BRCA1 tumor suppression depends on BRCT phosphoprotein binding, but not its E3 ligase activity

Germline mutations of the breast cancer 1 (BRCA1) gene are a major cause of familial breast and ovarian cancer. The BRCA1 protein displays E3 ubiquitin ligase activity, and this enzymatic function is thought to be required for tumor suppression. To test this hypothesis, we generated mice that express an enzymatically defective Brca1. We found that this mutant Brca1 prevents tumor formation to the same degree as does wild-type Brca1 in three different genetically engineered mouse (GEM) models of cancer. In contrast, a mutation that ablates phosphoprotein recognition by the BRCA C terminus (BRCT) domains of BRCA1 elicits tumors in each of the three GEM models. Thus, BRCT phosphoprotein recognition, but not the E3 ligase activity, is required for BRCA1 tumor suppression.     

A comparison of florfenicol and tulathromycine for the treatment of undifferentiated fever in feedlot calves

The purpose of this study was to compare the relative cost-effectiveness of florfenicol with that of tulathromycin for treatment of undifferentiated fever (UF) in feedlot calves at ultra-high risk of developing UF that receive metaphylactic tulathromycin on arrival at the feedlot. Calves that received therapeutic florfenicol had lower overall mortality (P=.045) and bovine respiratory disease mortality (P=.050) compared with calves that received therapeutic tulathromycin, but no significant differences were detected in feedlot performance, carcass characteristics, or other animal health variables. There was a net advantage of Can$41.19/treated animal in the florfenicol group versus the tulathromycin group. This study demonstrates that it is more cost-effective to use florfenicol than tulathromycin for the initial treatment of UF in feedlot calves at ultra-high risk of developing UF that receive on-arrival metaphylactic tulathromycin.     

Using synchrotron-based FTIR microspectroscopy to reveal chemical features of feather protein secondary structure: comparison with other feed protein sources

Studying the secondary structure of proteins leads to an understanding of the components that make up a whole protein. An understanding of the structure of the whole protein is often vital to understanding its digestive behavior in animals and nutritive quality. Usually protein secondary structures include alpha-helix and beta-sheet. The percentages of these two structures in protein secondary structures may influence feed protein quality and digestive behavior. Feathers are widely available as a potential protein supplement. They are very high in protein (84%), but the digestibility of the protein is very low (5%). The objective of this study was to use synchrotron-based Fourier transform infrared (FTIR) microspectroscopy to reveal chemical features of feather protein secondary structure within amide I at ultraspatial resolution (pixel size = 10 x 10 microm), in comparison with other protein sources from easily digested feeds such as barley, oat, and wheat tissue at endosperm regions (without destruction of their inherent structure). This experiment was performed at beamline U2B of the Albert Einstein Center for Synchrotron Biosciences at the National Synchrotron Light Source (NSLS) in Brookhaven National Laboratory (BNL), U.S. Dept of Energy (NSLS-BNL, Upton, NY). The results showed that ultraspatially resolved chemical imaging of feed protein secondary structure in terms of beta-sheet to alpha-helix peak height ratio by stepping in pixel-sized increments was obtained. Using synchrotron FTIR microspectroscopy can distinguish structures of protein amide I among the different feed protein sources. The results show that the secondary structure of feather protein differed from those of other feed protein sources in terms of the line-shape and position of amide I. The feather protein amide I peaked at approximately 1630 cm(-1). However, other feed protein sources showed a peak at approximately 1650 cm(-1). By using multicomponent peak modeling, the relatively quantitative amounts of alpha-helix and beta-sheet in protein secondary structure were obtained, which showed that feather contains 88% beta-sheet and 4% alpha-helix, barley contains 17% beta-sheet and 71% alpha-helix, oat contains 2% beta-sheet and 92% alpha-helix, and wheat contains 42% beta-sheet and 50% alpha-helix. The difference in percentage of protein secondary structure may be part of the reason for different feed protein digestive behaviors. These results demonstrate the potential of highly spatially resolved infrared microspectroscopy to reveal feed protein secondary structure. Information from this study by the infrared probing of feed protein secondary structure may be valuable as a guide for feed breeders to improve and maintain protein quality for animal use.     

