A unified approach to the estimation and interpretation of resistance costs in plants

Plants exhibit a number of adaptive defence traits that endow resistance to past and current abiotic and biotic stresses. It is generally accepted that these adaptations will incur a cost when plants are not challenged by the stress to which they have become adapted--the so-called 'cost of adaptation'. The need to minimise or account for allelic variation at other fitness-related loci (genetic background control) is frequently overlooked when assessing resistance costs associated with plant defence traits. We provide a synthesis of the various experimental protocols that accomplish this essential requirement. We also differentiate those methods that enable the identification of the trait-specific or mechanistic basis of costs (direct methods) from those that provide an estimate of the impact of costs by examining the evolutionary trajectories of resistance allele frequencies at the population level (indirect methods). The advantages and disadvantages for each proposed experimental design are discussed. We conclude that plant resistance systems provide an ideal model to address fundamental questions about the cost of adaptation to stress. We also propose some ways to expand the scope of future studies for further fundamental and applied insight into the significance of adaptation costs.     

Spatial and temporal population genetic structure of the butterfly aglais urticae L. (Lepidoptera, nymphalidae)

The genetic diversity and the temporal and spatial genetic population structure of the butterfly Aglais urticae, a highly mobile species, were studied by allozyme electrophoresis. High levels of allozyme diversity were found. Most of the total genetic diversity occurred at the within-population scale rather than at the between-population scale. This variation could not be accounted for by Wright's model of 'isolation by distance'. No significant temporal variation was observed for those populations that were sampled in different years. A process combining high movement rate between neighbouring patches, long-distance migration and rare extinction/recolonization is suggested to explain the observed genetic structure. This hypothesis is favoured over an island model of population structure because migration in A. urticae is uniform neither with distance nor with time.     

Expression in a RabGAP yeast mutant of two human homologues, one of which is an oncogene

The yeast proteins Msb3p and Msb4p are two Ypt/Rab-specific GTPase-activating proteins (GAPs) involved in cell growth polarization. Both proteins share with a wide variety of other proteins the highly conserved TBC domain forming the catalytically active RabGAP domain. In particular, Msb3p and Msb4p are similar to the human proteins oncTre210p (the 786-amino-acid product of the human Tre2 oncogene, implicated in Ewing's sarcoma) and RN-tre (a Rab5-GAP controlling endocytosis of the EGFR). To further understand the biochemical function of Tre2 oncogene, we expressed its cDNA and, as a control, the RN-tre cDNA, in an msb3 msb4 double mutant yeast strain. Complementation data show that RN-tre can, unlike Tre2, replace the function of the MSB3 and MSB4 genes. As two highly conserved amino acids, including the catalytic arginine, are mutated in the oncTre210p TBC domain, we restored these two amino acids and expressed the modified Tre2 cDNA in the yeast mutant.     

Genetic Polymorphisms Affect Mouse and Human Trace Amine-Associated Receptor 1 Function

Methamphetamine (MA) and neurotransmitter precursors and metabolites such as tyramine, octopamine, and β-phenethylamine stimulate the G protein-coupled trace amine-associated receptor 1 (TAAR1). TAAR1 has been implicated in human conditions including obesity, schizophrenia, depression, fibromyalgia, migraine, and addiction. Additionally TAAR1 is expressed on lymphocytes and astrocytes involved in inflammation and response to infection. In brain, TAAR1 stimulation reduces synaptic dopamine availability and alters glutamatergic function. TAAR1 is also expressed at low levels in heart, and may regulate cardiovascular tone. Taar1 knockout mice orally self-administer more MA than wild type and are insensitive to its aversive effects. DBA/2J (D2) mice express a non-synonymous single nucleotide polymorphism (SNP) in Taar1 that does not respond to MA, and D2 mice are predisposed to high MA intake, compared to C57BL/6 (B6) mice. Here we demonstrate that endogenous agonists stimulate the recombinant B6 mouse TAAR1, but do not activate the D2 mouse receptor. Progeny of the B6XD2 (BxD) family of recombinant inbred (RI) strains have been used to characterize the genetic etiology of diseases, but contrary to expectations, BXDs derived 30–40 years ago express only the functional B6 Taar1 allele whereas some more recently derived BXD RI strains express the D2 allele. Data indicate that the D2 mutation arose subsequent to derivation of the original RIs. Finally, we demonstrate that SNPs in human TAAR1 alter its function, resulting in expressed, but functional, sub-functional and non-functional receptors. Our findings are important for identifying a predisposition to human diseases, as well as for developing personalized treatment options.     

Trade-off between immunocompetence and growth in magpies: an experimental study

A trade-off between immunity and growth has repeatedly been suggested, mainly based on laboratory and poultry science, but also from experiments where parasitism intensity was manipulated in field bird populations. However, as resource allocation to different activities (or organs) during growth is difficult to manipulate, this trade-off has only been experimentally tested by studying the effects of non-pathogenic antigens. By providing some nestling magpies (Pica pica) with methionine, a sulphur amino acid that specifically enhances T-cell immune response in chickens, we investigated this trade-off by directly affecting allocation of limited resources during growth. Results were in accordance with the hypothetical trade-off because nestlings fed with methionine showed a lower growth rate during the four days of methionine administration, but a larger response when fledglings were challenged with phytohaemagglutinin (a measure of the intensity of T-lymphocyte-mediated immune responsiveness) than control nestlings. Surprisingly, we found that control and experimental nestlings fledged with similar body mass, size and condition, but experimental nestlings suffered less from blood parasites (Haemoproteus) and had fewer lymphocytes (a widely used measure of health status) than control nestlings, suggesting a negative effect of blood parasites or other pathogens on nestling growth.     

