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 Since 1916, GENETICS has sought to publish significant advances in the field. To be accepted for publication, manuscripts must provide new insights into a biological process or demonstrate novel and creative approaches to an important biological problem or describe development of new resources, methods, technologies, or tools. And the study must be of interest to a wide range of genetics and genomics investigators. The editors of GENETICS seek to attract and publish articles that they believe will have a high impact on the field.   While it has a long and illustrious history, GENETICS has changed: it is not your mentor's journal. The editors make decisions quickly – in around 32 days – without sacrificing the excellence and scholarship for which the journal has long been known. GENETICS is constantly innovating; some of the new and expanded types of content include: YeastBook – a comprehensive compendium of reviews that presents the current state of knowledge of the molecular biology, cellular biology, and genetics of the yeast Saccharomyces cerevisiae. Educational Primers – tied to a current article in GENETICS, these educational resources lay out the necessary background (i.e., what was the question and why did that question matter?), explain the hypothesis or approach, describe the methodology, guide the reader through the results, and provide a precise summation of the discussion. Genetic Toolbox Reviews – describes practical and intellectual resources available for the study of less commonly used experimental organisms. Thompson Reuters JCR Impact Factor (2014): 5.963 EigenFactor (2014): 0.06335 Cited Half-life (2014): >10 years        What's Inside the Current Issue of GENETICS   Tuesday, October 11 2016 08:45:40 AM ISSUE HIGHLIGHTS [Issue Highlights] Tuesday, October 11 2016 08:45:40 AM Multiparental Populations: A Call for Papers [Call for Papers] Tuesday, October 11 2016 08:45:40 AM Fumio Tajima and the Origin of Modern Population Genetics [Classics] Nielsen, R. Tuesday, October 11 2016 08:45:40 AM Pritchard, Stephens, and Donnelly on Population Structure [Classics] Novembre, J. Tuesday, October 11 2016 08:45:40 AM Probing the Depths of Biological Diversity During the Second Century of GENETICS [Commentary] Sandell, L., Otto, S. P. Tuesday, October 11 2016 08:45:40 AM Darwins Influence on Mendel: Evidence from a New Translation of Mendels Paper [Perspectives] Fairbanks, D. J., Abbott, S. Gregor Mendel’s classic paper, Versuche über Pflanzen-Hybriden (Experiments on Plant Hybrids), was published in 1866, hence 2016 is its sesquicentennial. Mendel completed his experiments in 1863 and shortly thereafter began compiling the results and writing his paper, which he presented in meetings of the Natural Science Society in Brünn in February and March of 1865. Mendel owned a personal copy of Darwin’s Origin of Species, a German translation published in 1863, and it contains his marginalia. Its publication date indicates that Mendel’s study of Darwin’s book could have had no influence while he was conducting his experiments but its publication date coincided with the period of time when he was preparing his paper, making it possible that Darwin’s writings influenced Mendel’s interpretations and theory. Based on this premise, we prepared a Darwinized English translation of Mendel’s paper by comparing German terms Mendel employed with the same terms in the German translation of Origin of Species in his possession, then using Darwin’s counterpart English words and phrases as much as possible in our translation. We found a substantially higher use of these terms in the final two (10th and 11th) sections of Mendel’s paper, particularly in one key paragraph, where Mendel reflects on evolutionary issues, providing strong evidence of Darwin’s influence on Mendel. Tuesday, October 11 2016 08:45:40 AM Experiments on Plant Hybrids by Gregor Mendel [Perspectives] Abbott, S., Fairbanks, D. J. Tuesday, October 11 2016 08:45:40 AM The Evolutionary Origin and Genetic Makeup of Domestic Horses [Review] Librado, P., Fages, A., Gaunitz, C., Leonardi, M., Wagner, S., Khan, N., Hanghoj, K., Alquraishi, S. A., Alfarhan, A. H., Al-Rasheid, K. A., Der Sarkissian, C., Schubert, M., Orlando, L. The horse was domesticated only 5.5 KYA, thousands of years after dogs, cattle, pigs, sheep, and goats. The horse nonetheless represents the domestic animal that most impacted human history; providing us with rapid transportation, which has considerably changed the speed and magnitude of the circulation of goods and people, as well as their cultures and diseases. By revolutionizing warfare and agriculture, horses also deeply influenced the politico-economic trajectory of human societies. Reciprocally, human activities have circled back on the recent evolution of the horse, by creating hundreds of domestic breeds through selective programs, while leading all wild populations to near extinction. Despite being tightly associated with humans, several aspects in the evolution of the domestic horse remain controversial. Here, we review recent advances in comparative genomics and paleogenomics that helped advance our understanding of the genetic foundation of domestic horses. Tuesday, October 11 2016 08:45:40 AM Dosage Compensation in Drosophila--a Model for the Coordinate Regulation of Transcription [Gene Expression] Kuroda, M. I., Hilfiker, A., Lucchesi, J. C. The sex chromosomes have special significance in the history of genetics. The chromosomal basis of inheritance was firmly established when Calvin Bridges demonstrated that exceptions to Mendel’s laws of segregation were accompanied at the cytological level by exceptional sex chromosome segregation. The morphological differences between X and Y exploited in Bridges’ experiments arose as a consequence of the evolution of the sex chromosomes. Originally a homologous chromosome pair, the degeneration of the Y chromosome has been accompanied by a requirement for increased expression of the single X chromosome in males. Drosophila has been a model for the study of this dosage compensation and has brought key strengths, including classical genetics, the exceptional cytology of polytene chromosomes, and more recently, comprehensive genomics. The impact of these studies goes beyond sex chromosome regulation, providing valuable