Elevated Prolactin during Pregnancy Drives a Phenotypic Switch in Mouse Hypothalamic Dopaminergic Neurons

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Next steps after 15 stimulating years of human gut microbiome research

Gut microbiome research has bloomed over the past 15 years. We have learnt a lot about the complex microbial communities that colonize our intestine. Promising avenues of research and microbiome-based applications are being implemented, with the goal of sustaining host health and applying personalized disease management strategies. Despite this exciting outlook, many fundamental questions about enteric microbial ecosystems remain to be answered. Organizational measures will also need to be taken to optimize the outcome of discoveries happening at an extremely rapid pace. This article highlights our own view of the field and perspectives for the next 15 years.     

Induction of drug metabolizing enzymes: a survey of in vitro methodologies and interpretations used in the pharmaceutical industry--do they comply with FDA recommendations?

The FDA has published guidelines by which to carry out and interpret in vitro induction studies using hepatocytes but do researchers in pharmaceutical companies actually follow these to the letter? In a survey of 30 participants in the pharmaceutical industry, 19 questions were posed regarding the species investigated, methodologies and interpretations of the data. Also addressed was the in-house decision making processes as a result of in vitro induction data. The survey showed that, although the basic methods were similar, no two researchers carried out and interpreted induction assays in exactly the same way. No single method was superior but all included enzyme activities as the major end point. Hepatocytes from animal species were used to confirm animal in vivo data but only human hepatocytes were used to predict human induction responses. If a compound was found to be positive in an in vitro induction assay, few would halt the development of the compound. The majority would consider other properties of the compound (bioavailability, clearance and therapeutic concentrations) and follow the FDA recommendation to conduct clinical drug-drug interaction studies. Overall, the results from this survey indicate that there is no standard pharmaceutical industry method or evaluation criterion by which in vitro assays are carried out. Rather than adhering to the FDA guidelines, some adapt methods and interpretation according to their own experience and need (whether screening or lead optimisation). There was general consensus that studies using human hepatocyte cultures currently provide the best indication of the in vivo induction potential of NCEs. In addition, the assessment of in vitro induction data from the literature suggest that the two-fold induction threshold and the percent of positive control criteria may not be the best methods to accurately assess the in vivo induction potential of a drug. Although the two-fold induction criterion is now obsolete, more predictive models for determining the clinical induction potential are needed. Alternative models are proposed and discussed herein.     

Functional roles for beta1,4-N-acetlygalactosaminyltransferase-A in Drosophila larval neurons and muscles

Adult Drosophila mutant for the glycosyltransferase beta1,4-N-acetlygalactosaminyltransferase-A (beta4GalNAcTA) display an abnormal locomotion phenotype, indicating a role for this enzyme, and the glycan structures that it generates, in the neuromuscular system. To investigate the functional role of this enzyme in more detail, we turned to the accessible larval neuromuscular system and report here that larvae mutant for beta4GalNAcTA display distinct nerve and muscle phenotypes. Mutant larvae exhibit abnormal backward crawling, reductions in nerve terminal bouton number, decreased spontaneous transmitter-release frequency, and short, wide muscles. This muscle shape change appears to result from hypercontraction since the individual sarcomeres are shorter in mutant muscles. Analysis of muscle calcium signals showed altered calcium handling in the mutant, suggesting a mechanism by which hypercontraction could occur. All of these phenotypes can be rescued by a transgene carrying the beta4GalNAcTA genomic region. Tissue-specific expression, using the Gal4-UAS system, reveals that neural expression rescues the mutant crawling phenotype, while muscle expression rescues the muscle defect. Tissue-specific expression did not appear to rescue the decrease in neuromuscular junction bouton number, suggesting that this defect arises from cooperation between nerve and muscle. Altogether, these results suggest that beta4GalNAcTA has at least three distinct functional roles.     

Insect pest management in the age of synthetic biology

Arthropod crop pests are responsible for 20% of global annual crop losses, a figure predicted to increase in a changing climate where the ranges of numerous species are projected to expand. At the same time, many insect species are beneficial, acting as pollinators and predators of pest species. For thousands of years, humans have used increasingly sophisticated chemical formulations to control insect pests but, as the scale of agriculture expanded to meet the needs of the global population, concerns about the negative impacts of agricultural practices on biodiversity have grown. While biological solutions, such as biological control agents and pheromones, have previously had relatively minor roles in pest management, biotechnology has opened the door to numerous new approaches for controlling insect pests. In this review, we look at how advances in synthetic biology and biotechnology are providing new options for pest control. We discuss emerging technologies for engineering resistant crops and insect populations and examine advances in biomanufacturing that are enabling the production of new products for pest control.     

