The size, shape, density and ring of the dwarf planet Haumea from a stellar occultation.

Haumea-one of the four known trans-Neptunian dwarf planets-is a very elongated and rapidly rotating body. In contrast to other dwarf planets, its size, shape, albedo and density are not well constrained. The Centaur Chariklo was the first body other than a giant planet known to have a ring system, and the Centaur Chiron was later found to possess something similar to Chariklo's rings. Here we report observations from multiple Earth-based observatories of Haumea passing in front of a distant star (a multi-chord stellar occultation). Secondary events observed around the main body of Haumea are consistent with the presence of a ring with an opacity of 0.5, width of 70 kilometres and radius of about 2,287 kilometres. The ring is coplanar with both Haumea's equator and the orbit of its satellite Hi'iaka. The radius of the ring places it close to the 3:1 mean-motion resonance with Haumea's spin period-that is, Haumea rotates three times on its axis in the time that a ring particle completes one revolution. The occultation by the main body provides an instantaneous elliptical projected shape with axes of about 1,704 kilometres and 1,138 kilometres. Combined with rotational light curves, the occultation constrains the three-dimensional orientation of Haumea and its triaxial shape, which is inconsistent with a homogeneous body in hydrostatic equilibrium. Haumea's largest axis is at least 2,322 kilometres, larger than previously thought, implying an upper limit for its density of 1,885 kilograms per cubic metre and a geometric albedo of 0.51, both smaller than previous estimates. In addition, this estimate of the density of Haumea is closer to that of Pluto than are previous estimates, in line with expectations. No global nitrogen- or methane-dominated atmosphere was detected.

Macrophage-stimulating protein polymorphism rs3197999 is associated with a gain of function: implications for inflammatory bowel disease.

Crohn's disease and ulcerative colitis, the two main types of inflammatory bowel disease (IBD), were reported to be associated with a variety of genetic polymorphisms. A subset of these polymorphisms was identified in both diseases and only three of them were found in primary sclerosing cholangitis (PSC). rs3197999 (Arg689Cys) located in the MST1 gene is one of the most convincingly replicated IBD/PSC-associated polymorphisms but its functional consequences have not been investigated, yet. We expressed both MST1 gene variants (Arg(689) (MSP(wt)) and Cys(689) (MSP(mut)) in a eukaryotic cell system and compared their stimulatory effects on macrophage-like THP-1 cells. Except for the rate of apoptosis that remained unchanged, MSP(mut) significantly increased the stimulatory effect of MSP (macrophage-stimulating protein) on chemotaxis and proliferation by THP-1 cells, indicating a gain of function associated with the Arg689Cys exchange. A broad set of evidence reported previously suggests that pro-inflammatory changes in macrophage function have a major role in the initiation of the inflammatory process in IBD and PSC. Therefore, the gain of function observed with rs3197999 in MST1 might provide a cellular mechanism for the consistent association of this polymorphism with an increased risk for IBD and PSC.

Independent glucocorticoid induction and repression of two contiguous responsive genes.

Specific DNA sequence elements which contain binding sites for the glucocorticoid receptor mediate the action of glucocorticoid hormones on gene transcription. In glucocorticoid-inducible genes, these glucocorticoid-responsive elements behave as hormone-inducible enhancers of transcription. We have taken advantage of the bovine papillomavirus (BPV) system to test the stringency of glucocorticoid regulation of transcription. BPV episomes were constructed to contain two hormone-regulated transcription units in close proximity; one transcription unit is under control of a glucocorticoid-inducible promoter (mouse mammary tumor virus) while the other is under control of a glucocorticoid-inhibited promoter (pro-opiomelanocortin). Glucocorticoids independently regulated transcription of the two physically linked transcription units, irrespective of their relative orientation and of their proximity on the BPV episomes. This result contrasts with the so-called position-independent activity of enhancers and suggests that the multicomponent organization of eucaryotic promoters restricts the action of hormone-responsive regulatory elements to a specific transcription unit, thus accounting for the stringency of hormonal regulation observed in vivo.