insights into mechanisms for the establishment and maintenance of chromatin domains, and for the coordinate regulation of transcription. Tuesday, October 11 2016 08:45:40 AM Next-Generation Sequencing-Based Approaches for Mutation Mapping and Identification in Caenorhabditis elegans [WormMethods] Doitsidou, M., Jarriault, S., Poole, R. J. The use of next-generation sequencing (NGS) has revolutionized the way phenotypic traits are assigned to genes. In this review, we describe NGS-based methods for mapping a mutation and identifying its molecular identity, with an emphasis on applications in Caenorhabditis elegans. In addition to an overview of the general principles and concepts, we discuss the main methods, provide practical and conceptual pointers, and guide the reader in the types of bioinformatics analyses that are required. Owing to the speed and the plummeting costs of NGS-based methods, mapping and cloning a mutation of interest has become straightforward, quick, and relatively easy. Removing this bottleneck previously associated with forward genetic screens has significantly advanced the use of genetics to probe fundamental biological processes in an unbiased manner. Tuesday, October 11 2016 08:45:40 AM Comparing the Statistical Fate of Paralogous and Orthologous Sequences [Statistical Genetics and Genomics] Massip, F., Sheinman, M., Schbath, S., Arndt, P. F. For several decades, sequence alignment has been a widely used tool in bioinformatics. For instance, finding homologous sequences with a known function in large databases is used to get insight into the function of nonannotated genomic regions. Very efficient tools like BLAST have been developed to identify and rank possible homologous sequences. To estimate the significance of the homology, the ranking of alignment scores takes a background model for random sequences into account. Using this model we can estimate the probability to find two exactly matching subsequences by chance in two unrelated sequences. For two homologous sequences, the corresponding probability is much higher, which allows us to identify them. Here we focus on the distribution of lengths of exact sequence matches between protein-coding regions of pairs of evolutionarily distant genomes. We show that this distribution exhibits a power-law tail with an exponent $$\alpha =-5.$$ Developing a simple model of sequence evolution by substitutions and segmental duplications, we show analytically and computationally that paralogous and orthologous gene pairs contribute differently to this distribution. Our model explains the differences observed in the comparison of coding and noncoding parts of genomes, thus providing a better understanding of statistical properties of genomic sequences and their evolution. Tuesday, October 11 2016 08:45:40 AM Statistical Methods for Testing Genetic Pleiotropy [Statistical Genetics and Genomics] Schaid, D. J., Tong, X., Larrabee, B., Kennedy, R. B., Poland, G. A., Sinnwell, J. P. Genetic pleiotropy is when a single gene influences more than one trait. Detecting pleiotropy and understanding its causes can improve the biological understanding of a gene in multiple ways, yet current multivariate methods to evaluate pleiotropy test the null hypothesis that none of the traits are associated with a variant; departures from the null could be driven by just one associated trait. A formal test of pleiotropy should assume a null hypothesis that one or no traits are associated with a genetic variant. For the special case of two traits, one can construct this null hypothesis based on the intersection-union (IU) test, which rejects the null hypothesis only if the null hypotheses of no association for both traits are rejected. To allow for more than two traits, we developed a new likelihood-ratio test for pleiotropy. We then extended the testing framework to a sequential approach to test the null hypothesis that $$k+1$$ traits are associated, given that the null of k traits are associated was rejected. This provides a formal testing framework to determine the number of traits associated with a genetic variant, while accounting for correlations among the traits. By simulations, we illustrate the type I error rate and power of our new methods; describe how they are influenced by sample size, the number of traits, and the trait correlations; and apply the new methods to multivariate immune phenotypes in response to smallpox vaccination. Our new approach provides a quantitative assessment of pleiotropy, enhancing current analytic practice. Tuesday, October 11 2016 08:45:40 AM A Comparison of One-Rate and Two-Rate Inference Frameworks for Site-Specific dN/dS Estimation [Statistical Genetics and Genomics] Spielman, S. J., Wan, S., Wilke, C. O. Two broad paradigms exist for inferring $$dN/dS,$$ the ratio of nonsynonymous to synonymous substitution rates, from coding sequences: (i) a one-rate approach, where $$dN/dS$$ is represented with a single parameter, or (ii) a two-rate approach, where $${d}_{\mathrm{N}}$$ and $${d}_{\mathrm{S}}$$ are estimated separately. The performances of these two approaches have been well studied in the specific context of proper model specification, i.e., when the inference model matches the simulation model. By contrast, the relative performances of one-rate vs. two-rate parameterizations when applied to data generated according to a different mechanism remain unclear. Here, we compare the relative merits of one-rate and two-rate approaches in the specific context of model misspecification by simulating alignments with mutation–selection models rather than with $$dN/dS$$-based models. We find that one-rate frameworks generally infer more accurate $$dN/dS$$ point estimates, even when $${d}_{\mathrm{S}}$$ varies among sites. In other words, modeling $${d}_{\mathrm{S}}$$ variation may substantially reduce accuracy of $$dN/dS$$ point estimates. These results appear to depend on the selective constraint operating at a given site. For sites under strong purifying selection ($$dN/dS\hspace{0.17em}\mathrm{\lesssim }\hspace{0.17em}0.3$$), one-rate and two-rate models show comparable performances. However, one-rate models significantly outperform two-rate models for sites under moderate-to-weak purifying selection. We attribute this distinction to the fact that, for these more quickly evolving sites, a given substitution is more likely