Structure of the pentameric ligand-gated ion channel ELIC cocrystallized with its competitive antagonist acetylcholine

ELIC, the pentameric ligand-gated ion channel from Erwinia chrysanthemi, is a prototype for Cys-loop receptors. Here we show that acetylcholine is a competitive antagonist for ELIC. We determine the acetylcholine–ELIC cocrystal structure to a 2.9-Å resolution and find that acetylcholine binding to an aromatic cage at the subunit interface induces a significant contraction of loop C and other structural rearrangements in the extracellular domain. The side chain of the pore-lining residue F247 reorients and the pore size consequently enlarges, but the channel remains closed. We attribute the inability of acetylcholine to activate ELIC primarily to weak cation-π and electrostatic interactions in the pocket, because an acetylcholine derivative with a simple quaternary-to-tertiary ammonium substitution activates the channel. This study presents a compelling case for understanding the structural underpinning of the functional relationship between agonism and competitive antagonism in the Cys-loop receptors, providing a new framework for developing novel therapeutic drugs

Characterizing Ligand-Gated Ion Channel Receptors with Genetically Encoded Ca++ Sensors

We present a cell based system and experimental approach to characterize agonist and antagonist selectivity for ligand-gated ion channels (LGIC) by developing sensor cells stably expressing a Ca2+ permeable LGIC and a genetically encoded Förster (or fluorescence) resonance energy transfer (FRET)-based calcium sensor. In particular, we describe separate lines with human α7 and human α4β2 nicotinic acetylcholine receptors, mouse 5-HT3A serotonin receptors and a chimera of human α7/mouse 5-HT3A receptors. Complete concentration-response curves for agonists and Schild plots of antagonists were generated from these sensors and the results validate known pharmacology of the receptors tested. Concentration-response relations can be generated from either the initial rate or maximal amplitudes of FRET-signal. Although assaying at a medium throughput level, this pharmacological fluorescence detection technique employs a clonal line for stability and has versatility for screening laboratory generated congeners as agonists or antagonists on multiple subtypes of ligand-gated ion channels. The clonal sensor lines are also compatible with in vivo usage to measure indirectly receptor activation by endogenous neurotransmitters

No behavioural response to kin competition in a lekking species

The processes of kin selection and competition may occur simultaneously if limited individual dispersal i.e. population viscosity, is the only cause of the interactions between kin. Therefore, the net indirect benefits of a specific behaviour may largely depend on the existence of mechanisms dampening the fitness costs of competing with kin. In lekking species, males may increase the mating success of their close relatives (and hence gain indirect fitness benefits) because female prefer large leks. At the same time, kin selection may also lead to the evolution of mechanisms that dampen the costs of kin competition. As this mechanism has largely been ignored to date, we used detailed behavioural and genetic data collected in the black grouse Lyrurus tetrix to test whether males mitigate the costs of kin competition through the modulation of their fighting behaviours according to kinship and the avoidance of close relatives when establishing a lek territory. We found that neighbouring males’ fighting behaviour was unrelated to kinship and males did not avoid settling down with close relatives on leks. As males’ current and future mating success are strongly related to their behaviour on the lek (including fighting behaviour and territory position), the costs of kin competition may be negligible relative to the direct benefits of successful male-male contests. As we previously showed that the indirect fitness benefits of group membership were very limited in this black grouse population, these behavioural data support the idea that direct fitness benefits gained by successful male-male encounters likely outbalance any indirect fitness benefits

Patterns of genetic diversity in southern and southeastern Araucaria angustifolia (Bert.) O. Kuntze relict populations

Habitat fragmentation and a decrease in population size may lead to a loss in population genetic diversity. For the first time, the reduction in genetic diversity in the northernmost limit of natural occurence (southeastern Brazil) of Araucaria angustifolia in comparison with populations in the main area of the species continuous natural distribution (southern Brazil), was tested. The 673 AFLPs markers revealed a high level of genetic diversity for the species (Ht = 0.27), despite anthropogenic influence throughout the last century, and a decrease of H in isolated populations of southeastern Brazil (H = 0.16), thereby indicating the tendency for higher genetic diversity in remnant populations of continuous forests in southern Brazil, when compared to natural isolated populations in the southeastern region. A strong differentiation among southern and southeastern populations was detected (AMOVA variance ranged from 10%-15%). From Bayesian analysis, it is suggested that the nine populations tested form five “genetic clusters” (K = 5). Five of these populations, located in the northernmost limit of distribution of the species, represent three “genetic clusters”. These results are in agreement with the pattern of geographic distribution of the studied populations

The Effect of Recurrent Floods on Genetic Composition of Marble Trout Populations

A changing global climate can threaten the diversity of species and ecosystems. We explore the consequences of catastrophic disturbances in determining the evolutionary and demographic histories of secluded marble trout populations in Slovenian streams subjected to weather extremes, in particular recurrent flash floods and debris flows causing massive mortalities. Using microsatellite data, a pattern of extreme genetic differentiation was found among populations (global FST of 0.716), which exceeds the highest values reported in freshwater fish. All locations showed low levels of genetic diversity as evidenced by low heterozygosities and a mean of only 2 alleles per locus, with few or no rare alleles. Many loci showed a discontinuous allele distribution, with missing alleles across the allele size range, suggestive of a population contraction. Accordingly, bottleneck episodes were inferred for all samples with a reduction in population size of 3–4 orders of magnitude. The reduced level of genetic diversity observed in all populations implies a strong impact of genetic drift, and suggests that along with limited gene flow, genetic differentiation might have been exacerbated by recurrent mortalities likely caused by flash flood and debris flows. Due to its low evolutionary potential the species might fail to cope with an intensification and altered frequency of flash flood events predicted to occur with climate change

Is Promiscuity Associated with Enhanced Selection on MHC-DQα in Mice (genus Peromyscus)?

Reproductive behavior may play an important role in shaping selection on Major Histocompatibility Complex (MHC) genes. For example, the number of sexual partners that an individual has may affect exposure to sexually transmitted pathogens, with more partners leading to greater exposure and, hence, potentially greater selection for variation at MHC loci. To explore this hypothesis, we examined the strength of selection on exon 2 of the MHC-DQα locus in two species of Peromyscus. While the California mouse (P. californicus) is characterized by lifetime social and genetic monogamy, the deer mouse (P. maniculatus) is socially and genetically promiscuous; consistent with these differences in mating behavior, the diversity of bacteria present within the reproductive tracts of females is significantly greater for P. maniculatus. To test the prediction that more reproductive partners and exposure to a greater range of sexually transmitted pathogens are associated with enhanced diversifying selection on genes responsible for immune function, we compared patterns and levels of diversity at the Class II MHC-DQα locus in sympatric populations of P. maniculatus and P. californicus. Using likelihood based analyses, we show that selection is enhanced in the promiscuous P. maniculatus. This study is the first to compare the strength of selection in wild sympatric rodents with known differences in pathogen milieu