First chromosome scale genomes of ithomiine butterflies (Nymphalidae: Ithomiini): Comparative models for mimicry genetic studies

The ithomiine butterflies (Nymphalidae: Danainae) represent the largest known radiation of Müllerian mimetic butterflies. They dominate by number the mimetic butterfly communities, which include species such as the iconic neotropical Heliconius genus. Recent studies on the ecology and genetics of speciation in Ithomiini have suggested that sexual pheromones, colour pattern and perhaps hostplant could drive reproductive isolation. However, no reference genome was available for Ithomiini, which has hindered further exploration on the genetic architecture of these candidate traits, and more generally on the genomic patterns of divergence. Here, we generated high-quality, chromosome-scale genome assemblies for two Melinaea species, M. marsaeus and M. menophilus, and a draft genome of the species Ithomia salapia. We obtained genomes with a size ranging from 396 to 503 Mb across the three species and scaffold N50 of 40.5 and 23.2 Mb for the two chromosome-scale assemblies. Using collinearity analyses we identified massive rearrangements between the two closely related Melinaea species. An annotation of transposable elements and gene content was performed, as well as a specialist annotation to target chemosensory genes, which is crucial for host plant detection and mate recognition in mimetic species. A comparative genomic approach revealed independent gene expansions in ithomiines and particularly in gustatory receptor genes. These first three genomes of ithomiine mimetic butterflies constitute a valuable addition and a welcome comparison to existing biological models such as Heliconius, and will enable further understanding of the mechanisms of adaptation in butterflies.     

The Himalayas: Barrier and conduit for gene flow

The Himalayan mountain range is strategically located at the crossroads of the major cultural centers in Asia, the Middle East and Europe. Although previous Y‐chromosome studies indicate that the Himalayas served as a natural barrier for gene flow from the south to the Tibetan plateau, this region is believed to have played an important role as a corridor for human migrations between East and West Eurasia along the ancient Silk Road. To evaluate the effects of the Himalayan mountain range in shaping the maternal lineages of populations residing on either side of the cordillera, we analyzed mitochondrial DNA variation in 344 samples from three Nepalese collections (Newar, Kathmandu and Tamang) and a general population of Tibet. Our results revealed a predominantly East Asian‐specific component in Tibet and Tamang, whereas Newar and Kathmandu are both characterized by a combination of East and South Central Asian lineages. Interestingly, Newar and Kathmandu harbor several deep‐rooted Indian lineages, including M2, R5, and U2, whose coalescent times from this study (U2, >40 kya) and previous reports (M2 and R5, >50 kya) suggest that Nepal was inhabited during the initial peopling of South Central Asia. Comparisons with our previous Y‐chromosome data indicate sex‐biased migrations in Tamang and a founder effect and/or genetic drift in Tamang and Newar. Altogether, our results confirm that while the Himalayas acted as a geographic barrier for human movement from the Indian subcontinent to the Tibetan highland, it also served as a conduit for gene flow between Central and East Asia. Am J Phys Anthropol 151:169–182, 2013. © 2013 Wiley Periodicals, Inc.     

Dealing with paralogy in RADseq data: in silico detection and single nucleotide polymorphism validation in Robinia pseudoacacia L.

The RADseq technology allows researchers to efficiently develop thousands of polymorphic loci across multiple individuals with little or no prior information on the genome. However, many questions remain about the biases inherent to this technology. Notably, sequence misalignments arising from paralogy may affect the development of single nucleotide polymorphism (SNP) markers and the estimation of genetic diversity. We evaluated the impact of putative paralog loci on genetic diversity estimation during the development of SNPs from a RADseq dataset for the nonmodel tree species Robinia pseudoacacia L. We sequenced nine genotypes and analyzed the frequency of putative paralogous RAD loci as a function of both the depth of coverage and the mismatch threshold allowed between loci. Putative paralogy was detected in a very variable number of loci, from 1% to more than 20%, with the depth of coverage having a major influence on the result. Putative paralogy artificially increased the observed degree of polymorphism and resulting estimates of diversity. The choice of the depth of coverage also affected diversity estimation and SNP validation: A low threshold decreased the chances of detecting minor alleles while a high threshold increased allelic dropout. SNP validation was better for the low threshold (4×) than for the high threshold (18×) we tested. Using the strategy developed here, we were able to validate more than 80% of the SNPs tested by means of individual genotyping, resulting in a readily usable set of 330 SNPs, suitable for use in population genetics applications.     

Decorin reverses the repressive effect of autocrine-produced TGF-beta on mouse macrophage activation

Several cytokines or growth factors induce macrophages to proliferate, become activated, differentiate, or die through apoptosis. Like the major macrophage activator IFN-gamma, the extracellular matrix protein decorin inhibits proliferation and protects macrophages from the induction of apoptosis. Decorin enhances the IFN-gamma-induced expression of the IAalpha and IAbeta MHC class II genes. Moreover, it increases the IFN-gamma- or LPS-induced expression of inducible NO synthase, TNF-alpha, IL-1beta, and IL-6 genes and the secretion of these cytokines. Using a number of extracellular matrix proteins, we found a negative correlation between adhesion and proliferation. However, the effects of decorin on macrophage activation do not seem to be mediated through its effect on adhesion or proliferation. Instead, this proteoglycan abolishes the binding of TGF-beta to macrophages, as shown by Scatchard analysis of (125)I-labeled TGF-beta, which, in the absence of decorin, showed a K(d) of 0.11 +/- 0.03 nM and approximately 5000 receptors/cell. This was confirmed when we treated macrophages with Abs to block the endogenously produced TGF-beta, which enhanced macrophage activation in a way similar to decorin. The increase in activation mediated by decorin demonstrates that macrophages are under negative regulation that can be reversed by proteins of the extracellular matrix.     

