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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   Thursday, November 10 2016 08:40:34 AM ISSUE HIGHLIGHTS [Issue Highlights] Thursday, November 10 2016 08:40:34 AM Charlesworth et al. on Background Selection and Neutral Diversity [Classics] Wright, S. I. Thursday, November 10 2016 08:40:34 AM Alfred Sturtevant Walks into a Bar: Gene Dosage, Gene Position, and Unequal Crossing Over in Drosophila [Classics] Wolfner, M. F., Miller, D. E. Thursday, November 10 2016 08:40:34 AM Selfish DNA and Epigenetic Repression Revisited [Commentary] Gasser, S. M. Thursday, November 10 2016 08:40:34 AM Why I Love Genetics: Essay on Occasion of Being Awarded the GSA Medal 2016 [The 2016 GSA Honors and Awards] Weigel, D. Thursday, November 10 2016 08:40:34 AM Susan Celniker: Foundational Resources To Study a Dynamic Genome [The 2016 GSA Honors and Awards] Celniker, S. Thursday, November 10 2016 08:40:34 AM The Genetics of Axon Guidance and Axon Regeneration in Caenorhabditis elegans [Neurobiology and Behavior] Chisholm, A. D., Hutter, H., Jin, Y., Wadsworth, W. G. The correct wiring of neuronal circuits depends on outgrowth and guidance of neuronal processes during development. In the past two decades, great progress has been made in understanding the molecular basis of axon outgrowth and guidance. Genetic analysis in Caenorhabditis elegans has played a key role in elucidating conserved pathways regulating axon guidance, including Netrin signaling, the slit Slit/Robo pathway, Wnt signaling, and others. Axon guidance factors were first identified by screens for mutations affecting animal behavior, and by direct visual screens for axon guidance defects. Genetic analysis of these pathways has revealed the complex and combinatorial nature of guidance cues, and has delineated how cues guide growth cones via receptor activity and cytoskeletal rearrangement. Several axon guidance pathways also affect directed migrations of non-neuronal cells in C. elegans, with implications for normal and pathological cell migrations in situations such as tumor metastasis. The small number of neurons and highly stereotyped axonal architecture of the C. elegans nervous system allow analysis of axon guidance at the level of single identified axons, and permit in vivo tests of prevailing models of axon guidance. C. elegans axons also have a robust capacity to undergo regenerative regrowth after precise laser injury (axotomy). Although such axon regrowth shares some similarities with developmental axon outgrowth, screens for regrowth mutants have revealed regeneration-specific pathways and factors that were not identified in developmental screens. Several areas remain poorly understood, including how major axon tracts are formed in the embryo, and the function of axon regeneration in the natural environment. Thursday, November 10 2016 08:40:34 AM CRISPR Technology Reveals RAD(51)-ical Mechanisms of Repair in Roundworms: An Educational Primer for Use with "Promotion of Homologous Recombination by SWS-1 in Complex with RAD-51 Paralogs in Caenorhabditis elegans" [Primer] Turcotte, C. A., Andrews, N. P., Sloat, S. A., Checchi, P. M. The mechanisms cells use to maintain genetic fidelity via DNA repair and the accuracy of these processes have garnered interest from scientists engaged in basic research to clinicians seeking improved treatment for cancer patients. Despite the continued advances, many details of DNA repair are still incompletely understood. In addition, the inherent complexity of DNA repair processes, even at the most fundamental level, makes it a challenging topic. This primer is meant to assist both educators and students in using a recent paper, "Promotion of homologous recombination by SWS-1 in complex with RAD-51 paralogs in Caenorhabditis elegans," to understand mechanisms of DNA repair. The goals of this primer are to highlight and clarify several key techniques utilized, with special emphasis on the clustered, regularly interspaced, short palindromic repeats technique and the ways in which it has revolutionized genetics research, as well as to provide questions for deeper in-class discussion. Thursday, November 10 2016 08:40:34 AM Heterozygosity Ratio, a Robust Global Genomic Measure of Autozygosity and Its Association with Height and Disease Risk [Statistical Genetics and Genomics] Samuels, D. C., Wang, J., Ye, F., He, J., Levinson, R. T., Sheng, Q., Zhao, S., Capra, J. A., Shyr, Y., Zheng, W., Guo, Y. Greater genetic variability in an individual is protective against recessive disease. However, existing quantifications of autozygosity, such as runs of homozygosity (ROH), have proved highly sensitive to genotyping density and have yielded inconclusive results about the relationship of diversity and disease risk. Using genotyping data from three data sets with >43,000 subjects, we demonstrated that an alternative approach to quantifying genetic variability, the heterozygosity ratio, is a robust measure of diversity and is positively associated with the nondisease trait height and several disease phenotypes in subjects of European ancestry. The heterozygosity ratio is the number of heterozygous sites in an individual divided by the number of nonreference homozygous sites and is strongly affected by the degree of genetic