A Method of Utilizing Accounting Records for Nurseries Producing Field Grown Stock

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L30A Mutation of Phospholemman Mimics Effects of Cardiac Glycosides in Isolated Cardiomyocytes

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Biochemistry, © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see http://pubs.acs.org/doi/abs/10.1021/acs.biochem.6b00633To determine if mutations introduced into phospholemman (PLM) could increase the level of PLM–Na,K-ATPase (NKA) binding, we performed scanning mutagenesis of the transmembrane domain of PLM and measured Förster resonance energy transfer (FRET) between each mutant and NKA. We observed an increased level of binding to NKA for several PLM mutants compared to that of the wild type (WT), including L27A, L30A, and I32A. In isolated cardiomyocytes, overexpression of WT PLM increased the amplitude of the Ca2+ transient compared to the GFP control. The Ca2+ transient amplitude was further increased by L30A PLM overexpression. The L30A mutation also delayed Ca2+ extrusion and increased the duration of cardiomyocyte contraction. This mimics aspects of the effect of cardiac glycosides, which are known to increase contractility through inhibition of NKA. No significant differences between WT and L30A PLM-expressing myocytes were observed after treatment with isoproterenol, suggesting that the superinhibitory effects of L30A are reversible with β-adrenergic stimulation. We also observed a decrease in the extent of PLM tetramerization with L30A compared to WT using FRET, suggesting that L30 is an important residue for mediating PLM–PLM binding. Molecular dynamics simulations revealed that the potential energy of the L30A tetramer is greater than that of the WT, and that the transmembrane α helix is distorted by the mutation. The results identify PLM residue L30 as an important determinant of PLM tetramerization and of functional inhibition of NKA by PLM.Peer reviewe

Differentiating between models of Epothilone binding to microtubules using tubulin mutagenesis, cytotoxicity, and molecular modeling

This is the peer reviewed version of the following article: Entwistle, R. A., Rizk, R. S., Cheng, D. M., Lushington, G. H., Himes, R. H., & Gupta, M. L. (2012). Differentiating between models of Epothilone binding to microtubules using tubulin mutagenesis, cytotoxicity, and molecular modeling. ChemMedChem, 7(9), 1580–1586. http://doi.org/10.1002/cmdc.201200286, which has been published in final form at doi.org/10.1002/cmdc.201200286. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.Microtubule stabilizers are powerful anti-mitotic compounds and represent a proven cancer treatment strategy. Several classes of compounds in clinical use or trials, such as the taxanes and epothilones, bind to the same region of β-tubulin. Determining how these molecules interact with tubulin and stabilize microtubules is important both for understanding the mechanism of action and enhancing chemotherapeutic potential, e.g. reducing side effects, increasing solubility, and overcoming resistance. Structural studies using nonpolymerized tubulin or stabilized polymers have produced different models of epothilone binding. Here, we used directed mutagenesis of the binding site on Saccharomyces cerevisiae β-tubulin to analyze interactions between Epothilone B and its biologically relevant substrate, dynamic microtubules. Five engineered amino acid changes contributed to a 125-fold increase in Epothilone B cytotoxicity independent of inherent microtubule stability. The mutagenesis of endogenous β-tubulin was done in otherwise isogenic strains. This facilitated the correlation of amino acid substitutions with altered cytotoxicity using molecular mechanics simulations. The results, which are based on the interaction between Epothilone B and dynamic microtubules, most strongly support the binding mode determined by NMR spectroscopy-based studies. This work establishes a system for discriminating between potential binding modes and among various compounds and/or analogues using a sensitive biological activity-based readout

In situ carbon uptake of marine macrophytes is highly variable among species, taxa, and morphology

Macroalgae form important coastal ecosystems and are considered to be highly productive, yet individual macrophyte carbon uptake rates are poorly documented and methodologies for in situ assessments of productivity are not well developed. In this study, we employ a 13C enrichment method in benthic chambers to calculate carbon uptake rates and assess δ13C signatures of a large stock of nearshore benthic macroalgae varying in taxa and morphology in Southern California. Our objectives are to 1) identify the variability of carbon uptake and inorganic carbon use among individuals of the same species or morphology, and 2) establish accurate and accessible carbon uptake procedures for coastal benthic primary producers. We found no significant relationship between the observed ranges of environmental factors such as nutrient concentrations, PAR, temperature, conductivity, and productivity rates, suggesting that unique physiological complexions underpin the high variability of carbon uptake and δ13C in studied macrophyte samples. We consider three reasons our experimental carbon uptake rates are 3–4 orders of magnitude lower than existing literature, which reports carbon uptake in the same units despite using different methods: 1) underrepresentation of Pmax, 2) incomplete carbon fractionation corrections, and 3) reduced hydrodynamics within the benthic chambers

Effect of antenatal milk expression education on lactation outcomes in birthing people with pre-pregnancy body mass index ≥ 25: Protocol for a randomized, controlled trial

Background: Birthing people with pre-pregnancy body mass indices (BMIs) ≥ 25 kg/m2, particularly those without prior breastfeeding experience, are at increased risk for suboptimal lactation outcomes. Antenatal milk expression (AME) may be one way to counteract the negative effects of early infant formula supplementation common in this population. Methods: This ongoing, randomized controlled trial in the United States evaluates the efficacy of a telelactation-delivered AME education intervention versus an attention control condition on lactation outcomes to 1 year postpartum among 280 nulliparous-to-primiparous, non-diabetic birthing people with pre-pregnancy BMI ≥ 25 kg/m2. The assigned study treatment is delivered via four weekly online video consultations between gestational weeks 37-40. Participants assigned to AME meet with study personnel and a lactation consultant to learn and practice AME. Instructions are provided for home practice of AME between study visits. Control group participants view videos on infant care/development at study visits. Participants complete emailed surveys at enrollment (340/7-366/7 gestational weeks) and 2 weeks, 6 weeks, 12 weeks, 6 months, and 12 months postpartum. Surveys assess lactation and infant feeding practices; breastfeeding self-efficacy, attitudes, and satisfaction; perception of insufficient milk; onset of lactogenesis-II; lactation support and problems; and reasons for breastfeeding cessation. Surveys also assess factors associated with lactation outcomes, including demographic characteristics, health problems, birth trauma, racial discrimination, and weight stigma. Health information and infant feeding data are abstracted from the pregnancy and birth center electronic health record. Milk samples are collected from the intervention group at each study visit and from both groups at each postpartum follow-up for future analyses. Qualitative interviews are conducted at 6 weeks postpartum to understand AME experiences. Primary outcomes of interest are breastfeeding exclusivity and breastfeeding self-efficacy scores at 2 weeks postpartum. Outcomes will be examined longitudinally with generalized linear mixed-effects modeling. Discussion: This is the first adequately powered trial evaluating the effectiveness of AME among U.S. birthing people and within a non-diabetic population with pre-pregnancy BMI ≥ 25 kg/m2. This study will also provide the first evidence of acceptability and effectiveness of telelactation-delivered AME. Trial registration: ClinicalTrials.gov: NCT04258709

Fisheries Exclusion Zones: Value and its sensitivity to data uncertainty

This paper focuses on the findings of the project that pertain particularly to the determination of the value of fisheries exclusion zones and the sensitivity of value estimation to data uncertainty. It necessarily draws on both the lite rature review and the case studies and in terms of its content extracts, presents and su mmarises the pertinent material therein covered. By collating this material a more directed consid eration of the topic of value is facilitated and the findings of the study as a whole more clear ly highlighted, and through consideration of uncertainty, also qualified