Two regions of the mouse mammary tumor virus long terminal repeat regulate the activity of its promoter in mammary cell lines.

In vivo expression of the mouse mammary tumor virus (MMTV) is restricted to a few organs, with the highest rate of transcription found in the mammary gland. Using a series of mammary and nonmammary murine cell lines, we have identified two regulatory elements, located upstream of the hormone responsive element, that specifically regulate the MMTV promoter. The first element displays an enhancerlike activity and is coincident with the binding of a nuclear factor (designated MP4; position -1078 to -1052 in the long terminal repeat) whose presence is apparently restricted to mammary cell lines. The second regulatory region mediates a repressive activity and is mapped to the long terminal repeat segment from -415 to -483. This repression is specific for a particular subtype of mammary cells (RAC cells) able to grow under two differentiation states (A. Sonnenberg, H. Daams, J. Calafat, and J. Hilgers, Cancer Res. 46:5913-5922, 1986). The MMTV promoter in mammary cell lines thus appears to be modulated by two cis-acting elements that are likely to be involved in tissue-specific expression in vivo.

Mouse mammary tumor virus chromatin in human breast cancer cells is constitutively hypersensitive and exhibits steroid hormone-independent loading of transcription factors in vivo.

We have stably introduced a reporter gene under the control of the mouse mammary tumor virus (MMTV) long terminal repeat (LTR) into human T47D breast cancer cells to study the action of the progesterone receptor (PR) on transcription from a chromatin template. Unexpectedly, the chromatin organization of the MMTV LTR in these human breast cancer cells differed markedly from what we have observed previously. The region adjacent to the transcription start site (-221 to -75) was found to be constitutively hypersensitive to restriction enzyme cleavage in the absence of hormone. This region is normally encompassed within the second nucleosome of a phased array of six nucleosomes that is assembled when the MMTV LTR is stably maintained in mouse cells. Characteristically, in these rodent cells, the identical DNA sequences show increased restriction enzyme cleavage only in the presence of glucocorticoid. The increased access of restriction enzymes observed in the human PR+ cells was not observed in adjacent nucleosomes and was unaffected by treatment with the progesterone antagonist RU486. In addition, exonuclease III-dependent stops corresponding to the binding sites for nuclear factor 1 and the PR were observed before and after hormone treatment. These results indicate that MMTV chromatin replicated in these cells is organized into a constitutively open architecture and that this open chromatin state is accompanied by hormone-independent loading of a transcription factor complex that is normally excluded from uninduced chromatin.

Glucocorticoid regulation of transcription at an amplified, episomal promoter.

The mouse mammary tumor virus long terminal repeat (MMTV LTR) has been introduced into cultured murine cells, using the 69% transforming fragment of bovine papilloma virus type 1 (BPV). Transformed cells contain up to 200 copies of the chimeric molecules per diploid genome. The restriction endonuclease map of the acquired recombinants, as well as the physical structure of the DNA, indicates that the LTR-BPV molecules present in these cells occur exclusively as unintegrated, extrachromosomal episome. When a 72-base pair direct repeat "enhancer" element (derived from the Harvey sarcoma retrovirus) was included in the MMTV LTR-BPV chimeric plasmids, DNA acquired through transfection, with a single exception, was integrated or rearranged or both. The transcriptional potential of the episomal MMTV promoter present in these cells was tested in two ways. First, steady-state levels of MMTV-initiated RNA were measured by quantitative S1 mapping. Second, the relative number of transcription complexes initiated in vivo was determined by using a subnuclear fraction highly enriched for MMTV-BPV minichromosomes in an in vitro transcription extension assay. Both approaches showed that the MMTV LTR present in the episomal state was capable of supporting glucocorticoid hormone-regulated transcription. We have therefore demonstrated the hormone response for the first time in a totally defined primary sequence environment. Significant differences both in the basal level of MMTV-initiated transcription and in the extent of glucocorticoid induction were observed in individual cell lines with similar episomal copy numbers. These phenotypic variations suggest that epigenetic structure is an important component of the mechanism of regulation.