admixture of the population and varies across human populations. Unlike quantifications of ROH, the heterozygosity ratio is not sensitive to the density of genotyping performed. Our results establish the heterozygosity ratio as a powerful new statistic for exploring the patterns and phenotypic effects of different levels of genetic variation in populations. Thursday, November 10 2016 08:40:34 AM Different Mechanisms of Longevity in Long-Lived Mouse and Caenorhabditis elegans Mutants Revealed by Statistical Analysis of Mortality Rates [Statistical Genetics and Genomics] Hughes, B. G., Hekimi, S. Mouse and Caenorhabditis elegans mutants with altered life spans are being used to investigate the aging process and how genes determine life span. The survival of a population can be modeled by the Gompertz function, which comprises two parameters. One of these parameters ("G") describes the rate at which mortality accelerates with age and is often described as the "rate of aging." The other parameter ("A") may correspond to the organism’s baseline vulnerability to deleterious effects of disease and the environment. We show that, in mice, life-span-extending mutations systematically fail to affect the age-dependent acceleration of mortality (G), but instead affect only baseline vulnerability (A). This remains true even when comparing strains maintained under identical environmental conditions. In contrast, life-span-extending mutations in C. elegans were associated with decreases in G. These observations on mortality rate kinetics suggest that the mechanisms of aging in mammals might fundamentally differ from those in nematodes. Thursday, November 10 2016 08:40:34 AM Boosting Gene Mapping Power and Efficiency with Efficient Exact Variance Component Tests of Single Nucleotide Polymorphism Sets [Statistical Genetics and Genomics] Zhou, J. J., Hu, T., Qiao, D., Cho, M. H., Zhou, H. Single nucleotide polymorphism (SNP) set tests have been a powerful method in analyzing next-generation sequencing (NGS) data. The popular sequence kernel association test (SKAT) method tests a set of variants as random effects in the linear mixed model setting. Its P-value is calculated based on asymptotic theory that requires a large sample size. Therefore, it is known that SKAT is conservative and can lose power at small or moderate sample sizes. Given the current cost of sequencing technology, scales of NGS are still limited. In this report, we derive and implement computationally efficient, exact (nonasymptotic) score (eScore), likelihood ratio (eLRT), and restricted likelihood ratio (eRLRT) tests, ExactVCTest, that can achieve high power even when sample sizes are small. We perform simulation studies under various genetic scenarios. Our ExactVCTest (i.e., eScore, eLRT, eRLRT) exhibits well-controlled type I error. Under the alternative model, eScore P-values are universally smaller than those from SKAT. eLRT and eRLRT demonstrate significantly higher power than eScore, SKAT, and SKAT optimal (SKAT-o) across all scenarios and various samples sizes. We applied these tests to an exome sequencing study. Our findings replicate previous results and shed light on rare variant effects within genes. The software package is implemented in the open source, high-performance technical computing language Julia, and is freely available at https://github.com/Tao-Hu/VarianceComponentTest.jl. Analysis of each trait in the exome sequencing data set with 399 individuals and 16,619 genes takes around 1 min on a desktop computer. Thursday, November 10 2016 08:40:34 AM Incorporating Functional Annotations for Fine-Mapping Causal Variants in a Bayesian Framework Using Summary Statistics [Statistical Genetics and Genomics] Chen, W., McDonnell, S. K., Thibodeau, S. N., Tillmans, L. S., Schaid, D. J. Functional annotations have been shown to improve both the discovery power and fine-mapping accuracy in genome-wide association studies. However, the optimal strategy to incorporate the large number of existing annotations is still not clear. In this study, we propose a Bayesian framework to incorporate functional annotations in a systematic manner. We compute the maximum a posteriori solution and use cross validation to find the optimal penalty parameters. By extending our previous fine-mapping method CAVIARBF into this framework, we require only summary statistics as input. We also derived an exact calculation of Bayes factors using summary statistics for quantitative traits, which is necessary when a large proportion of trait variance is explained by the variants of interest, such as in fine mapping expression quantitative trait loci (eQTL). We compared the proposed method with PAINTOR using different strategies to combine annotations. Simulation results show that the proposed method achieves the best accuracy in identifying causal variants among the different strategies and methods compared. We also find that for annotations with moderate effects from a large annotation pool, screening annotations individually and then combining the top annotations can produce overly optimistic results. We applied these methods on two real data sets: a meta-analysis result of lipid traits and a cis-eQTL study of normal prostate tissues. For the eQTL data, incorporating annotations significantly increased the number of potential causal variants with high probabilities. Thursday, November 10 2016 08:40:34 AM Hydroxyurea Induces Cytokinesis Arrest in Cells Expressing a Mutated Sterol-14{alpha}-Demethylase in the Ergosterol Biosynthesis Pathway [Genome Integrity and Transmission] Xu, Y.