necessary for the acquisition and production of learned vocalizations are known, and they all respond to acoustic stimulation. However, the circuits by which conspecific auditory signals are perceived, processed, and stored in long-term memory have not been well documented. In particular, no evidence exists of direct connections between auditory and vocal motor pathways, and two newly identified centers for auditory processing, caudomedial neostriatum (Ncm) and caudomedial hyperstriatum ventrale (cmHV), have no documented place among known auditory circuits. Our goal was to describe anatomically the auditory pathways in adult zebra finch males and, specifically, to show the projections by which Ncm and vocal motor centers may receive auditory input. By using injections of different kinds of neuroanatomical tracers (biotinylated dextran amines, rhodamine-linked dextran amines, biocytin, fluorogold, and rhodamine-linked latex beads), we have shown that, as in other avian groups, the neostriatal field L complex in caudal telencephalon is the primary forebrain relay for pathways originating in the auditory thalamus, i.e., the nucleus ovoidalis complex (Ov). In addition, Ncm and cmHV also receive input from the Ov complex. Ov has been broken down into two parts, the Ov "core" and "shell," which project in parallel to different targets in the caudal telencephalon. Parts of the field L complex are connected among themselves and to Ncm, cmHV, and caudolateral Hv (clHV) through a complex web of largely reciprocal pathways. In addition, clHV and parts of the field L complex project strongly to the "shelf" of neostriatum underneath the song control nucleus high vocal center (HVC) and to the "cup" of archistriatum rostrodorsal to another song-control nucleus, the robust nucleus of the archistriatum (RA). We have documented two points at which the vocal motor pathway may pick up auditory signals: the HVC-shelf interface and a projection from clHV to the nucleus interfacialis (NIf), which projects to HVC. These data represent the most complete survey to date of auditory pathways in the adult male zebra finch brain, and of their projections to motor stations of the song system.