Proopiomelanocortin gene promoter elements required for constitutive and glucocorticoid-repressed transcription.

The POMC gene is expressed predominantly in the anterior pituitary. The high level of POMC transcription in this tissue is modulated by peptide hormones and repressed by glucocorticoids. In this present study we have investigated promoter elements required for the high basal transcription and glucocorticoid repression using transient transfection and in vitro transcription assays. We first determined that the region between -77 to -51 of the promoter, which has previously been shown to harbor a glucocorticoid receptor-binding site, is required for high basal expression both in vivo and in vitro. This promoter domain is also required for glucocorticoid repression of transcription in vivo. Two site-directed mutants within this area both decreased basal transcription, but were fully repressed by glucocorticoids, implying that the -77 to -51 region is a complex regulatory region harboring separable basal and glucocorticoid-repressible elements. Electrophoretic mobility shift and exonuclease III footprinting analysis revealed the existence of two factors that bind in this region. We also examined the effect of broad promoter deletions on basal expression and glucocorticoid repression. These experiments revealed that the region between -480 and -320 is also required for glucocorticoid repression. Taken together, the data suggest a model in which high basal transcription is generated by direct interaction of factors binding between -480 to -320 and -77 to -51. Glucocorticoid repression could occur by direct receptor disruption of these interactions.

Differential steroid hormone induction of transcription from the mouse mammary tumor virus promoter.

The Mouse Mammary Tumor Virus (MMTV) contains sequences in its proximal promoter region to which both glucocorticoid and progesterone receptors can bind. In transient transfection experiments both hormones are able to stimulate transcription from reporter plasmids containing either native or consensus hormone response elements (glucocorticoid response element/progesterone response element). Previous experiments have demonstrated that the MMTV long terminal repeat is reproducibly assembled into a phased array of nucleosomes when stably introduced into cells. Stimulation by glucocorticoids of endogenous templates led to a rapid but transient increase in transcription initiation and mRNA accumulation that can be correlated with increased sensitivity to restriction enzymes. In contrast, experiments using progesterone or a truncated glucocorticoid receptor failed to elicit a similar increase in mRNA levels as dexamethasone from stable chromatin templates. In an attempt to understand this differential response, we have compared the responsiveness of the MMTV promoter to glucocorticoids and progesterone when it is organized in either stable chromatin or in transiently acquired plasmids. Our results demonstrate that the native chromatin structure prevents activation of this locus by progesterone, but permits stimulation by glucocorticoids.

Circulatory and metabolic effects of alpha-adrenergic blockade in the hyperinsulinemic ovine fetus.

OBJECTIVES: Fetuses of diabetic women exhibit hypoxemia, elevated catecholamine concentrations at birth, and increased incidence of death. Our previous findings suggested that experimental fetal hyperinsulinemia results in a surge in catecholamines with cardiovascular changes supported by increased beta-adrenergic activity. The present experiments were designed to assess the contribution of alpha-adrenergic stimulation to the hemodynamic changes in the hyperinsulinemic ovine fetus. METHODS: Combined ventricular output, regional organ blood flow, vascular resistance, metabolism, and catecholamine concentrations were measured before and during an infusion of insulin and during continued infusion with alpha-adrenergic blockade (phentolamine) in eight chronically catheterized fetal sheep. RESULTS: Fetal insulin infusion produced hyperinsulinemic-hypoglycemia, a surge in epinephrine and norepinephrine concentration, and increases in the combined ventricular output (blood flow to the fetus plus placenta) and regional blood flow to the fetus, heart, stomach, gastrointestinal tract, fat, and carcass. In the hyperinsulinemic state, alpha-adrenergic blockade was associated with additional increases in fetal norepinephrine concentration and no major changes in combined ventricular output or blood flow to the body of the fetus, except for decreased blood flow to the stomach and lungs, and a decrease in stroke volume. CONCLUSIONS: Because vasodilation characterizes the hyperinsulinemic state, alpha-adrenergic stimulation contributes less to compensatory cardiovascular changes in the hyperinsulinemic fetus than that which we previously have shown for beta-adrenergic stimulation.