-j., Singh, A., Alter, G. M. Hydroxyurea (HU) has been used for the treatment of multiple diseases, such as cancer. The therapeutic effect is generally believed to be due to the suppression of ribonucleotide reductase (RNR), which slows DNA polymerase movement at replication forks and induces an S phase cell cycle arrest in proliferating cells. Although aberrant mitosis and DNA damage generated at collapsed forks are the likely causes of cell death in the mutants with defects in replication stress response, the mechanism underlying the cytotoxicity of HU in wild-type cells remains poorly understood. While screening for new fission yeast mutants that are sensitive to replication stress, we identified a novel mutation in the erg11 gene encoding the enzyme sterol-14α-demethylase in the ergosterol biosynthesis pathway that dramatically sensitizes the cells to chronic HU treatment. Surprisingly, HU mainly arrests the erg11 mutant cells in cytokinesis, not in S phase. Unlike the reversible S phase arrest in wild-type cells, the cytokinesis arrest induced by HU is relatively stable and occurs at low doses of the drug, which likely explains the remarkable sensitivity of the mutant to HU. We also show that the mutation causes sterol deficiency, which may predispose the cells to the cytokinesis arrest and lead to cell death. We hypothesize that in addition to the RNR, HU may have a secondary unknown target(s) inside cells. Identification of such a target(s) may greatly improve the chemotherapies that employ HU or help to expand the clinical usage of this drug for additional pathological conditions. Thursday, November 10 2016 08:40:34 AM Dosage Mutator Genes in Saccharomyces cerevisiae: A Novel Mutator Mode-of-Action of the Mph1 DNA Helicase [Genome Integrity and Transmission] Ang, J. S., Duffy, S., Segovia, R., Stirling, P. C., Hieter, P. Mutations that cause genome instability are considered important predisposing events that contribute to initiation and progression of cancer. Genome instability arises either due to defects in genes that cause an increased mutation rate (mutator phenotype), or defects in genes that cause chromosome instability (CIN). To extend the catalog of genome instability genes, we systematically explored the effects of gene overexpression on mutation rate, using a forward-mutation screen in budding yeast. We screened ~5100 plasmids, each overexpressing a unique single gene, and characterized the five strongest mutators, MPH1 (mutator phenotype 1), RRM3, UBP12, PIF1, and DNA2. We show that, for MPH1, the yeast homolog of Fanconi Anemia complementation group M (FANCM), the overexpression mutator phenotype is distinct from that of mph1. Moreover, while four of our top hits encode DNA helicases, the overexpression of 48 other DNA helicases did not cause a mutator phenotype, suggesting this is not a general property of helicases. For Mph1 overexpression, helicase activity was not required for the mutator phenotype; in contrast Mph1 DEAH-box function was required for hypermutation. Mutagenesis by MPH1 overexpression was independent of translesion synthesis (TLS), but was suppressed by overexpression of RAD27, a conserved flap endonuclease. We propose that binding of DNA flap structures by excess Mph1 may block Rad27 action, creating a mutator phenotype that phenocopies rad27. We believe this represents a novel mutator mode-of-action and opens up new prospects to understand how upregulation of DNA repair proteins may contribute to mutagenesis. Thursday, November 10 2016 08:40:34 AM Synaptonemal Complex Components Are Required for Meiotic Checkpoint Function in Caenorhabditis elegans [Communications] Bohr, T., Ashley, G., Eggleston, E., Firestone, K., Bhalla, N. Synapsis involves the assembly of a proteinaceous structure, the synaptonemal complex (SC), between paired homologous chromosomes, and is essential for proper meiotic chromosome segregation. In Caenorhabditis elegans, the synapsis checkpoint selectively removes nuclei with unsynapsed chromosomes by inducing apoptosis. This checkpoint depends on pairing centers (PCs), cis-acting sites that promote pairing and synapsis. We have hypothesized that the stability of homolog pairing at PCs is monitored by this checkpoint. Here, we report that SC components SYP-3, HTP-3, HIM-3, and HTP-1 are required for a functional synapsis checkpoint. Mutation of these components does not abolish PC function, demonstrating they are bona fide checkpoint components. Further, we identify mutant backgrounds in which the instability of homolog pairing at PCs does not correlate with the synapsis checkpoint response. Altogether, these data suggest that, in addition to homolog pairing, SC assembly may be monitored by the synapsis checkpoint. Thursday, November 10 2016 08:40:34 AM To Break or Not To Break: Sex Chromosome Hemizygosity During Meiosis in Caenorhabditis [Genetics of Sex] Van, M. V., Larson, B. J., Engebrecht, J. Meiotic recombination establishes connections between homologous chromosomes to promote segregation. Hemizygous regions of sex