Target selection and determination of function in structural genomics

The first crucial step in any structural genomics project is the selection and prioritization of target proteins for structure determination. There may be a number of selection criteria to be satisfied, including that the proteins have novel folds, that they be representatives of large families for which no structure is known, and so on. The better the selection at this stage, the greater is the value of the structures obtained at the end of the experimental process. This value can be further enhanced once the protein structures have been solved if the functions of the given proteins can also be determined. Here we describe the methods used at either end of the experimental process: firstly, sensitive sequence comparison techniques for selecting a high-quality list of target proteins, and secondly the various computational methods that can be applied to the eventual 3D structures to determine the most likely biochemical function of the proteins in question.     

Analysis of the CC chemokine receptor 5 (CCR5) delta-32 polymorphism in inflammatory bowel disease

The inflammatory bowel diseases (IBD) Crohn's disease (CD) and ulcerative colitis (UC) are complex multifactorial traits involving both environmental and genetic factors. Recent studies have shown the important role of pro-inflammatory cytokines and chemokines, including RANTES, in IBD. RANTES is the natural ligand for the CC-chemokine receptor 5 (CCR5). The chromosomal location of the CCR5 gene on 3p21 coincides with an IBD-susceptibility locus identified by genome-wide scanning. A 32-bp deletion (A32) in the CCR5 gene results in a nonfunctional receptor and is found with high frequency in Caucasians. In this study, we investigated the presence of the CCR5delta32 allele in a large cohort of IBD patients and in a healthy control population. Blood samples were obtained from 538 unselected IBD cases (433 unrelated IBD patients: 289 CD, 142 UC, 2 indeterminate colitis; 105 affected first-degree relatives) and 135 unaffected first-degree family members. Of the IBD patients, 36% had familial IBD with at least two members being affected. There were no significant differences in the CCR5delta32 mutation frequency between IBD patients and healthy controls, nor between CD and UC patients. There was no correlation between the CCR5delta32 genotype and the age at IBD-diagnosis, the frequency of surgical intervention, or disease localization. Only the association between CCR5delta32 homozygosity and the presence of anal lesions in CD patients was statistically significant (P=0.007). Analysis by the transmission/disequilibrium test showed no significant transmission distortion to the probands or their clinically silent siblings. Based on these results, it is unlikely that the CCR5delta32 allele is an important marker for predisposition to IBD.     

The Effect of Positive and Negative Feedback on Risk-Taking across Different Contexts

Preferences for risky choices have often been shown to be unstable and context-dependent. Though people generally avoid gambles with mixed outcomes, a phenomenon often attributed to loss aversion, contextual factors can impact this dramatically. For example, people typically prefer risky options after a financial loss, while generally choosing safer options after a monetary gain. However, it is unclear what exactly contributes to these preference shifts as a function of prior outcomes, as these gain/loss outcomes are usually confounded with participant performance, and therefore it is unclear whether these effects are driven purely by the monetary gains or losses, or rather by success or failure at the actual task. Here, we experimentally separated the effects of monetary gains/losses from performance success/failure prior to a standard risky choice. Participants performed a task in which they experienced contextual effects: 1) monetary gain or loss based directly on performance, 2) monetary gain or loss that was randomly awarded and was, crucially, independent from performance, and 3) success or failure feedback based on performance, but without any monetary incentive. Immediately following these positive/negative contexts, participants were presented with a gain-loss gamble that they had to decide to either play or pass. We found that risk preferences for identical sets of gambles were biased by positive and negative contexts containing monetary gains and losses, but not by contexts containing performance feedback. This data suggests that the observed framing effects are driven by aversion for monetary losses and not simply by the positive or negative valence of the context, or by potential moods resulting from positive or negative contexts. These results highlight the specific context dependence of risk preferences.     

Using Structural Information to Change the Phosphotransfer Specificity of a Two-Component Chemotaxis Signalling Complex

Two-component signal transduction pathways comprising histidine protein kinases (HPKs) and their response regulators (RRs) are widely used to control bacterial responses to environmental challenges. Some bacteria have over 150 different two-component pathways, and the specificity of the phosphotransfer reactions within these systems is tightly controlled to prevent unwanted crosstalk. One of the best understood two-component signalling pathways is the chemotaxis pathway. Here, we present the 1.40 Å crystal structure of the histidine-containing phosphotransfer domain of the chemotaxis HPK, CheA₃, in complex with its cognate RR, CheY₆. A methionine finger on CheY₆ that nestles in a hydrophobic pocket in CheA₃ was shown to be important for the interaction and was found to only occur in the cognate RRs of CheA₃, CheY₆, and CheB₂. Site-directed mutagenesis of this methionine in combination with two adjacent residues abolished binding, as shown by surface plasmon resonance studies, and phosphotransfer from CheA₃-P to CheY₆. Introduction of this methionine and an adjacent alanine residue into a range of noncognate CheYs, dramatically changed their specificity, allowing protein interaction and rapid phosphotransfer from CheA₃-P. The structure presented here has allowed us to identify specificity determinants for the CheA–CheY interaction and subsequently to successfully reengineer phosphotransfer signalling. In summary, our results provide valuable insight into how cells mediate specificity in one of the most abundant signalling pathways in biology, two-component signal transduction.