Towards a density functional treatment of chemical reactions in complex media.

We discuss two techniques involving density functional theory (i.e., ab initio molecular dynamics simulations and frozen density functional theory) which show promise for applications directed towards understanding reactions in complex media. Preliminary results for the simulation of the conformational dynamics of an isolated analogue of alanine dipeptide and for the interaction between F- and H2O are included. These results represent the first steps towards a combined theoretical approach of reactions in complex media.

Electrical stimulation of the superior olivary complex can produce cortical evoked potential and behavioral discrimination correlates of pitch perception in the rat.

Patterned electrical stimulation of the superior olivary complex (SOC) which simulated the neural frequency following response (FFR) extracellular potential was used as a stimulus in behavioral frequency discrimination and cortical evoked potential studies. Behavioral judgments of SOC stimulation frequency were found to be as accurate as those obtained for 80 dB acoustic stimuli within the spectral band of the FFR (200 to 3800 Hz). Cortical evoked potentials elicited by acoustic and electrical stimulation of the SOC were then compared for preservation of waveform similarity. Frequency dependent similarity was observed in slow wave events elicited by stimuli with frequencies in the FFR band. A 3 msec time lag was found between acoustic and SOC stimulation produced waveforms which can be accounted for by forward stimulation of the auditory pathway. Our study supports the idea that integrated extracellular waveforms of the FFR index low frequency representations in the auditory brainstem, perhaps by selecting patches of SOC cell transmembrane potential changes. Because bilateral cochlear damage did not prevent behavioral discrimination of SOC electrical stimulation, feedback to the ear is not necessary for perceptual significance of simulated FFR extracellular field potentials in the SOC.

Circulatory and metabolic effects of anemia in hyperinsulinemic ovine fetuses.

Infants born to women with poorly controlled diabetes mellitus have an increased incidence of perinatal asphyxia, cardiovascular abnormalities, elevated catecholamines, and sudden fetal death. Although hyperinsulinemic fetuses of diabetic women often exhibit polycythemia, they may also develop anemia because of pregnancy- and/or delivery-related complications. Experimental fetal hyperinsulinemia results in cardiovascular changes and a surge in catecholamines. We hypothesized that reductions in fetal O2 availability via anemic hypoxia limits O2 transport and compromises the hemodynamically and metabolically stressed but compensated hyperinsulinemic fetus. Chronically catheterized fetuses receiving insulin (n = 9) or placebo (n = 5) for 48 h were rendered anemic by an isovolemic exchange transfusion. In the hyperinsulinemic state, anemic-hypoxia augmented the insulin-mediated surge in norepinephrine concentration and increases in blood flow to brain, heart, and adrenal glands. Insulin-related increase in the combined ventricular output was sustained during anemia. O2 delivery to the fetus decreased, extraction increased, and O2 uptake did not change. Regional O2 delivery to the brain, kidney, gastrointestinal tract, muscle, fat, pancreas, spleen, and carcass decreased. Hyperinsulinemic ovine fetus exposed to anemic hypoxia demonstrated an accentuated surge in norepinephrine, a sustained increase in the combined ventricular output, preservation of systemic O2 uptake, and compromised regional O2 delivery to certain vascular regions. We conclude that the hyperinsulinemic fetus was able to compensate for anemic hypoxia by increased or sustained regional vascular perfusion.

Circulatory and metabolic effects of beta-adrenergic blockade in the hyperinsulinemic ovine fetus.