to be nonsynonymous than synonymous. The data will therefore be relatively enriched for nonsynonymous changes, and modeling $${d}_{\mathrm{S}}$$ contributes excessive noise to $$dN/dS$$ estimates. We additionally find that high levels of divergence among sequences, rather than the number of sequences in the alignment, are more critical for obtaining precise point estimates. Tuesday, October 11 2016 08:45:40 AM Estimation of Gene Insertion/Deletion Rates with Missing Data [Statistical Genetics and Genomics] Dang, U. J., Devault, A. M., Mortimer, T. D., Pepperell, C. S., Poinar, H. N., Golding, G. B. Lateral gene transfer is an important mechanism for evolution among bacteria. Here, genome-wide gene insertion and deletion rates are modeled in a maximum-likelihood framework with the additional flexibility of modeling potential missing data. The performance of the models is illustrated using simulations and a data set on gene family phyletic patterns from Gardnerella vaginalis that includes an ancient taxon. A novel application involving pseudogenization/genome reduction magnitudes is also illustrated, using gene family data from Mycobacterium spp. Finally, an R package called indelmiss is available from the Comprehensive R Archive Network at https://cran.r-project.org/package=indelmiss, with support documentation and examples. Tuesday, October 11 2016 08:45:40 AM Biochemical Activities and Genetic Functions of the Drosophila melanogaster Fancm Helicase in DNA Repair [Genome Integrity and Transmission] Romero, N.-E., Matson, S. W., Sekelsky, J. Repair of DNA damage is essential to the preservation of genomic stability. During repair of double-strand breaks, several helicases function to promote accurate repair and prevent the formation of crossovers through homologous recombination. Among these helicases is the Fanconi anemia group M (FANCM) protein. FANCM is important in the response to various types of DNA damage and has been suggested to prevent mitotic crossovers during double-strand break repair. The helicase activity of FANCM is believed to be important in these functions, but no helicase activity has been detected in vitro. We report here a genetic and biochemical study of Drosophila melanogaster Fancm. We show that purified Fancm is a 3' to 5' ATP-dependent helicase that can disassemble recombination intermediates, but only through limited lengths of duplex DNA. Using transgenic flies expressing full-length or truncated Fancm, each with either a wild-type or mutated helicase domain, we found that there are helicase-independent and C-terminal-independent functions in responding to DNA damage and in preventing mitotic crossovers. Tuesday, October 11 2016 08:45:40 AM Accurate Chromosome Segregation at First Meiotic Division Requires AGO4, a Protein Involved in RNA-Dependent DNA Methylation in Arabidopsis thaliana [Genome Integrity and Transmission] Oliver, C., Santos, J. L., Pradillo, M. The RNA-directed DNA methylation (RdDM) pathway is important for the transcriptional repression of transposable elements and for heterochromatin formation. Small RNAs are key players in this process by regulating both DNA and histone methylation. Taking into account that methylation underlies gene silencing and that there are genes with meiosis-specific expression profiles, we have wondered whether genes involved in RdDM could play a role during this specialized cell division. To address this issue, we have characterized meiosis progression in pollen mother cells from Arabidopsis thaliana mutant plants defective for several proteins related to RdDM. The most relevant results were obtained for ago4-1. In this mutant, meiocytes display a slight reduction in chiasma frequency, alterations in chromatin conformation around centromeric regions, lagging chromosomes at anaphase I, and defects in spindle organization. These abnormalities lead to the formation of polyads instead of tetrads at the end of meiosis, and might be responsible for the fertility defects observed in this mutant. Findings reported here highlight an involvement of AGO4 during meiosis by ensuring accurate chromosome segregation at anaphase I. Tuesday, October 11 2016 08:45:40 AM Restructuring of Holocentric Centromeres During Meiosis in the Plant Rhynchospora pubera [Genome Integrity and Transmission] Marques, A., Schubert, V., Houben, A., Pedrosa-Harand, A. Centromeres are responsible for the correct segregation of chromosomes during mitosis and meiosis. Holocentric chromosomes, characterized by multiple centromere units along each chromatid, have particular adaptations to ensure regular disjunction during meiosis. Here we show by detecting CENH3, CENP-C, tubulin, and centromeric repeats that holocentromeres may be organized differently in mitosis and meiosis of Rhynchospora pubera. Contrasting to the mitotic linear holocentromere organization, meiotic centromeres show several clusters of centromere units (cluster-holocentromeres) during meiosis I. They accumulate along the poleward surface of bivalents where spindle fibers perpendicularly attach. During meiosis II, the cluster-holocentromeres are mostly present in the midregion of each chromatid. A linear holocentromere organization is restored after meiosis during pollen mitosis. Thus, a not yet described case of a cluster-holocentromere organization, showing a clear centromere restructuration between mitosis and meiosis, was identified in a holocentric organism. Tuesday, October 11 2016 08:45:40 AM Nicotinamide Suppresses the DNA Damage Sensitivity of Saccharomyces cerevisiae Independently of Sirtuin Deacetylases [Genome Integrity and Transmission] Rossl, A., Bentley-DeSousa, A., Tseng, Y.