Black cohosh acts as a mixed competitive ligand and partial agonist of the serotonin receptor

Extracts of the rhizome of black cohosh [Actaea racemosa L., formerly called Cimicifuga racemosa (L.) Nutt.] were evaluated for potential mechanisms of action in the alleviation of menopausal hot flashes. Ovariectomized Sprague-Dawley rats were administered a 40% 2-propanol extract of black cohosh [4, 40, and 400 mg/(kg.day)] by gavage for 2 weeks with or without estradiol [50 microg/(kg.day)] to determine if black cohosh could act as an estrogen or antiestrogen on the basis of an increase in uterine weight or vaginal cellular cornification. No effects were observed on uterine weight or on vaginal cellular cornification in rats treated with black cohosh alone or in combination with 17beta-estradiol, indicating this black cohosh extract had no estrogenic or antiestrogenic properties in the ovariectomized rat model. To evaluate other potential pathways by which black cohosh might reduce menopausal hot flashes, serotonin activity was first assessed by the inhibition of radioligand binding to cell membrane preparations containing recombinant human serotonin receptor (5-HT) subtypes. A 40% 2-propanol extract of black cohosh was tested against 10 subtypes of the serotonin receptor, revealing the presence of compounds with strong binding to the 5-HT(1A), 5-HT(1D), and 5-HT(7) subtypes. Subsequent binding studies were carried out using 5-HT(1A) and 5-HT(7) receptors because of their association with the hypothalamus, which has been implicated in the generation of hot flashes. The black cohosh 40% 2-propanol extract inhibited [(3)H]lysergic acid diethylamide (LSD) binding to the human 5-HT(7) receptor (IC(50) = 2.4 +/- 0.4 microg/mL) with greater potency than binding of [(3)H]-8-hydroxy-2-(di-N-propylamino)tetralin to the rat 5-HT(1A) receptor (IC(50) = 13.9 +/- 0.6 microg/mL). Analysis of ligand binding data indicated that components of a black cohosh methanol extract functioned as a mixed competitive ligand of the 5-HT(7) receptor. In addition, a black cohosh methanol extract elevated cAMP levels in 293T-5-HT(7)-transfected HEK cells, suggesting the extract acted as a partial agonist at the receptor. The elevation in cAMP mediated by the black cohosh extract could be reversed in the presence of the antagonist methiothepin, indicating a receptor-mediated process. These data suggest that reductions in hot flashes in some women taking black cohosh may not be due to estrogenic properties. This study identifies other possible biological targets of black cohosh that could account for reported biological effects.     

The influence of landscape characteristics and home-range size on the quantification of landscape-genetics relationships

A common approach used to estimate landscape resistance involves comparing correlations of ecological and genetic distances calculated among individuals of a species. However, the location of sampled individuals may contain some degree of spatial uncertainty due to the natural variation of animals moving through their home range or measurement error in plant or animal locations. In this study, we evaluate the ways that spatial uncertainty, landscape characteristics, and genetic stochasticity interact to influence the strength and variability of conclusions about landscape-genetics relationships. We used a neutral landscape model to generate 45 landscapes composed of habitat and non-habitat, varying in percent habitat, aggregation, and structural connectivity (patch cohesion). We created true and alternate locations for 500 individuals, calculated ecological distances (least-cost paths), and simulated genetic distances among individuals. We compared correlations between ecological distances for true and alternate locations. We then simulated genotypes at 15 neutral loci and investigated whether the same influences could be detected in simple Mantel tests and while controlling for the effects of isolation-by-distance using the partial Mantel test. Spatial uncertainty interacted with the percentage of habitat in the landscape, but led to only small reductions in correlations. Furthermore, the strongest correlations occurred with low percent habitat, high aggregation, and low to intermediate levels of cohesion. Overall genetic stochasticity was relatively low and was influenced by landscape characteristics.     

