The influence of population structure on gene expression and flowering time variation in the ubiquitous weed Capsella bursa‐pastoris (Brassicaceae)

Population structure is a potential problem when testing for adaptive phenotypic differences among populations. The observed phenotypic differences among populations can simply be due to genetic drift, and if the genetic distance between them is not considered, the differentiation may be falsely interpreted as adaptive. Conversely, adaptive and demographic processes might have been tightly associated and correcting for the population structure may lead to false negatives. Here, we evaluated this problem in the cosmopolitan weed Capsella bursa‐pastoris. We used RNA‐Seq to analyse gene expression differences among 24 accessions, which belonged to a much larger group that had been previously characterized for flowering time and circadian rhythm and were genotyped using genotyping‐by‐sequencing (GBS) technique. We found that clustering of accessions for gene expression retrieved the same three clusters that were obtained with GBS data previously, namely Europe, the Middle East and Asia. Moreover, the three groups were also differentiated for both flowering time and circadian rhythm variation. Correction for population genetic structure when analysing differential gene expression analysis removed all differences among the three groups. This may suggest that most differences are neutral and simply reflect population history. However, geographical variation in flowering time and circadian rhythm indicated that the distribution of adaptive traits might be confounded by population structure. To bypass this confounding effect, we compared gene expression differentiation between flowering ecotypes within the genetic groups. Among the differentially expressed genes, FLOWERING LOCUS C was the strongest candidate for local adaptation in regulation of flowering time.     

The WEE1 regulators CPEB1 and miR-15b switch from inhibitor to activators at G2/M

MicroRNAs (miRNAs) in the AGO-containing RISC complex control messenger RNA (mRNA) translation by binding to mRNA 3′ untranslated region (3′UTR). The relationship between miRNAs and other regulatory factors that also bind to mRNA 3′UTR, such as CPEB1 (cytoplasmic polyadenylation element-binding protein), remains elusive. We found that both CPEB1 and miR-15b control the expression of WEE1, a key mammalian cell cycle regulator. Together, they repress WEE1 protein expression during G1 and S-phase. Interestingly, the 2 factors lose their inhibitory activity at the G2/M transition, at the time of the cell cycle when WEE1 expression is maximal, and, moreover, rather activate WEE1 translation in a synergistic manner. Our data show that translational regulation by RISC and CPEB1 is essential in cell cycle control and, most importantly, is coordinated, and can be switched from inhibition to activation during the cell cycle.     

Antiviral activity of benzimidazole derivatives. II. Antiviral activity of 2-phenylbenzimidazole derivatives

Seventy-six 2-phenylbenzimidazole derivatives were synthesized and evaluated in cell-based assays for cytotoxicity and antiviral activity against a panel of 10 RNA and DNA viruses. The most commonly affected viruses were, in decreasing order, CVB-2, BVDV, Sb-1, HSV-1, and YFV, while HIV-1 and VSV were not affected, and RSV, VV and Reo-1 were only susceptible to a few compounds. Thirty-nine compounds exhibited high activity (EC₅₀ = 0.1–10 μM) against at least one virus, and four of them were outstanding for their high and selective activity against VV (24, EC₅₀ = 0.1 μM) and BVDV (50, 51, and 53 with EC₅₀ = 1.5, 0.8, and 1.0 μM, respectively). The last compounds inhibited at low micromolar concentrations the NS5B RdRp of BVDV and also of HCV, the latter sharing structural similarity with the former. The considered compounds represent attractive leads for the development of antiviral agents against poxviruses, pestiviruses and even HCV, which are important human and veterinary pathogens.     

Origin and diversification dynamics of self-incompatibility haplotypes

Self-incompatibility (SI) is a genetic system found in some hermaphrodite plants. Recognition of pollen by pistils expressing cognate specificities at two linked genes leads to rejection of self pollen and pollen from close relatives, i.e., to avoidance of self-fertilization and inbred matings, and thus increased outcrossing. These genes generally have many alleles, yet the conditions allowing the evolution of new alleles remain mysterious. Evolutionary changes are clearly necessary in both genes, since any mutation affecting only one of them would result in a nonfunctional self-compatible haplotype. Here, we study diversification at the S-locus (i.e., a stable increase in the total number of SI haplotypes in the population, through the incorporation of new SI haplotypes), both deterministically (by investigating analytically the fate of mutations in an infinite population) and by simulations of finite populations. We show that the conditions allowing diversification are far less stringent in finite populations with recurrent mutations of the pollen and pistil genes, suggesting that diversification is possible in a panmictic population. We find that new SI haplotypes emerge fastest in populations with few SI haplotypes, and we discuss some implications for empirical data on S-alleles. However, allele numbers in our simulations never reach values as high as observed in plants whose SI systems have been studied, and we suggest extensions of our models that may reconcile the theory and data.     