-C., Nwosu, C., Downey, M. Nicotinamide is both a reaction product and an inhibitor of the conserved sirtuin family of deacetylases, which have been implicated in a broad range of cellular functions in eukaryotes from yeast to humans. Phenotypes observed following treatment with nicotinamide are most often assumed to stem from inhibition of one or more of these enzymes. Here, we used this small molecule to inhibit multiple sirtuins at once during treatment with DNA damaging agents in the Saccharomyces cerevisiae model system. Since sirtuins have been previously implicated in the DNA damage response, we were surprised to observe that nicotinamide actually increased the survival of yeast cells exposed to the DNA damage agent MMS. Remarkably, we found that enhanced resistance to MMS in the presence of nicotinamide was independent of all five yeast sirtuins. Enhanced resistance was also independent of the nicotinamide salvage pathway, which uses nicotinamide as a substrate to generate NAD+, and of a DNA damage-induced increase in the salvage enzyme Pnc1. Our data suggest a novel and unexpected function for nicotinamide that has broad implications for its use in the study of sirtuin biology across model systems. Tuesday, October 11 2016 08:45:40 AM Divergent Transactivation of Maize Storage Protein Zein Genes by the Transcription Factors Opaque2 and OHPs [Gene Expression] Yang, J., Ji, C., Wu, Y. Maize transcription factors (TFs) opaque2 (O2) and the O2 heterodimerizing proteins (OHP1 and OHP2) originated from an ancient segmental duplication. The 22-kDa (z1C) and 19-kDa (z1A, z1B, and z1D) α-zeins are the most abundant storage proteins in maize endosperm. O2 is known to regulate α-zein gene expression, but its target motifs in the 19-kDa α-zein gene promoters have not been identified. The mechanisms underlying the regulation of α-zein genes by these TFs are also not well understood. In this study, we found that the O2 binding motifs in the α-zein gene promoters are quite flexible, with ACGT being present in the z1C and z1A promoters and a variant, ACAT, being present in the z1B and z1D promoters. OHPs recognized and transactivated all of the α-zein promoters, although to much lower levels than did O2. In the presence of O2, the suppression of OHPs did not cause a significant reduction in the transcription of α-zein genes, but in the absence of O2, OHPs were critical for the expression of residual levels of α-zeins. These findings demonstrated that O2 is the primary TF and that OHPs function as minor TFs in this process. This relationship is the converse of that involved in 27-kDa -zein gene regulation, indicating that the specificities of O2 and the OHPs for regulating zein genes diverged after gene duplication. The prolamine-box binding factor by itself has limited transactivation activity, but it promotes the binding of O2 to O2 motifs, resulting in the synergistic transactivation of α-zein genes. Tuesday, October 11 2016 08:45:40 AM Promoter Architecture and Sex-Specific Gene Expression in Daphnia pulex [Gene Expression] Raborn, R. T., Spitze, K., Brendel, V. P., Lynch, M. Large-scale transcription start site (TSS) profiling produces a high-resolution, quantitative picture of transcription initiation and core promoter locations within a genome. However, application of TSS profiling to date has largely been restricted to a small set of prominent model systems. We sought to characterize the cis-regulatory landscape of the water flea Daphnia pulex, an emerging model arthropod that reproduces both asexually (via parthenogenesis) and sexually (via meiosis). We performed Cap Analysis of Gene Expression (CAGE) with RNA isolated from D. pulex within three developmental states: sexual females, asexual females, and males. Identified TSSs were utilized to generate a "Daphnia Promoter Atlas," i.e., a catalog of active promoters across the surveyed states. Analysis of the distribution of promoters revealed evidence for widespread alternative promoter usage in D. pulex, in addition to a prominent fraction of compactly-arranged promoters in divergent orientations. We carried out de novo motif discovery using CAGE-defined TSSs and identified eight candidate core promoter motifs; this collection includes canonical promoter elements (e.g., TATA and Initiator) in addition to others lacking obvious orthologs. A comparison of promoter activities found evidence for considerable state-specific differential gene expression between states. Our work represents the first global definition of transcription initiation and promoter architecture in crustaceans. The Daphnia Promoter Atlas presented here provides a valuable resource for comparative study of cis-regulatory regions in metazoans, as well as for investigations into the circuitries that underpin meiosis and parthenogenesis. Tuesday, October 11 2016 08:45:40 AM Mitochondrial-Nuclear Interactions Mediate Sex-Specific Transcriptional Profiles in Drosophila [Gene Expression] Mossman, J. A., Tross, J. G., Li, N., Wu, Z., Rand, D. M. The assembly and function of mitochondria require coordinated expression from two distinct genomes, the mitochondrial DNA (mtDNA) and nuclear DNA (nDNA). Mutations in either genome can be a source of phenotypic variation, yet their coexpression has been largely overlooked as a source of variation, particularly in the emerging paradigm of mitochondrial replacement therapy. Here we tested how the transcriptome responds to mtDNA and nDNA variation, along with mitonuclear interactions (mtDNA x nDNA) in Drosophila melanogaster. We used two mtDNA haplotypes that differ in a substantial number of single nucleotide polymorphisms, with >100 amino acid differences. We placed each haplotype on each of two D. melanogaster nuclear backgrounds and tested for transcription differences in both sexes. We found that large numbers of transcripts were differentially expressed between nuclear backgrounds, and that mtDNA type altered the expression of nDNA genes, suggesting a retrograde, trans effect of mitochondrial genotype. Females were generally more sensitive to genetic perturbation than males, and males demonstrated an asymmetrical effect of mtDNA in each nuclear background; mtDNA effects were nuclear-background specific. mtDNA-sensitive genes were not enriched in male- or female-limited expression space in either sex. Using a variety of differential expression analyses, we show the responses to mitonuclear covariation to be substantially different between the sexes, yet the mtDNA genes were consistently differentially expressed across nuclear backgrounds and sexes. Our results provide evidence that the main mtDNA effects can be consistent across nuclear backgrounds, but the interactions between mtDNA and nDNA can lead to sex-specific global transcript responses. Tuesday, October 11 2016 08:45:40 AM Production of Small Noncoding RNAs from the flamenco Locus Is Regulated by the gypsy Retrotransposon of Drosophila melanogaster [Gene Expression] Guida, V., Cernilogar, F. M., Filograna, A., De Gregorio, R., Ishizu, H., Siomi, M. C., Schotta, G., Bellenchi, G. C., Andrenacci, D. Protective