Offspring of women with poorly controlled diabetes exhibit hypoxemia, elevated catecholamine concentration at birth, and an increased incidence of fetal death. Experimental fetal hyperinsulinemia results in increased catecholamine concentration and hemodynamic changes including increased combined ventricular output and vasodilation of select fetal organs. We hypothesized that insulin-induced catecholamine-mediated beta-adrenergic stimulation supports some of these hemodynamic changes in the hyperinsulinemic ovine fetus. To study this, 24 chronically instrumented fetal sheep receiving insulin for 24 h were exposed to beta-(propranolol),beta 1-(metoprolol), and beta 2-(ICI 118,551) adrenergic blockade. Insulin infusion resulted in hyperinsulinemic-hypoglycemia, a surge in epinephrine and norepinephrine concentration, and increases in the combined ventricular output and regional blood flow to the heart, adrenal glands, kidney, gastrointestinal tract, liver, fat, muscle, carcass, and placenta. In the hyperinsulinemic state, beta-adrenergic blockade was associated with significant reductions in the combined ventricular output and blood flow to fat, carcass, lungs, and the placenta; beta 1-blockade was associated with reductions in the combined ventricular output and blood flow to the lungs; and beta 2-adrenergic blockade was associated with reductions in blood flow to muscle and lungs. Because beta-adrenergic blockade was associated with reductions in placental blood flow during hyperinsulinemia, oxygen and glucose metabolism were also compromised. We conclude that in the hyperinsulinemic-hypoglycemic normoxemic ovine fetus, insulin-induced catecholamine-mediated hemodynamic changes are modulated in part by beta-adrenergic receptor stimulation.

Plasticity of dendritic spine formation: a state-dependent stochastic process.

This study proposes that plasticity of dendritic spine formation may be modeled as distribution patterns imbedded in a spine length-dependent and density-dependent stochastic process. Modeling the jewel fish tectal interneuron revealed a critical 10-36 micron region where spine length plasticity was predicted to be most detectable. This hypothesis was tested by comparing neurons sampled from jewel fish reared for 4 years in a crowded environment (1 fish/5.64 l) with uncrowded controls (1 fish/25 l). The interaction between fish groups and the location of spine length differences was significant (p less than 0.01) within the basal 10-30 micron dendritic segment. Spine head widths were also significantly smaller (p less than 0.01) in the crowded fish over the entire dendrite. These findings suggest two modes of neuronal plasticity: (1) plasticity of spine length during formation, and (2) plasticity in spine head width after the spine is formed.

Specific transcriptional initiation in vitro on murine type C retrovirus promoters.

We have investigated the ability of molecularly cloned murine type C retroviral DNA to direct accurate initiation of RNA synthesis when added to cell-free extracts. Two different cloned proviruses were used. The first was derived from an integrated molecule of AKR murine leukemia virus and contains adjacent host information. The origin of the second was an unintegrated permuted copy of Harvey murine sarcoma virus. We found that the leukemia virus cloned provirus, as predicted by structural considerations, contained two functional RNA polymerase II promoters located in the U3 region present at either end of the molecule. These promoters initiate transcription at equal rates in vitro. We also found that the permuted sarcoma virus clone contained an RNA polymerase II promoter in the U3 region. Removal of viral sequences 49 bases upstream of the in vitro sarcoma virus initiation site by restriction cleavage results in loss of specific transcription, indicating a role for this information in in vitro promotion. The 5' ends of in vitro and in vivo viral RNA were compared by nuclease mapping techniques and found to be identical. Based on this evidence, we conclude that murine retroviral genomes contain sufficient information to initiate transcription independent of any host information in vitro and that these viral promoters are probably also active in vivo. In addition to the promoter in U3, Harvey murine sarcoma virus contains a second promoter in vitro that initiates near the 5' boundary of the transformation-specific (src) region of the virus. Initiation by this promoter was insensitive to low levels of alpha-amanitin, and the RNA transcript could be terminated to yield a 340-nucleotide product.

Circulatory and metabolic effects of hypoxia in the hyperinsulinemic ovine fetus.

