Machine learning reveals sex‐specific 17β‐estradiol‐responsive expression patterns in white perch (Morone americana) plasma proteins

With growing abundance and awareness of endocrine disrupting compounds (EDCs) in the environment, there is a need for accurate and reliable detection of EDC exposure. Our objective in the present study was to observe differences within and between the global plasma proteomes of sexually mature male and female white perch (Morone americana) before (Initial Control, IC) and after 17β‐estradiol (E₂) induction. Semiquantitative nanoLC‐MS/MS data were analyzed by machine learning support vector machines (SVMs) and by two‐way ANOVA. By ANOVA, the expression levels of 44, 77, and 57 proteins varied significantly by gender, treatment, and the interaction of gender and treatment, respectively. SVMs perfectly classified male and female perch IC and E₂‐induced plasma samples using the protein expression data. E₂‐induced male and female perch plasma proteomes contained significantly higher levels of the yolk precursors vitellogenin Aa and Ab (VtgAa, VtgAb), as well as latrophilin and seven transmembrane domain‐containing protein 1 (Eltd1) and kininogen 1 (Kng1). This is the first report that Eltd1 and Kng1 may be E₂‐responsive proteins in fishes and therefore may be useful indicators of estrogen induction.     

Exploring the Interaction of SV2A with Racetams Using Homology Modelling,Molecular Dynamics and Site-Directed Mutagenesis

The putative Major Facilitator Superfamily (MFS) transporter, SV2A, is the target for levetiracetam (LEV), which is a successful anti-epileptic drug. Furthermore, SV2A knock out mice display a severe seizure phenotype and die after a few weeks. Despite this, the mode of action of LEV is not known at the molecular level. It would be extremely desirable to understand this more fully in order to aid the design of improved anti-epileptic compounds. Since there is no structure for SV2A, homology modelling can provide insight into the ligand-binding site. However, it is not a trivial process to build such models, since SV2A has low sequence identity to those MFS transporters whose structures are known. A further level of complexity is added by the fact that it is not known which conformational state of the receptor LEV binds to, as multiple conformational states have been inferred by tomography and ligand binding assays or indeed, if binding is exclusive to a single state. Here, we explore models of both the inward and outward facing conformational states of SV2A (according to the alternating access mechanism for MFS transporters). We use a sequence conservation analysis to help guide the homology modelling process and generate the models, which we assess further with Molecular Dynamics (MD). By comparing the MD results in conjunction with docking and simulation of a LEV-analogue used in radioligand binding assays, we were able to suggest further residues that line the binding pocket. These were confirmed experimentally. In particular, mutation of D670 leads to a complete loss of binding. The results shed light on the way LEV analogues may interact with SV2A and may help with the on-going design of improved anti-epileptic compounds.     

Can Butterflies Evade Fire? Pupa Location and Heat Tolerance in Fire Prone Habitats of Florida

Butterflies such as the atala hairstreak, Eumaeus atala Poey, and the frosted elfin, Callophrys irus Godart, are restricted to frequently disturbed habitats where their larval host plants occur. Pupae of these butterflies are noted to reside at the base of host plants or in the leaf litter and soil, which may allow them to escape direct mortality by fire, a prominent disturbance in many areas they inhabit. The capacity of these species to cope with fire is a critical consideration for land management and conservation strategies in the locations where they are found. Survival of E. atala pupae in relation to temperature and duration of heat pulse was tested using controlled water bath experiments and a series of prescribed fire field experiments. Survival of E. atala pupae was correlated to peak temperature and heat exposure in both laboratory and field trials. In addition, E. atala survival following field trials was correlated to depth of burial; complete mortality was observed for pupae at the soil surface. Fifty percent of E. atala survived the heat generated by prescribed fire when experimentally placed at depths ≥ 1.75cm, suggesting that pupation of butterflies in the soil at depth can protect from fatal temperatures caused by fire. For a species such as E. atala that pupates above ground, a population reduction from a burn event is a significant loss, and so decreasing the impact of prescribed fire on populations is critical.     