mechanisms based on RNA silencing directed against the propagation of transposable elements are highly conserved in eukaryotes. The control of transposable elements is mediated by small noncoding RNAs, which derive from transposon-rich heterochromatic regions that function as small RNA-generating loci. These clusters are transcribed and the precursor transcripts are processed to generate Piwi-interacting RNAs (piRNAs) and endogenous small interfering RNAs (endo-siRNAs), which silence transposable elements in gonads and somatic tissues. The flamenco locus is a Drosophila melanogaster small RNA cluster that controls gypsy and other transposable elements, and has played an important role in understanding how small noncoding RNAs repress transposable elements. In this study, we describe a cosuppression mechanism triggered by new euchromatic gypsy insertions in genetic backgrounds carrying flamenco alleles defective in gypsy suppression. We found that the silencing of gypsy is accompanied by the silencing of other transposons regulated by flamenco, and of specific flamenco sequences from which small RNAs against gypsy originate. This cosuppression mechanism seems to depend on a post-transcriptional regulation that involves both endo-siRNA and piRNA pathways and is associated with the occurrence of developmental defects. In conclusion, we propose that new gypsy euchromatic insertions trigger a post-transcriptional silencing of gypsy sense and antisense sequences, which modifies the flamenco activity. This cosuppression mechanism interferes with some developmental processes, presumably by influencing the expression of specific genes. Tuesday, October 11 2016 08:45:40 AM Mms21 SUMO Ligase Activity Promotes Nucleolar Function in Saccharomyces cerevisiae [Cellular Genetics] Kim, D.-H., Harris, B., Wang, F., Seidel, C., McCroskey, S., Gerton, J. L. The budding yeast E3 SUMO ligase Mms21, also known as Nse2, is a component of the Smc5/6 complex, which regulates sister chromatid cohesion, DNA replication, and repair. Our study shows that the mms21RING mutant exhibits (1) reduced ribosomal RNA production; (2) nuclear accumulation of ribosomal proteins; (3) elevated Gcn4 translation, indicating translational stress; and (4) upregulation of Gcn4 targets. Genes involved in ribosome biogenesis and translation are downregulated in the mms21RING mutant. We identified RPL19A as a novel genetic suppressor of the mms21RING mutant. Deletion of RPL19A partially suppresses growth defects in both smc5-6 and mms21RING mutants as well as nuclear accumulation of ribosome subunits in the mms21RING mutant. Deletion of a previously identified strong suppressor, MPH1, rescues both the accumulation of ribosome subunits and translational stress. This study suggests that the Smc5/6 complex supports nucleolar function. Tuesday, October 11 2016 08:45:40 AM Identification of the Target of the Retrograde Response that Mediates Replicative Lifespan Extension in Saccharomyces cerevisiae [Cellular Genetics] Jiang, J. C., Stumpferl, S. W., Tiwari, A., Qin, Q., Rodriguez-Quinones, J. F., Jazwinski, S. M. The retrograde response signals mitochondrial status to the nucleus, compensating for accumulating mitochondrial dysfunction during Saccharomyces cerevisiae aging and extending replicative lifespan. The histone acetylase Gcn5 is required for activation of nuclear genes and lifespan extension in the retrograde response. It is part of the transcriptional coactivators SAGA and SLIK, but it is not known which of these complexes is involved. Genetic manipulation showed that these complexes perform interchangeably in the retrograde response. These results, along with the finding that the histone deacetylase Sir2 was required for a robust retrograde response informed a bioinformatics screen that reduced to four the candidate genes causal for longevity of the 410 retrograde response target genes. Of the four, only deletion of PHO84 suppressed lifespan extension. Retrograde-response activation of PHO84 displayed some preference for SAGA. Increased PHO84 messenger RNA levels from a second copy of the gene in cells in which the retrograde response is not activated achieved >80% of the lifespan extension observed in the retrograde response. Our studies resolve questions involving the roles of SLIK and SAGA in the retrograde response, pointing to the cooperation of these complexes in gene activation. They also finally pinpoint the gene that is both necessary and sufficient to extend replicative lifespan in the retrograde response. The finding that this gene is PHO84 opens up a new set of questions about the mechanisms involved, as this gene is known to have pleiotropic effects. Tuesday, October 11 2016 08:45:40 AM Development of the Cellular Immune System of Drosophila Requires the Membrane Attack Complex/Perforin-Like Protein Torso-Like [Cellular Genetics] Forbes-Beadle, L., Crossman, T., Johnson, T. K., Burke, R., Warr, C. G., Whisstock, J. C. Pore-forming members of the membrane attack complex/perforin-like (MACPF) protein superfamily perform well-characterized roles as mammalian immune effectors. For example, complement component 9 and perforin function to directly form pores in the membrane of Gram-negative pathogens or virally infected/transformed cells, respectively. In contrast, the only known MACPF protein in Drosophila melanogaster, Torso-like, plays crucial roles during development in embryo patterning and larval growth. Here, we report that in addition to these functions, Torso-like plays an important role in Drosophila immunity. However, in contrast to a hypothesized effector function in, for example, elimination of Gram-negative pathogens, we find that torso-like null mutants instead show increased susceptibility to certain Gram-positive pathogens such as Staphylococcus aureus and Enterococcus faecalis. We further show that this deficit is due to a severely reduced number of circulating immune cells and, as a consequence, an impaired ability to phagocytose bacterial particles. Together these data suggest that Torso-like plays an important role in controlling the development of the Drosophila cellular immune system. Tuesday, October 11 2016 08:45:40 AM Complex Locomotion Behavior Changes Are Induced in Caenorhabditis elegans by the Lack of the Regulatory Leak K+ Channel TWK-7 [Developmental and Behavioral Genetics] Luersen, K., Gottschling, D.