chromosomes have no homologous chromosome to recombine with, yet must be transmitted through meiosis. An extreme case of hemizygosity exists in the genus Caenorhabditis, where males have a single X chromosome that completely lacks a homologous partner. To determine whether similar strategies have evolved to accommodate hemizygosity of the X during male meiosis in Caenorhabditis with distinct modes of sexual reproduction, we examined induction and processing of meiotic double strand breaks (DSBs) in androdioecious (hermaphrodite/male) Caenorhabditis elegans and C. briggsae, and gonochoristic (female/male) C. remanei and C. brenneri. Analysis of the recombinase RAD-51 suggests more meiotic DSBs are induced in gonochoristic vs. androdioecious species. However, in late prophase in all species, chromosome pairs are restructured into bivalents around a single axis, suggesting that the holocentric nature of Caenorhabditis chromosomes dictates a single crossover per bivalent regardless of the number of DSBs induced. Interestingly, RAD-51 foci were readily observed on the X chromosome of androdioecious male germ cells, while very few were detected in gonochoristic male germ cells. As in C. elegans, the X chromosome in C. briggsae male germ cells undergoes transient pseudosynapsis and flexibility in DSB repair pathway choice. In contrast, in C. remanei and C. brenneri male germ cells, the X chromosome does not undergo pseudosynapsis and appears refractory to SPO-11-induced breaks. Together our results suggest that distinct strategies have evolved to accommodate sex chromosome hemizygosity during meiosis in closely related Caenorhabditis species. Thursday, November 10 2016 08:40:34 AM Extraction of the Constituent Subgenomes of the Natural Allopolyploid Rapeseed (Brassica napus L.) [Genome Integrity and Transmission] Zhu, B., Tu, Y., Zeng, P., Ge, X., Li, Z. As the dynamic nature of progenitor genomes accompanies the speciation by interspecific hybridization, the extraction of the constituent subgenome(s) from a natural allopolyploid species of long history and then restitution of the progenitor(s) provides the unique opportunity to study the genome evolution and interplay. Herein, the A subgenome from the allotetraploid oilseed rape (Brassica napus L., AACC) was extracted through inducing the preferential elimination of C-subgenome chromosomes in intertribal crosses and the progenitor B. rapa was restituted (RBR). Then by crossing and backcrossing RBR with B. napus donor, the C subgenome was in situ dissected by adding each of its nine chromosomes to the extracted A subgenome and establishing the whole set of monosonic alien addition lines (MAALs). RBR from spring-type B. napus genotype "Oro" expressed a phenotype resembling some type of B. rapa never observed before, but showed a winter-type flowering habit. This RBR had weaker growth vigor and suffered more seriously from biotic and abiotic stresses compared with Oro. The phenotypes specific for these MAALs showed the location of the related genes on the particular C-subgenome chromosomes. These MAALs exhibited obviously different frequencies in homeologous pairing and transmission of additional C-subgenome chromosomes, which were associated with the distinct degrees of their relatedness, and even with the possible genetic regulation for meiotic pairing evolved in B. napus. Finally, large scaffolds undetermined for sequence assembly of B. napus were anchored to specific C-subgenome chromosomes using MAALs. Thursday, November 10 2016 08:40:34 AM Identification of Tension Sensing Motif of Histone H3 in Saccharomyces cerevisiae and Its Regulation by Histone Modifying Enzymes [Genome Integrity and Transmission] Luo, J., Deng, X., Buehl, C., Xu, X., Kuo, M.-H. To ensure genome stability during cell division, all chromosomes must attach to spindles emanating from the opposite spindle pole bodies before segregation. The tension between sister chromatids generated by the poleward pulling force is an integral part of chromosome biorientation. In budding yeast, the residue Gly44 of histone H3 is critical for retaining the conserved Shugoshin protein Sgo1p at the pericentromeres for monitoring the tension status during mitosis. Studies carried out in this work showed that Lys42, Gly44, and Thr45 of H3 form the core of a tension sensing motif (TSM). Similar to the previously reported G44S mutant, K42A, G44A, and T45A alleles all rendered cells unable to respond to erroneous spindle attachment, a phenotype suppressed by Sgo1p overexpression. TSM functions by physically recruiting or retaining Sgo1p at pericentromeres as evidenced by chromatin immunoprecipitation and by in vitro pulldown experiments. Intriguingly, the function of TSM is likely regulated by multiple histone modifying enzymes, including the histone acetyltransferase Gcn5p, and deacetylases Rpd3p and Hos2p. Defects caused by TSM mutations can be suppressed by the expression of a catalytically inactive mutant of Gcn5p. Conversely, G44S mutant cells exhibit prominent chromatin instability phenotype in the absence of RPD3. Importantly, the gcn5– suppressor restores the tension sensing function in tsm– background in a fashion that bypasses the need of stably associating Sgo1p with chromatin. These results demonstrate that the TSM of histone H3 is a key component of a mechanism that ensures faithful segregation, and that interaction with chromatin modifying enzymes may be an important part of the mitotic quality control process. Thursday, November 10 2016 08:40:34 AM A Multigenic Network of ARGONAUTE4 Clade Members Controls Early Megaspore Formation in Arabidopsis [Gene Expression] Hernandez-Lagana, E., Rodriguez-Leal, D., Lua, J., Vielle-Calzada, J.