Unusual N-glycan structures required for trafficking Toxoplasma gondii GAP50 to the inner membrane complex regulate host cell entry through parasite motility

Toxoplasma gondii motility, which is essential for host cell entry, migration through host tissues, and invasion, is a unique form of actin-dependent gliding. It is powered by a motor complex mainly composed of myosin heavy chain A, myosin light chain 1, gliding associated proteins GAP45, and GAP50, the only integral membrane anchor so far described. In the present study, we have combined glycomic and proteomic approaches to demonstrate that all three potential N-glycosylated sites of GAP50 are occupied by unusual N-glycan structures that are rarely found on mature mammalian glycoproteins. Using site-directed mutagenesis, we show that N-glycosylation is a prerequisite for GAP50 transport from the endoplasmic reticulum to the Golgi apparatus and for its subsequent delivery into the inner complex membrane. Assembly of key partners into the gliding complex, and parasite motility are severely impaired in the unglycosylated GAP50 mutants. Furthermore, comparative affinity purification using N-glycosylated and unglycosylated GAP50 as bait identified three novel hypothetical proteins including the recently described gliding associated protein GAP40, and we demonstrate that N-glycans are required for efficient binding to gliding partners. Collectively, these results provide the first detailed analyses of T. gondii N-glycosylation functions that are vital for parasite motility and host cell entry.     

Superparasitism evolution: adaptation or manipulation?

Superparasitism refers to the oviposition behavior of parasitoid females who lay their eggs in an already parasitized host. This often yields intense competition among larvae that are sharing the same host. Why would a female oviposit in such hostile habitat instead of looking for a better quality, unparasitized host? Here we present a continuous-time model of host-parasitoid interaction and discuss alternative scenarios. This model is first used to analyze the evolution of the superparasitism behavior of a solitary proovigenic parasitoid under both time and egg limitation. Then, following the recent discovery by Varaldi et al., we allow the parasitoid to be infected by a virus that alters the superparasitism behavior of its host to enhance its own horizontal transmission. The analysis of the coevolution of this manipulative behavior with the oviposition behavior of uninfected females clarifies and quantifies the conflict that emerges between the parasitoid and its virus. The model also yields new testable predictions. For example, we expect that uninfected parasitoids should superparasite less after coevolving with the manipulative virus. More generally, this model provides a theoretical framework for analyzing the evolution of the manipulation of parasitoid life-history traits by microparasites.     

Leptospirosis in Rio Grande do Sul,Brazil: An Ecosystem Approach in the Animal-Human Interface

Background

Leptospirosis is an epidemic-prone neglected disease that affects humans and animals, mostly in vulnerable populations. The One Health approach is a recommended strategy to identify drivers of the disease and plan for its prevention and control. In that context, the aim of this study was to analyze the distribution of human cases of leptospirosis in the State of Rio Grande do Sul, Brazil, and to explore possible drivers. Additionally, it sought to provide further evidence to support interventions and to identify hypotheses for new research at the human-animal-ecosystem interface.

Methodology and findings

The risk for human infection was described in relation to environmental, socioeconomic, and livestock variables. This ecological study used aggregated data by municipality (all 496). Data were extracted from secondary, publicly available sources. Thematic maps were constructed and univariate analysis performed for all variables. Negative binomial regression was used for multivariable statistical analysis of leptospirosis cases. An annual average of 428 human cases of leptospirosis was reported in the state from 2008 to 2012. The cumulative incidence in rural populations was eight times higher than in urban populations. Variables significantly associated with leptospirosis cases in the final model were: Parana/Paraiba ecoregion (RR: 2.25; CI_95%: 2.03–2.49); Neossolo Litolítico soil (RR: 1.93; CI_95%: 1.26–2.96); and, to a lesser extent, the production of tobacco (RR: 1.10; CI_95%: 1.09–1.11) and rice (RR: 1.003; CI_95%: 1.002–1.04).