-C., Doring, F. The change of locomotion activity in response to external cues is a considerable achievement of animals and is required for escape responses, foraging, and other complex behaviors. Little is known about the molecular regulators of such an adaptive locomotion. The conserved eukaryotic two-pore domain potassium (K2P) channels have been recognized as regulatory K+ channels that modify the membrane potential of cells, thereby affecting, e.g., rhythmic muscle activity. By using the Caenorhabditis elegans system combined with cell-type-specific approaches and locomotion in-depth analyses, here, we found that the loss of K2P channel TWK-7 increases the locomotor activity of worms during swimming and crawling in a coordinated mode. Moreover, loss of TWK-7 function results in a hyperactive state that (although less pronounced) resembles the fast, persistent, and directed forward locomotion behavior of stimulated C. elegans. TWK-7 is expressed in several head neurons as well as in cholinergic excitatory and GABAergic inhibitory motor neurons. Remarkably, the abundance of TWK-7 in excitatory B-type and inhibitory D-type motor neurons affected five central aspects of adaptive locomotion behavior: velocity/frequency, wavelength/amplitude, direction, duration, and straightness. Hence, we suggest that TWK-7 activity might represent a means to modulate a complex locomotion behavior at the level of certain types of motor neurons. Tuesday, October 11 2016 08:45:41 AM Growth Coordination During Drosophila melanogaster Imaginal Disc Regeneration Is Mediated by Signaling Through the Relaxin Receptor Lgr3 in the Prothoracic Gland [Developmental and Behavioral Genetics] Jaszczak, J. S., Wolpe, J. B., Bhandari, R., Jaszczak, R. G., Halme, A. Damage to Drosophila melanogaster imaginal discs activates a regeneration checkpoint that (1) extends larval development and (2) coordinates the regeneration of the damaged disc with the growth of undamaged discs. These two systemic responses to damage are both mediated by Dilp8, a member of the insulin/insulin-like growth factor/relaxin family of peptide hormones, which is released by regenerating imaginal discs. Growth coordination between regenerating and undamaged imaginal discs is dependent on Dilp8 activation of nitric oxide synthase (NOS) in the prothoracic gland (PG), which slows the growth of undamaged discs by limiting ecdysone synthesis. Here we demonstrate that the Drosophila relaxin receptor homolog Lgr3, a leucine-rich repeat-containing G-protein-coupled receptor, is required for Dilp8-dependent growth coordination and developmental delay during the regeneration checkpoint. Lgr3 regulates these responses to damage via distinct mechanisms in different tissues. Using tissue-specific RNA-interference disruption of Lgr3 expression, we show that Lgr3 functions in the PG upstream of NOS, and is necessary for NOS activation and growth coordination during the regeneration checkpoint. When Lgr3 is depleted from neurons, imaginal disc damage no longer produces either developmental delay or growth inhibition. To reconcile these discrete tissue requirements for Lgr3 during regenerative growth coordination, we demonstrate that Lgr3 activity in both the CNS and PG is necessary for NOS activation in the PG following damage. Together, these results identify new roles for a relaxin receptor in mediating damage signaling to regulate growth and developmental timing. Tuesday, October 11 2016 08:45:41 AM Nationwide Genomic Study in Denmark Reveals Remarkable Population Homogeneity [Population and Evolutionary Genetics] Athanasiadis, G., Cheng, J. Y., Vilhjalmsson, B. J., Jorgensen, F. G., Als, T. D., Le Hellard, S., Espeseth, T., Sullivan, P. F., Hultman, C. M., Kjaergaard, P. C., Schierup, M. H., Mailund, T. Denmark has played a substantial role in the history of Northern Europe. Through a nationwide scientific outreach initiative, we collected genetic and anthropometrical data from ~800 high school students and used them to elucidate the genetic makeup of the Danish population, as well as to assess polygenic predictions of phenotypic traits in adolescents. We observed remarkable homogeneity across different geographic regions, although we could still detect weak signals of genetic structure reflecting the history of the country. Denmark presented genomic affinity with primarily neighboring countries with overall resemblance of decreasing weight from Britain, Sweden, Norway, Germany, and France. A Polish admixture signal was detected in Zealand and Funen, and our date estimates coincided with historical evidence of Wend settlements in the south of Denmark. We also observed considerably diverse demographic histories among Scandinavian countries, with Denmark having the smallest current effective population size compared to Norway and Sweden. Finally, we found that polygenic prediction of self-reported adolescent height in the population was remarkably accurate (R2 = 0.639 ± 0.015). The high homogeneity of the Danish population could render population structure a lesser concern for the upcoming large-scale gene-mapping studies in the country. Tuesday, October 11 2016 08:45:41 AM Estimating the Effective Population Size from Temporal Allele Frequency Changes in Experimental Evolution [Population and Evolutionary Genetics] Jonas, A., Taus, T., Kosiol, C., Schlotterer, C., Futschik, A. The effective population size ($${N}_{\mathrm{e}}$$) is a major factor determining allele frequency changes in natural and experimental populations. Temporal methods provide a powerful and simple approach to estimate short-term $${N}_{\mathrm{e}}.$$ They use allele frequency shifts between temporal samples to calculate the standardized variance, which is directly related to $${N}_{\mathrm{e}}.