-P. The development of gametophytes relies on the establishment of a haploid gametophytic generation that initiates with the specification of gametophytic precursors. The majority of flowering plants differentiate a single gametophytic precursor in the ovule: the megaspore mother cell. Here we show that, in addition to argonaute9 (ago9), mutations in other ARGONAUTE (AGO) genes such as ago4, ago6, and ago8, also show abnormal configurations containing supernumerary gametophytic precursors in Arabidopsis thaliana. Double homozygous ago4 ago9 individuals showed a suppressive effect on the frequency of ovules with multiple gametophytic precursors across three consecutive generations, indicating that genetic interactions result in compensatory mechanisms. Whereas overexpression of AGO6 in ago9 and ago4 ago9 confirms strong regulatory interactions among genes involved in RNA-directed DNA methylation, AGO8 is overexpressed in premeiotic ovules of ago4 ago9 individuals, suggesting that the regulation of this previously presumed pseudogene responds to the compensatory mechanism. The frequency of abnormal meiotic configurations found in ago4 ago9 individuals is dependent on their parental genotype, revealing a transgenerational effect. Our results indicate that members of the AGO4 clade cooperatively participate in preventing the abnormal specification of multiple premeiotic gametophytic precursors during early ovule development in A. thaliana. Thursday, November 10 2016 08:40:34 AM Discovering Single Nucleotide Polymorphisms Regulating Human Gene Expression Using Allele Specific Expression from RNA-seq Data [Gene Expression] Kang, E. Y., Martin, L. J., Mangul, S., Isvilanonda, W., Zou, J., Ben-David, E., Han, B., Lusis, A. J., Shifman, S., Eskin, E. The study of the genetics of gene expression is of considerable importance to understanding the nature of common, complex diseases. The most widely applied approach to identifying relationships between genetic variation and gene expression is the expression quantitative trait loci (eQTL) approach. Here, we increased the computational power of eQTL with an alternative and complementary approach based on analyzing allele specific expression (ASE). We designed a novel analytical method to identify cis-acting regulatory variants based on genome sequencing and measurements of ASE from RNA-sequencing (RNA-seq) data. We evaluated the power and resolution of our method using simulated data. We then applied the method to map regulatory variants affecting gene expression in lymphoblastoid cell lines (LCLs) from 77 unrelated northern and western European individuals (CEU), which were part of the HapMap project. A total of 2309 SNPs were identified as being associated with ASE patterns. The SNPs associated with ASE were enriched within promoter regions and were significantly more likely to signal strong evidence for a regulatory role. Finally, among the candidate regulatory SNPs, we identified 108 SNPs that were previously associated with human immune diseases. With further improvements in quantifying ASE from RNA-seq, the application of our method to other datasets is expected to accelerate our understanding of the biological basis of common diseases. Thursday, November 10 2016 08:40:34 AM Donor Preference Meets Heterochromatin: Moonlighting Activities of a Recombinational Enhancer in Saccharomyces cerevisiae [Gene Expression] Dodson, A. E., Rine, J. In Saccharomyces cerevisiae, a small, intergenic region known as the recombination enhancer regulates donor selection during mating-type switching and also helps shape the conformation of chromosome III. Using an assay that detects transient losses of heterochromatic repression, we found that the recombination enhancer also acts at a distance in cis to modify the stability of gene silencing. In a mating-type-specific manner, the recombination enhancer destabilized the heterochromatic repression of a gene located ~17 kbp away. This effect depended on a subregion of the recombination enhancer that is largely sufficient to determine donor preference. Therefore, this subregion affects both recombination and transcription from a distance. These observations identify a rare example of long-range transcriptional regulation in yeast and raise the question of whether other cis elements also mediate dual effects on recombination and gene expression. Thursday, November 10 2016 08:40:34 AM A Common Suite of Coagulation Proteins Function in Drosophila Muscle Attachment [Cellular Genetics] Green, N., Odell, N., Zych, M., Clark, C., Wang, Z.