The stereoselective metabolism of halofantrine to desbutylhalofantrine in the rat: evidence of tissue-specific enantioselectivity in microsomal metabolism

The pharmacokinetics of the antimalarial drug (+/-)-halofantrine are stereoselective in humans and rats. To better understand the stereoselective metabolism of the drug to its primary metabolite, desbutylhalofantrine (DHF), a series of in vitro and in vivo experiments were undertaken in the rat. Formation of (-)-DHF exceeded that of (+)-DHF in liver microsomes [(-):(+) ratio of intrinsic formation clearances = 1.4]. In contrast, in intestinal microsomes no significant stereoselectivity was noted in the formation of the DHF enantiomers. Intestinal microsomes were also less efficient at producing the DHF enantiomers than were liver microsomes. Based on kinetic analysis of the DHF formation, there appeared to be more than one enzyme involved in the biotransformation. (+/-)-Ketoconazole (KTZ) effectively inhibited the formation of both DHF enantiomers by both liver and intestinal microsomes, although the reduction was more marked in liver microsomes. Through a combination of the use of CYP antibodies and recombinant CYP isoenzymes, the involvement of CYP 2B1/2, 3A1, 3A2, 1A1, 2C11, 2C6, 2D1, and 2D2 were implicated in the metabolism of halofantrine to DHF. Of these, CYP3A1/2 and CYP2C11 appeared to be the primary isoenzymes involved, although CYP2C11 showed greater (+)-DHF than (-)-DHF formation, whereas for CYP3A1 it was similar to the isolated rat liver microsomes. In vivo, oral (+/-)-KTZ caused significant increases in plasma halofantrine and decreases in DHF enantiomer plasma concentrations.     

Sugar receptors in Drosophila

The detection and discrimination of chemical compounds in potential foods are essential sensory processes when animals feed. The fruit fly Drosophila melanogaster employs 68 different gustatory receptors (GRs) for the detection of mostly nonvolatile chemicals that include sugars, a diverse group of toxic compounds present in many inedible plants and spoiled foods, and pheromones [1-6]. With the exception of a trehalose (GR5a) and a caffeine (GR66a) receptor [7-9], the functions of GRs involved in feeding are unknown. Here, we show that the Gr64 genes encode receptors for numerous sugars. We generated a fly strain that contained a deletion for all six Gr64 genes (DeltaGr64) and showed that these flies exhibit no or a significantly diminished proboscis extension reflex (PER) response when stimulated with glucose, maltose, sucrose, and several other sugars. The only considerable response was detected when Gr64 mutant flies were stimulated with fructose. Interestingly, response to trehalose is also abolished in these flies, even though they contain a functional Gr5a gene, which has been previously shown to encode a receptor for this sugar [8, 9]. This observation indicates that two or more Gr genes are necessary for trehalose detection, suggesting that GRs function as multimeric receptor complexes. Finally, we present evidence that some members of the Gr64 gene family are transcribed as a polycistronic mRNA, providing a mechanism for the coexpression of multiple sugar receptors in the same taste neurons.     

Breathing with big babies: ventilation and oxygen consumption during pregnancy in the lizard Tiliqua rugosa

We determined the effects of high gestational loads on ventilation and the rate of oxygen consumption (VO2) in the scincid lizard Tiliqua rugosa. Tiliqua rugosa is a large viviparous lizard that gives birth to one to four young after 6-7 mo gestation. Pregnant females gave birth to large young, weighing 89.5+/-5.9 g, which represents 21.6%+/-2.6% of maternal body mass. As the embryos developed, they occupied an increasingly large proportion of the body cavity, decreasing food consumption and compressing the gastrointestinal tract. Computerized axial tomography scans demonstrated that the lungs were compressed and/or regionally collapsed by the developing embryos, potentially compromising ventilation. Both minute ventilation (VE) and tidal volume decreased as gestation progressed, but no compensatory changes in breathing frequency or in the duration of the nonventilatory period were observed. The total rate of VO2, consisting of contributions from both maternal and fetal tissues, did not change during gestation, suggesting that maternal VO2 decreases as fetal VO2 increases. Pregnant females demonstrated a decreased ventilatory response to increased respiratory drive (triggered via inhalation of hypoxic hypercapnic gas), which may be associated with the increased energetic cost of ventilating a compressed lung or the desensitization of chemoreceptors during gestation. The decreased ability of the respiratory system to respond to increased respiratory drive may have important consequences for locomotor performance and predator avoidance in pregnant lizards.