$$ Here we focus on experimental evolution studies that often rely on repeated sequencing of samples in pools (Pool-seq). Pool-seq is cost-effective and often outperforms individual-based sequencing in estimating allele frequencies, but it is associated with atypical sampling properties: Additional to sampling individuals, sequencing DNA in pools leads to a second round of sampling, which increases the variance of allele frequency estimates. We propose a new estimator of $${N}_{\mathrm{e}},$$ which relies on allele frequency changes in temporal data and corrects for the variance in both sampling steps. In simulations, we obtain accurate $${N}_{\mathrm{e}}$$ estimates, as long as the drift variance is not too small compared to the sampling and sequencing variance. In addition to genome-wide $${N}_{\mathrm{e}}$$ estimates, we extend our method using a recursive partitioning approach to estimate $${N}_{\mathrm{e}}$$ locally along the chromosome. Since the type I error is controlled, our method permits the identification of genomic regions that differ significantly in their $${N}_{\mathrm{e}}$$ estimates. We present an application to Pool-seq data from experimental evolution with Drosophila and provide recommendations for whole-genome data. The estimator is computationally efficient and available as an R package at https://github.com/ThomasTaus/Nest. Tuesday, October 11 2016 08:45:41 AM Commonalities in Development of Pure Breeds and Population Isolates Revealed in the Genome of the Sardinian Fonni's Dog [Population and Evolutionary Genetics] Dreger, D. L., Davis, B. W., Cocco, R., Sechi, S., Di Cerbo, A., Parker, H. G., Polli, M., Marelli, S. P., Crepaldi, P., Ostrander, E. A. The island inhabitants of Sardinia have long been a focus for studies of complex human traits due to their unique ancestral background and population isolation reflecting geographic and cultural restriction. Population isolates share decreased genomic diversity, increased linkage disequilibrium, and increased inbreeding coefficients. In many regions, dogs and humans have been exposed to the same natural and artificial forces of environment, growth, and migration. Distinct dog breeds have arisen through human-driven selection of characteristics to meet an ideal standard of appearance and function. The Fonni’s Dog, an endemic dog population on Sardinia, has not been subjected to an intensive system of artificial selection, but rather has developed alongside the human population of Sardinia, influenced by geographic isolation and unregulated selection based on its environmental adaptation and aptitude for owner-desired behaviors. Through analysis of 28 dog breeds, represented with whole-genome sequences from 13 dogs and ~170,000 genome-wide single nucleotide variants from 155 dogs, we have produced a genomic illustration of the Fonni’s Dog. Genomic patterns confirm within-breed similarity, while population and demographic analyses provide spatial identity of Fonni’s Dog to other Mediterranean breeds. Investigation of admixture and fixation indices reveals insights into the involvement of Fonni’s Dogs in breed development throughout the Mediterranean. We describe how characteristics of population isolates are reflected in dog breeds that have undergone artificial selection, and are mirrored in the Fonni’s Dog through traditional isolating factors that affect human populations. Lastly, we show that the genetic history of Fonni’s Dog parallels demographic events in local human populations. Tuesday, October 11 2016 08:45:41 AM Repeated Duplication of Argonaute2 Is Associated with Strong Selection and Testis Specialization in Drosophila [Population and Evolutionary Genetics] Lewis, S. H., Webster, C. L., Salmela, H., Obbard, D. J. Argonaute2 (Ago2) is a rapidly evolving nuclease in the Drosophila melanogaster RNA interference (RNAi) pathway that targets viruses and transposable elements in somatic tissues. Here we reconstruct the history of Ago2 duplications across the D. obscura group and use patterns of gene expression to infer new functional specialization. We show that some duplications are old, shared by the entire species group, and that losses may be common, including previously undetected losses in the lineage leading to D. pseudoobscura. We find that while the original (syntenic) gene copy has generally retained the ancestral ubiquitous expression pattern, most of the novel Ago2 paralogs have independently specialized to testis-specific expression. Using population genetic analyses, we show that most testis-specific paralogs have significantly lower genetic diversity than the genome-wide average. This suggests recent positive selection in three different species, and model-based analyses provide strong evidence of recent hard selective sweeps in or near four of the six D. pseudoobscura Ago2 paralogs. We speculate that the repeated evolution of testis specificity in obscura group Ago2 genes, combined with their dynamic turnover and strong signatures of adaptive evolution, may be associated with highly derived roles in the suppression of transposable elements or meiotic drive. Our study highlights the lability of RNAi pathways, even within well-studied groups such as Drosophila, and suggests that strong selection may act quickly after duplication in RNAi pathways, potentially giving rise to new and unknown RNAi functions in nonmodel species. Tuesday, October 11 2016 08:45:41 AM Imputation of DNA Methylation Levels in the Brain Implicates a Risk Factor for Parkinsons Disease [Genetics of Complex Traits] Rawlik, K., Rowlatt, A., Tenesa, A. Understanding how genetic variation affects intermediate phenotypes, like DNA methylation or gene expression, and how these in turn vary with complex human disease provides valuable insight into disease etiology. However, intermediate phenotypes are typically tissue and developmental stage specific, making relevant phenotypes difficult to assay. Assembling large case–control cohorts, necessary to achieve sufficient statistical power to assess associations between complex traits and relevant intermediate phenotypes, has therefore remained challenging. Imputation of such intermediate phenotypes represents a practical alternative in this context. We used a mixed linear model to impute DNA methylation (DNAm) levels of four brain tissues at up to 1826 methylome-wide sites in 6259 patients with Parkinson’s disease and 9452 controls from across five genome-wide association studies (GWAS). Six sites, in two regions, were found to associate with Parkinson’s disease for at least one tissue. While a majority of identified sites were within an established risk region for Parkinson’s disease, suggesting a role of DNAm in mediating previously observed genetic effects at this locus, we also identify an association with four CpG sites in chromosome 16p11.2. Direct measures of DNAm in the substantia nigra of 39 cases and 13 control samples were used to independently