-H., Biersmith, B., Bajzek, C., Cook, K. R., Dushay, M. S., Geisbrecht, E. R. The organization and stability of higher order structures that form in the extracellular matrix (ECM) to mediate the attachment of muscles are poorly understood. We have made the surprising discovery that a subset of clotting factor proteins are also essential for muscle attachment in the model organism Drosophila melanogaster. One such coagulation protein, Fondue (Fon), was identified as a novel muscle mutant in a pupal lethal genetic screen. Fon accumulates at muscle attachment sites and removal of this protein results in decreased locomotor behavior and detached larval muscles. A sensitized genetic background assay reveals that fon functions with the known muscle attachment genes Thrombospondin (Tsp) and Tiggrin (Tig). Interestingly, Tig is also a component of the hemolymph clot. We further demonstrate that an additional clotting protein, Larval serum protein 1 (Lsp1), is also required for muscle attachment stability and accumulates where muscles attach to tendons. While the local biomechanical and organizational properties of the ECM vary greatly depending on the tissue microenvironment, we propose that shared extracellular protein–protein interactions influence the strength and elasticity of ECM proteins in both coagulation and muscle attachment. Thursday, November 10 2016 08:40:34 AM The Roles of {beta}-Oxidation and Cofactor Homeostasis in Peroxisome Distribution and Function in Arabidopsis thaliana [Cellular Genetics] Rinaldi, M. A., Patel, A. B., Park, J., Lee, K., Strader, L. C., Bartel, B. Key steps of essential metabolic pathways are housed in plant peroxisomes. We conducted a microscopy-based screen for anomalous distribution of peroxisomally targeted fluorescence in Arabidopsis thaliana. This screen uncovered 34 novel alleles in 15 genes affecting oil body mobilization, fatty acid β-oxidation, the glyoxylate cycle, peroxisome fission, and pexophagy. Partial loss-of-function of lipid-mobilization enzymes conferred peroxisomes clustered around retained oil bodies without other notable defects, suggesting that this microscopy-based approach was sensitive to minor perturbations, and that fatty acid β-oxidation rates in wild type are higher than required for normal growth. We recovered three mutants defective in PECTIN METHYLESTERASE31, revealing an unanticipated role in lipid mobilization for this cytosolic enzyme. Whereas mutations reducing fatty acid import had peroxisomes of wild-type size, mutations impairing fatty acid β-oxidation displayed enlarged peroxisomes, possibly caused by excess fatty acid β-oxidation intermediates in the peroxisome. Several fatty acid β-oxidation mutants also displayed defects in peroxisomal matrix protein import. Impairing fatty acid import reduced the large size of peroxisomes in a mutant defective in the PEROXISOMAL NAD+ TRANSPORTER (PXN), supporting the hypothesis that fatty acid accumulation causes pxn peroxisome enlargement. The diverse mutants isolated in this screen will aid future investigations of the roles of β-oxidation and peroxisomal cofactor homeostasis in plant development. Thursday, November 10 2016 08:40:34 AM Trs33-Containing TRAPP IV: A Novel Autophagy-Specific Ypt1 GEF [Cellular Genetics] Lipatova, Z., Majumdar, U., Segev, N. Thursday, November 10 2016 08:40:34 AM Working-for-Food Behaviors: A Preclinical Study in Prader-Willi Mutant Mice [Developmental and Behavioral Genetics] Lassi, G., Maggi, S., Balzani, E., Cosentini, I., Garcia-Garcia, C., Tucci, V. Abnormal feeding behavior is one of the main symptoms of Prader-Willi syndrome (PWS). By studying a PWS mouse mutant line, which carries a paternally inherited deletion of the small nucleolar RNA 116 (Snord116), we observed significant changes in working-for-food behavioral responses at various timescales. In particular, we report that PWS mutant mice show a significant delay compared to wild-type littermate controls in responding to both hour-scale and seconds-to-minutes-scale time intervals. This timing shift in mutant mice is associated with better performance in the working-for-food task, and results in better decision making in these mutant mice. The results of our study reveal a novel aspect of the organization of feeding behavior, and advance the understanding of the interplay between the metabolic functions and cognitive mechanisms of PWS. Thursday, November 10 2016 08:40:34 AM Establishment of the Muscle-Tendon Junction During Thorax Morphogenesis in Drosophila Requires the Rho-Kinase [Developmental and Behavioral Genetics] Vega-Macaya, F., Manieu, C., Valdivia, M., Mlodzik, M., Olguin, P. The assembly of the musculoskeletal system in Drosophila relies on the integration of chemical and mechanical signaling between the developing muscles with ectodermal cells specialized as "tendon cells." Mechanical tension generated at the junction of flight muscles and tendon cells of the notum epithelium is required for muscle morphogenesis, and is balanced by the epithelium in order to not deform. We report that Drosophila Rho kinase (DRok) is necessary in tendon cells to assemble stable myotendinous junctions (MTJ), which are required for muscle morphogenesis and survival. In addition, DRok is required in tendon cells to maintain epithelial shape and cell orientation in the notum, independently of chascon (chas). Loss of DRok function in tendon cells results in mis-orientation of tendon cell extensions and abnormal accumulation of Thrombospondin and βPS-integrin, which may cause abnormal myotendinous junction formation and muscle morphogenesis. This role does not depend exclusively on nonmuscular Myosin-II activation (Myo-II), indicating that other DRok targets are key in this process. We propose that DRok function in tendon cells is key to promote the establishment of MTJ attachment and to balance mechanical tension generated at the MTJ by muscle compaction. Thursday, November 10 2016 08:40:34 AM The Neuropeptides FLP-2 and PDF-1 Act in Concert To Arouse Caenorhabditis elegans Locomotion [Developmental and Behavioral Genetics] Chen, D., Taylor, K. P., Hall, Q., Kaplan, J. M. During larval molts, Caenorhabditis elegans exhibits a sleep-like state (termed lethargus) that is characterized by the absence of feeding and profound locomotion quiescence. The rhythmic pattern of locomotion quiescence and arousal linked to the molting cycle is mediated by reciprocal changes in sensory responsiveness, whereby arousal is associated with increased responsiveness. Sensory neurons arouse locomotion via release of a neuropeptide (PDF-1) and glutamate. Here we identify a second arousing neuropeptide (FLP-2). We show that FLP-2 acts via an orexin-like receptor (FRPR-18), and that FLP-2 and PDF-1 secretion are regulated by reciprocal positive feedback. These results suggest that the aroused behavioral state is stabilized by positive feedback between two neuropeptides. Thursday, November 10 2016 08:40:34 AM A Plastic Vegetative Growth Threshold Governs Reproductive Capacity in Aspergillus nidulans [Developmental and Behavioral Genetics] Noble, L. M., Holland, L. M., McLauchlan, A. J., Andrianopoulos, A. Ontogenetic phases separating growth from reproduction are a common feature of cellular life. Long recognized for flowering plants and animals, early literature suggests this life-history component may also be prevalent among multicellular fungi. We establish the basis of developmental competence—the capacity to respond to induction of asexual development—in the filamentous saprotroph Aspergillus nidulans, describing environmental influences, including genotype-by-environment interactions among precocious mutants, gene expression associated with wild type and precocious competence acquisition, and the genetics of competence timing. Environmental effects are consistent with a threshold driven by metabolic rate and organism density, with pH playing a particularly strong role in determining competence timing. Gene expression diverges significantly over the competence window, despite a lack of overt morphological change, with differentiation in key metabolic, signaling, and cell trafficking processes. We identify five genes for which mutant alleles advance competence timing, including the conserved GTPase RasB (AN5832) and ambient pH sensor PalH (AN6886). In all cases examined, inheritance of competence timing is complex and non-Mendelian, with F1 progeny showing highly variable transgressive timing and dominant parental effects with a weak contribution from progeny genotype. Competence provides a new model for nutrient-limited life-cycle phases, and their elaboration from unicellular origins. Further work is required to establish the hormonal and bioenergetic basis of the trait across fungi, and underlying mechanisms of variable inheritance. Thursday, November 10 2016 08:40:34 AM Mitotic Spindle Positioning in the EMS Cell of Caenorhabditis elegans Requires LET-99 and LIN-5/NuMA [Developmental and Behavioral Genetics] Liro, M. J., Rose, L. S. Asymmetric divisions produce daughter cells with different fates, and thus are critical for animal development. During asymmetric divisions, the mitotic spindle must be positioned on a polarized axis to ensure the differential segregation of cell fate determinants into the daughter cells. In many cell types, a cortically localized complex consisting of Gα, GPR-1/2, and LIN-5 (Gαi/Pins/Mud, Gαi/LGN/NuMA) mediates the recruitment of dynactin/dynein, which exerts pulling forces on astral microtubules to physically position the spindle. The conserved PAR polarity proteins are known to regulate both cytoplasmic asymmetry and spindle positioning in many cases. However, spindle positioning also occurs in response to cell signaling cues that appear to be PAR-independent. In the four-cell Caenorhabditis elegans embryo, Wnt and Mes-1/Src-1 signaling pathways act partially redundantly to align the spindle on the anterior/posterior axis of the endomesodermal (EMS) precursor cell. It is unclear how those extrinsic signals individually contribute to spindle positioning and whether either pathway acts via conserved spindle positioning regulators. Here, we genetically test the involvement of Gα, LIN-5, and their negative regulator LET-99, in transducing EMS spindle positioning polarity cues. We also examined whether the C. elegans ortholog of another spindle positioning regulator, DLG-1, is required. We show that LET-99 acts in the Mes-1/Src-1 pathway for spindle positioning. LIN-5 is also required for EMS spindle positioning, possibly through a Gα- and DLG-1-independent mechanism. Thursday, November 10 2016 08:40:34 AM Inferring Past Effective Population Size from Distributions of Coalescent Times [Population and Evolutionary Genetics] Gattepaille, L., Gunther, T., Jakobsson, M. Inferring and understanding changes in effective population size over time is a major challenge for population genetics. Here we investigate some theoretical properties of