Conclusion

Urban cases were concentrated in the capital and rural cases in a specific ecoregion. The major drivers identified in this study were related to environmental and production processes that are permanent features of the state. This study contributes to the basic knowledge on leptospirosis distribution and drivers in the state and encourages a comprehensive approach to address the disease in the animal-human-ecosystem interface.     

Cellular and biochemical features of skeletal muscle in obese Yucatan minipigs

Objective: To examine cellular and biochemical features of skeletal muscle in response to dietary‐induced obesity in a novel Yucatan minipig model of childhood obesity. Research Methods and Procedures: From 4 to 16 months of age, minipigs were fed either a recommended human‐type diet (NF; n = 4) or were overfed a western‐type diet with saturated fat and high‐glycemic index carbohydrates (OF, n = 4). Muscle samples (biceps femoris) were histochemically stained for the identification of intramuscular adipocytes, intramyocellular lipid aggregates (oil red O), and myofiber types (myosin ATPase, succinate dehydrogenase). Gene expressions and/or activities of factors involved in lipogenesis, lipolysis, or energetic metabolism were quantified in muscle. Results: Cross‐sectional areas of myofibers paralleled pig body weight (r = 0.86, p < 0.01). The size of intramuscular adipocytes, the relative proportion of oil red O‐stained fibers, and total muscle lipid content tended (p ≤ 0.10) to increase in response to OF diet. Hormone‐sensitive lipase, carnitine palmityl transferase‐I, and uncoupling protein 2 mRNA levels were lower (p < 0.05) in OF pigs than in NF pigs. Activities of β‐hydroxyacyl‐coenzyme A dehydrogenase and citrate synthase assessing post‐carnitine palmityl transferase I events and the proportion of oxidative myofibers were not altered by OF diet. Activity and gene expression of fatty acid synthase were lower (p < 0.02) in OF pigs than in NF pigs. Discussion: Overfeeding in Yucatan minipigs reduced the expression levels of three catabolic steps in skeletal muscle that are involved also in the etiology of human obesity.     

Crucial role of conserved lysine 277 in the fidelity of tRNA aminoacylation by Escherichia coli valyl-tRNA synthetase

Valyl-tRNA synthetase (ValRS) from Escherichia coli undergoes covalent valylation by a donor valyl adenylate synthesized by the enzyme itself. ValRS could also be modified, although to a lesser extent, by the noncognate isosteric substrate L-threonine from a donor threonyl adenylate synthesized by the synthetase itself, or by the nonsubstrate methionine from methionyl adenylate produced by catalytic amounts of methionyl-tRNA synthetase. MALDI mass spectrometry analysis designated lysines 154, 162, 170, 533, 554, 593, 894, 930, and 940 of ValRS as the target residues for the attachment of valine. Following autothreonylation, lysines 162, 170, 178, 277, 291, 554, 580, 593, 861, 894, and 930 were found to be modified. Finally, L-Met-labeled residues were lysines 118, 162, 170, 178, 277, and 938. Alignment of the available ValRS amino acid sequences showed that lysines 277 and 554 are strictly conserved (with the exception concerning replacement of Lys-277 with a methionine or a tyrosine in archaebacteria), suggesting that these residues might be functionally significant. Indeed, lysine 554 of ValRS is the first lysine of the Lys-Met-Ser-Lys-Ser signature of the catalytic site of class I aminoacyl-tRNA synthetases. Lys-277 which is labeled by L-threonine or L-methionine, and not by L-valine, is located at or near the editing site, in the three-dimensional structure of ValRS. The role of lysine 277 was evaluated by site-directed mutagenesis. The Lys277Ala mutant (K277A) exhibited a posttransfer Thr-tRNA(Val) editing rate that was significantly lower than that observed for the wild-type enzyme. In addition, the K277A substitution altered amino acid discrimination in the editing site, resulting in hydrolysis of the correctly charged cognate Val-tRNA(Val). Finally, significant amounts of mischarged Thr-tRNA(Val) were produced by the K277A mutant, and not by wild-type ValRS. Altogether, our results designate Lys-277 as a likely candidate for nucleophilic attack of misacylated tRNA in the editing site of ValRS.