Common and specific responses to iron and phosphorus deficiencies in roots of apple tree (Malus × domestica)

Plant Molecular Biology - Iron and phosphorus are abundant elements in soils but poorly available for plant nutrition. The availability of these two nutrients represents a major constraint for...     

Proline-specific aminopeptidase P prevents replication-associated genome instability

Genotoxic stress during DNA replication constitutes a serious threat to genome integrity and causes human diseases. Defects at different steps of DNA metabolism are known to induce replication stress, but the contribution of other aspects of cellular metabolism is less understood. We show that aminopeptidase P (APP1), a metalloprotease involved in the catabolism of peptides containing proline residues near their N-terminus, prevents replication-associated genome instability. Functional analysis of C. elegans mutants lacking APP-1 demonstrates that germ cells display replication defects including reduced proliferation, cell cycle arrest, and accumulation of mitotic DSBs. Despite these defects, app-1 mutants are competent in repairing DSBs induced by gamma irradiation, as well as SPO-11-dependent DSBs that initiate meiotic recombination. Moreover, in the absence of SPO-11, spontaneous DSBs arising in app-1 mutants are repaired as inter-homologue crossover events during meiosis, confirming that APP-1 is not required for homologous recombination. Thus, APP-1 prevents replication stress without having an apparent role in DSB repair. Depletion of APP1 (XPNPEP1) also causes DSB accumulation in mitotically-proliferating human cells, suggesting that APP1’s role in genome stability is evolutionarily conserved. Our findings uncover an unexpected role for APP1 in genome stability, suggesting functional connections between aminopeptidase-mediated protein catabolism and DNA replication.     

Examination of the Kinetics of Herpes Simplex Virus Glycoprotein D Binding to the Herpesvirus Entry Mediator, Using Surface Plasmon Resonance

Previously, we showed that truncated soluble forms of herpes simplex virus (HSV) glycoprotein D (gDt) bound directly to a truncated soluble form of the herpesvirus entry mediator (HveAt, formerly HVEMt), a cellular receptor for HSV. The purpose of the present study was to determine the affinity of gDt for HveAt by surface plasmon resonance and to compare and contrast the kinetics of an expanded panel of gDt variants in binding to HveAt in an effort to better understand the mechanism of receptor binding and virus entry. Both HveAt and gDt are dimers in solution and interact with a 2:1 stoichiometry. With HveAt, gD1(306t) (from the KOS strain of HSV-1) had a dissociation constant (K_D) of 3.2 × 10⁻⁶ M and gD2(306t) had a K_D of 1.5 × 10⁻⁶ M. The interaction between gDt and HveAt fits a 1:1 Langmuir binding model, i.e., two dimers of HveAt may act as one binding unit to interact with one dimer of gDt as the second binding unit. A gD variant lacking all signals for N-linked oligosaccharides had an affinity for HveAt similar to that of gD1(306t). A variant lacking the bond from cysteine 1 to cysteine 5 had an affinity for HveAt that did not differ from that of the wild type. However, variants with double cysteine mutations that eliminated either of the other two disulfide bonds showed decreased affinity for HveAt. This result suggests that two of the three disulfide bonds of gD are important for receptor binding. Four nonfunctional gDt variants, each representing one functional domain of gD, were also studied. Mutations in functional regions I and II drastically decreased the affinity of gDt for HveAt. Surprisingly, a variant with an insertion in functional region III had a wild-type level of affinity for HveAt, suggesting that this domain may function in virus entry at a step other than receptor binding. A variant with a deletion in functional region IV [gD1(Δ290-299t)] exhibited a 100-fold enhancement in affinity for HveAt (K_D = 3.3 × 10⁻⁸ M) due mainly to a 40-fold increase in its kinetic on rate. This agrees with the results of other studies showing the enhanced ability of gD1(Δ290-299t) to block infection. Interestingly, all the variants with decreased affinities for HveAt exhibited decreased kinetic on rates but only minor changes in their kinetic off rates. The results suggest that once the complex between gDt and HveAt forms, its stability is unaffected by a variety of changes in gD.     

The need of specific standards for head dimensions of urban Sardinian boys

In the 1997-1998 and 1998-1999 school years we measured head dimensions of 1600 boys from 6 to 13 years attending elementary and middle schools in towns of the Cagliari area (Sardinia, Italy). For each age, we compared the mean values for circumference, length and width of the head with Canadian standards, widely used by Sardinian pediatricians. The t-test shows that the means of the three variables are significantly lower in the Cagliari boys than in their Canadian contemporaries, with the exception of head circumference in 6 and 7 year-olds, and of head width in 10 year-olds. Therefore, it is necessary to produce specific growth charts for circumference, length and width of the head of Sardinian children rather than evaluate their growth using standards of populations with different ethnic, geographical and socio-economic backgrounds.