Fetuses of women whose diabetes is poorly controlled often exhibit hypoxemia and elevated catecholamine concentrations at birth. In the ovine fetus, experimental hyperinsulinemia results in hypoxemia, elevated catecholamine concentrations, and hemodynamic changes. Limited oxygen availability occurring during pregnancy-related complications and/or delivery may present an additional risk to the hyperinsulinemic fetus. In this study, we tested the hypothesis that hypoxia induced via acutely limiting oxygen availability compromises the hemodynamically and metabolically stressed but compensated hyperinsulinemic ovine fetus. Fetuses receiving insulin (n = 8) or placebo (n = 5) for 48 h were exposed to maternally induced hypoxia. Hypoxic hypoxia was associated with a surge in catecholamines in the hyperinsulinemic fetuses. During hypoxia, this group exhibited insulin-related sustained increases in the combined ventricular output and fetal body blood flow, accentuation of the prior insulin-related increase in blood flow to the heart, decreased systemic oxygen delivery, accentuation of the insulin-related increased oxygen extraction, reductions in the insulin-related increased systemic oxygen uptake, sustained increases in regional oxygen delivery to the heart and adrenal glands, reductions in the insulin-related increased delivery to the carcass, and decreased oxygen delivery to the kidneys and gastrointestinal tract. We conclude that, in the hyperinsulinemic ovine fetus, hypoxic hypoxia attenuates the hyperinsulinemia-mediated increased systemic oxygen uptake. Regional oxygen transport is preserved to vital regions (brain, heart, and adrenal glands) by increased perfusion and compromised to certain other regions (kidneys and gastrointestinal tract), because the increases in perfusion are insufficient to offset the limited oxygen availability.

Glucocorticoid receptor-dependent disruption of a specific nucleosome on the mouse mammary tumor virus promoter is prevented by sodium butyrate.

Our laboratory has previously developed cell lines derived from mouse NIH 3T3 fibroblasts and C127 mammary tumor cells that stably express mouse mammary tumor virus (MMTV) long terminal repeat fusion genes in bovine papillomavirus-based episomes. Glucocorticoid hormone strongly activates transcription from episomes and induces the disruption of a single nucleosome in an array of phased nucleosomes on the MMTV promoter. Sodium butyrate inhibits the glucocorticoid hormone-dependent development of a nuclease-hypersensitive site that is due to the displacement of this nucleosome, and inhibits induction of RNA transcripts from episomes. Saturation binding studies show that butyrate treatment does not significantly affect the amount or the hormone-binding affinity of the glucocorticoid receptor. In a transient transfection assay, glucocorticoid hormone can activate transcription from a MMTV long terminal repeat-driven luciferase gene construct equivalently in untreated and butyrate-treated cells, indicating that the soluble factors necessary for transactivation of the MMTV promoter are unaffected by butyrate. The differential effect of butyrate on the induction of stable chromatin templates and transiently expressed plasmids suggests that butyrate prevents nucleosome displacement and represses transcription by inducing a modification of chromatin.

Glucocorticoid regulation of the Ha-MuSV p21 gene conferred by sequences from mouse mammary tumor virus.

Molecular chimeras with the p21 transforming gene of Harvey murine sarcoma virus linked to DNA containing the long terminal repeat (LTR) or mouse mammary tumor virus (MMTV) have been constructed. Transformants of NIH 3T3 cells induced by transfection with MMTV LTR-p21 hybrid DNA have been identified that express the normal p21 gene product. The levels of p21 RNA and protein in these transformants are regulated by physiological concentrations of dexamethasone, a synthetic glucocorticoid hormone. Hybrid transcripts containing p21 gene sequences originate at the normal MMTV viral initiation site. It is concluded that sequences necessary for hormonal control of transcription are completely specified by the viral genome and probably map within the viral LTR.

Effects of neonatal thyroxine, genotype, and noise on the ear and audiogenic seizures.

Seyfried, Glasser, and Yu recently reported that the DBA/2J mouse genotype, which is innately susceptible to audiogenic seizures, has high neonatal levels of thyroxine (T4) and that neonatal injections of T4 induce susceptibility in the C57BL/6J mouse. In the C57BL/6J mouse, neonatal T4 injections produced a temporary peripheral auditory dysfunction which appeared to be conductive in nature. A cochlear dysfunction was also seen in the DBA/2J mouse and in the acoustically primed C57BL/6J mouse. Since a peripheral auditory threshold elevation in these latter groups of mice appears to be causally related to their susceptibility to audiogenic seizures, it is likely that at least a portion of the susceptibility that Seyfried et al. reported was due to the effects of T4 on the ear.