Intracellular zinc release and ERK phosphorylation are required upstream of 12-lipoxygenase activation in peroxynitrite toxicity to mature rat oligodendrocytes

Peroxynitrite toxicity has been implicated in the pathogenesis of white matter injury. The mechanisms of peroxynitrite toxicity to oligodendrocytes (OLs), the major cell type of the white matter, are unknown. Using primary cultures of mature OLs that express myelin basic protein, we found that 3-morpholinosydnonimine, a peroxynitrite generator, caused toxicity to OLs. N,N,N',N'-tetrakis (2-pyridylmethyl)ethylenediamine, a zinc chelator, completely blocked peroxynitrite-induced toxicity. Use of FluoZin-3, a specific fluorescence zinc indicator, demonstrated the liberation of zinc from intracellular stores by peroxynitrite. Peroxynitrite caused the sequential activation of extracellular signal-regulated kinase 42/44 (ERK42/44), 12-lipoxygenase, and generation of reactive oxygen species, which were all dependent upon the intracellular release of zinc. The same cell death pathway was also activated when exogenous zinc was used. These results suggest that in addition to preventing the formation of peroxynitrite, useful strategies in preventing disease progression in pathologies in which peroxynitrite toxicity plays a critical role might include maintaining intracellular zinc homeostasis, blocking phosphorylation of ERK42/44, inhibiting activation of 12-lipoxygenase, and eliminating the accumulation of reactive oxygen species.     

PS1 activates PI3K thus inhibiting GSK-3 activity and tau overphosphorylation: effects of FAD mutations

Phosphatidylinositol 3-kinase (PI3K) promotes cell survival and communication by activating its downstream effector Akt kinase. Here we show that PS1, a protein involved in familial Alzheimer's disease (FAD), promotes cell survival by activating the PI3K/Akt cell survival signaling. This function of PS1 is unaffected by gamma-secretase inhibitors. Pharmacological and genetic evidence indicates that PS1 acts upstream of Akt, at or before PI3K kinase. PS1 forms complexes with the p85 subunit of PI3K and promotes cadherin/PI3K association. Furthermore, conditions that inhibit this association prevent the PS1-induced PI3K/Akt activation, indicating that PS1 stimulates PI3K/Akt signaling by promoting cadherin/PI3K association. By activating PI3K/Akt signaling, PS1 promotes phosphorylation/inactivation of glycogen synthase kinase-3 (GSK-3), suppresses GSK-3-dependent phosphorylation of tau at residues overphosphorylated in AD and prevents apoptosis of confluent cells. PS1 FAD mutations inhibit the PS1-dependent PI3K/Akt activation, thus promoting GSK-3 activity and tau overphosphorylation at AD-related residues. Our data raise the possibility that PS1 may prevent development of AD pathology by activating the PI3K/Akt signaling pathway. In contrast, FAD mutations may promote AD pathology by inhibiting this pathway.     

The cytoplasmic tails of Uukuniemi Virus (Bunyaviridae) G(N) and G(C) glycoproteins are important for intracellular targeting and the budding of virus-like particles

Functional motifs within the cytoplasmic tails of the two glycoproteins G(N) and G(C) of Uukuniemi virus (UUK) (Bunyaviridae family) were identified with the help of our recently developed virus-like particle (VLP) system for UUK virus (A. K. Overby, V. Popov, E. P. Neve, and R. F. Pettersson, J. Virol. 80:10428-10435, 2006). We previously reported that information necessary for the packaging of ribonucleoproteins into VLPs is located within the G(N) cytoplasmic tail (A. K. Overby, R. F. Pettersson, and E. P. Neve, J. Virol. 81:3198-3205, 2007). The G(N) glycoprotein cytoplasmic tail specifically interacts with the ribonucleoproteins and is critical for genome packaging. In addition, two other regions in the G(N) cytoplasmic tail, encompassing residues 21 to 25 and 46 to 50, were shown to be important for particle generation and release. By the introduction of point mutations within these two regions, we demonstrate that leucines at positions 23 and 24 are crucial for the initiation of VLP budding, while leucine 46, glutamate 47, and leucine 50 are important for efficient exit from the endoplasmic reticulum and subsequent transport to the Golgi complex. We found that budding and particle generation are highly dependent on the intracellular localization of both glycoproteins. The short cytoplasmic tail of UUK G(C) contains a lysine at position -3 from the C terminus that is highly conserved among members of the Phlebovirus, Hantavirus, and Orthobunyavirus genera. Mutating this single amino acid residue in G(C) resulted in the mislocalization of not only G(C) but also G(N) to the plasma membrane, and VLP generation was compromised in cells expressing this mutant. Together, these results demonstrate that the cytoplasmic tails of both G(N) and G(C) contain specific information necessary for efficient virus particle generation.     

Adaptation at different points along antibiotic concentration gradients

Antibiotic concentrations vary dramatically in the body and the environment. Hence, understanding the dynamics of resistance evolution along antibiotic concentration gradients is critical for predicting and slowing the emergence and spread of resistance. While it has been shown that increasing the concentration of an antibiotic slows resistance evolution, how adaptation to one antibiotic concentration correlates with fitness at other points along the gradient has not received much attention. Here, we selected populations of Escherichia coli at several points along a concentration gradient for three different antibiotics, asking how rapidly resistance evolved and whether populations became specialized to the antibiotic concentration they were selected on. Populations selected at higher concentrations evolved resistance more slowly but exhibited equal or higher fitness across the whole gradient. Populations selected at lower concentrations evolved resistance rapidly, but overall fitness in the presence of antibiotics was lower. However, these populations readily adapted to higher concentrations upon subsequent selection. Our results indicate that resistance management strategies must account not only for the rates of resistance evolution but also for the fitness of evolved strains.