Guayule as a feedstock for lignocellulosic biorefineries using ammonia fiber expansion (AFEX) pretreatment

Natural rubber latex extraction from guayule leaves behind greater than 90% (by weight) of agricultural residue as a feedstock suitable for conversion to biofuels via a thermochemical or biochemical route. Untreated guayule shrub and bagasse (after latex extraction) has shown to be very recalcitrant to enzymatic hydrolysis, necessitating application of a chemical pretreatment to enhance cellulase accessibility. The objective of this work was to carry out detailed compositional analysis, ammonia fiber expansion (AFEX¹) pretreatment, enzymatic hydrolysis and ethanol fermentation for various guayule-derived biomass fractions. Plant feedstocks tested were derived from two sources; (a) a mature 2007 AZ-2 whole guayule shrub plant obtained from USDA/ARS² research fields, and (b) the guayule latex-extracted commercial grade bagasse (62505) from Yulex Corporation. Compositional analysis and enzymatic hydrolysis were carried out using standard NREL³ protocols (www.nrel.gov/biomass/analytical_procedures.html). AFEX pretreatment was carried out using concentrated ammonium hydroxide at elevated temperatures for desired residence times in a pressurized reactor. Yeast fermentations on biomass hydrolyzates were carried out micro-aerobically using Saccharomyces cerevisiae (424A strain) in shake flasks.AFEX pretreatment was found to substantially improve overall enzymatic digestibility by 4-20 fold for both untreated guayule shrub and latex-extracted bagasse. Maximum glucan and xylan conversion achieved for the latex-extracted bagasse was 40% and 50%, respectively. The yeast was readily able to ferment both glucose and xylose to ethanol from the guayule bagasse hydrolyzate with or without external nutrient supplementation (i.e., yeast extract and tryptone). Our results highlight the possible utilization of guayule as a feedstock for lignocellulosic refineries co-producing natural rubber latex and biofuels. However, further process improvements (e.g., lignin/resin extraction and cellulose decrystallization using a modified AFEX process) are necessary to increase the effectiveness of ammonia-based pretreatments for further enhancing enzymatic digestibility of guayule-derived hardwood biomass.     

Effects of repeated hCG injections on reproductive efficiency in mares

Thirty reproductively sound mares were divided into treatment and control groups. In the treatment group, consisting of 14 mares, 2500 I.U. of human chorionic gonadotropin (hCG) was administered intravenously during estrus, in the presence of a 35 mm follicle over five successive cycles in 1987, and at least two cycles in 1988. Beginning with the second cycle of treatment in 1988, these mares were bred to a fertile stallion. The control group, consisting of 16 mares, was followed for two to five cycles in either the 1987 or 1988 season and six of these mares were bred to fertile stallions. Throughout the study period, blood was collected from the mares in the treatment group for analysis of anti-hCG antibodies and cross reactivity of the antibody to purified equine lutenizing hormone (eLH) and equine chorionic gonadotropin (eCG).In 1987, after the first three injections of hCG, mean duration of estrus in treated mares tended to be shorter than in control mares (P<.10). After all five hCG injections in 1987, mean ovulation time for treated mares was shorter than in control mares (P<0.01). However, after two to five hCG injections in 1987, seven treated mares (50%) had some individual ovulation times that did not differ from the control mares.Initially, following the first three injections of hCG in 1988, mean duration of estrus tended to be shorter (P<0.1) in treated mares compared to control mares. A reduction in mean ovulation time was observed after the first two hCG injections of 1988 (P<0.01). However, after one to four hCG injections in 1988, eight treated mares (57.1%) had some individual ovulation times that did not differ from controls.In 1987, all 14 treated mares developed significant levels of antibodies to hCG after one to four injections, and again in 1988, were positive for anti-hCG antibodies after one to three injections. However, no correlation was observed between magnitude of the immune response and duration of ovulation time or pregnancy rate. In cross reactivity studies, no significant binding of plasma anti-hCG antibodies to either eLH or eCG was observed in vitro.Overall, pregnancy and foaling rates of treated (85.7%) and control mares (83.3%) did not differ. Additionally, no difference was observed in number of inseminations per estrus between treated and control mares. In this study, with successive injections of hCG, the expected shortened time to ovulation was not elicited consistently in all mares. However, mares continued to ovulate, conceive and foal in the presence of significant levels of anti-hCG antibodies.