replicate these four associations. Only the association at cg10917602 replicated with a concordant direction of effect (P = 0.02). cg10917602 is 87 kb away from the closest reported GWAS hit. The employed imputation methodology implies that variation of DNAm levels at cg10917602 is predictive for Parkinson’s disease risk, suggesting a possible causal role for methylation at this locus. More generally this study demonstrates the feasibility of identifying predictive epigenetic markers of disease risk from readily available data sets. Tuesday, October 11 2016 08:45:41 AM Gene and Network Analysis of Common Variants Reveals Novel Associations in Multiple Complex Diseases [Genetics of Complex Traits] Nakka, P., Raphael, B. J., Ramachandran, S. Genome-wide association (GWA) studies typically lack power to detect genotypes significantly associated with complex diseases, where different causal mutations of small effect may be present across cases. A common, tractable approach for identifying genomic elements associated with complex traits is to evaluate combinations of variants in known pathways or gene sets with shared biological function. Such gene-set analyses require the computation of gene-level P-values or gene scores; these gene scores are also useful when generating hypotheses for experimental validation. However, commonly used methods for generating GWA gene scores are computationally inefficient, biased by gene length, imprecise, or have low true positive rate (TPR) at low false positive rates (FPR), leading to erroneous hypotheses for functional validation. Here we introduce a new method, PEGASUS, for analytically calculating gene scores. PEGASUS produces gene scores with as much as 10 orders of magnitude higher numerical precision than competing methods. In simulation, PEGASUS outperforms existing methods, achieving up to 30% higher TPR when the FPR is fixed at 1%. We use gene scores from PEGASUS as input to HotNet2 to identify networks of interacting genes associated with multiple complex diseases and traits; this is the first application of HotNet2 to common variation. In ulcerative colitis and waist–hip ratio, we discover networks that include genes previously associated with these phenotypes, as well as novel candidate genes. In contrast, existing methods fail to identify these networks. We also identify networks for attention-deficit/hyperactivity disorder, in which GWA studies have yet to identify any significant SNPs. Tuesday, October 11 2016 08:45:41 AM Insights into Interspecific Hybridization Events in Allotetraploid Cotton Formation from Characterization of a Gene-Regulating Leaf Shape [Genetics of Complex Traits] Chang, L., Fang, L., Zhu, Y., Wu, H., Zhang, Z., Liu, C., Li, X., Zhang, T. The morphology of cotton leaves varies considerably. Phenotypes, including okra, sea-island, super-okra, and broad leaf, are controlled by a multiple allele locus, L2. Okra leaf (L2°) is an incomplete mutation that alters leaf shape by increasing the length of lobes with deeper sinuses. Using a map-based cloning strategy, we cloned the L2 locus gene, which encodes a LATE MERISTEM IDENTITY 1 (LMI1)-like transcription factor (GhOKRA). Silencing GhOKRA leads to a change in phenotype from okra to broad leaf. Overexpression of GhOKRA in Arabidopsis thaliana greatly increases the degree of the leaf lobes and changes the leaf shape. Premature termination of translation in GhOKRA results in the production of broad leaves. The sequences of OKRA from diploid progenitor D-genome species, and wild races and domesticated allotetraploid cottons in Gossypium hirsutum show that a premature termination mutation occurred before and after the formation of tetraploid cotton, respectively. This study provides genomic insights into the two interspecific hybridization events: one produced the present broad leaf and another formed okra leaf phenotype with complete OKRA, that occurred during allotetraploid cotton formation. Tuesday, October 11 2016 08:45:41 AM A Synthetic Dosage Lethal Genetic Interaction Between CKS1B and PLK1 Is Conserved in Yeast and Human Cancer Cells [Genome and Systems Biology] Reid, R. J. D., Du, X., Sunjevaric, I., Rayannavar, V., Dittmar, J., Bryant, E., Maurer, M., Rothstein, R. The CKS1B gene located on chromosome 1q21 is frequently amplified in breast, lung, and liver cancers. CKS1B codes for a conserved regulatory subunit of cyclin–CDK complexes that function at multiple stages of cell cycle progression. We used a high throughput screening protocol to mimic cancer-related overexpression in a library of Saccharomyces cerevisiae mutants to identify genes whose functions become essential only when CKS1 is overexpressed, a synthetic dosage lethal (SDL) interaction. Mutations in multiple genes affecting mitotic entry and mitotic exit are highly enriched in the set of SDL interactions. The interactions between Cks1 and the mitotic entry checkpoint genes require the inhibitory activity of Swe1 on the yeast cyclin-dependent kinase (CDK), Cdc28. In addition, the SDL interactions of overexpressed CKS1 with mutations in the mitotic exit network are suppressed by modulating expression of the CDK inhibitor Sic1. Mutation of the polo-like kinase Cdc5, which functions in both the mitotic entry and mitotic exit pathways, is lethal in combination with overexpressed CKS1. Therefore we investigated the effect of targeting the human Cdc5 ortholog, PLK1, in breast cancers with various expression levels of human CKS1B. Growth inhibition by PLK1 knockdown correlates with increased CKS1B expression in published tumor cell data sets, and this correlation was confirmed using shRNAs against PLK1 in tumor cell lines. In addition, we overexpressed CKS1B in multiple cell lines and found increased sensitivity to PLK1 knockdown and PLK1 drug inhibition. Finally, combined inhibition of WEE1 and PLK1 results in less apoptosis than predicted based on an additive model of the individual inhibitors, showing an epistatic interaction and confirming a prediction of the yeast data. Thus, identification of a yeast SDL interaction uncovers conserved genetic interactions that can affect human cancer cell viability. Tuesday, October 11 2016 08:45:41 AM Taking Exception to Human Eugenics [Letters] Roth, F. P., Wakeley, J. Tuesday, October 11 2016 08:45:41 AM Mutation, Eugenics, and the Boundaries of Science [Letters] Lynch, M.