random-mating populations with varying size over time. In particular, we present an exact solution to compute the population size as a function of time, $${\mathit{N}}_{\mathrm{e}}\left(\mathit{t}\right)$$, based on distributions of coalescent times of samples of any size. This result reduces the problem of population size inference to a problem of estimating coalescent time distributions. To illustrate the analytic results, we design a heuristic method using a tree-inference algorithm and investigate simulated and empirical population-genetic data. We investigate the effects of a range of conditions associated with empirical data, for instance number of loci, sample size, mutation rate, and cryptic recombination. We show that our approach performs well with genomic data (≥ 10,000 loci) and that increasing the sample size from 2 to 10 greatly improves the inference of $${\mathit{N}}_{\mathrm{e}}\left(\mathit{t}\right)$$ whereas further increase in sample size results in modest improvements, even under a scenario of exponential growth. We also investigate the impact of recombination and characterize the potential biases in inference of $${\mathit{N}}_{\mathrm{e}}\left(\mathit{t}\right)$$. The approach can handle large sample sizes and the computations are fast. We apply our method to human genomes from four populations and reconstruct population size profiles that are coherent with previous finds, including the Out-of-Africa bottleneck. Additionally, we uncover a potential difference in population size between African and non-African populations as early as 400 KYA. In summary, we provide an analytic relationship between distributions of coalescent times and $${\mathit{N}}_{\mathrm{e}}\left(\mathit{t}\right)$$, which can be incorporated into powerful approaches for inferring past population sizes from population-genomic data. Thursday, November 10 2016 08:40:34 AM Effects of Linked Selective Sweeps on Demographic Inference and Model Selection [Population and Evolutionary Genetics] Schrider, D. R., Shanku, A. G., Kern, A. D. The availability of large-scale population genomic sequence data has resulted in an explosion in efforts to infer the demographic histories of natural populations across a broad range of organisms. As demographic events alter coalescent genealogies, they leave detectable signatures in patterns of genetic variation within and between populations. Accordingly, a variety of approaches have been designed to leverage population genetic data to uncover the footprints of demographic change in the genome. The vast majority of these methods make the simplifying assumption that the measures of genetic variation used as their input are unaffected by natural selection. However, natural selection can dramatically skew patterns of variation not only at selected sites, but at linked, neutral loci as well. Here we assess the impact of recent positive selection on demographic inference by characterizing the performance of three popular methods through extensive simulation of data sets with varying numbers of linked selective sweeps. In particular, we examined three different demographic models relevant to a number of species, finding that positive selection can bias parameter estimates of each of these models—often severely. We find that selection can lead to incorrect inferences of population size changes when none have occurred. Moreover, we show that linked selection can lead to incorrect demographic model selection, when multiple demographic scenarios are compared. We argue that natural populations may experience the amount of recent positive selection required to skew inferences. These results suggest that demographic studies conducted in many species to date may have exaggerated the extent and frequency of population size changes. Thursday, November 10 2016 08:40:34 AM The Fitness Effects of Spontaneous Mutations Nearly Unseen by Selection in a Bacterium with Multiple Chromosomes [Population and Evolutionary Genetics] Dillon, M. M., Cooper, V. S. Mutation accumulation (MA) experiments employ the strategy of minimizing the population size of evolving lineages to greatly reduce effects of selection on newly arising mutations. Thus, most mutations fix within MA lines independently of their fitness effects. This approach, more recently combined with genome sequencing, has detailed the rates, spectra, and biases of different mutational processes. However, a quantitative understanding of the fitness effects of mutations virtually unseen by selection has remained an untapped opportunity. Here, we analyzed the fitness of 43 sequenced MA lines of the multi-chromosome bacterium Burkholderia cenocepacia that had each undergone 5554 generations of MA and accumulated an average of 6.73 spontaneous mutations. Most lineages exhibited either neutral or deleterious fitness in three different environments in comparison with their common ancestor. The only mutational class that was significantly overrepresented in lineages with reduced fitness was the loss of the plasmid, though nonsense mutations, missense mutations, and coding insertion-deletions were also overrepresented in MA lineages whose fitness had significantly declined. Although the overall distribution of fitness effects was similar between the three environments, the magnitude and even the sign of the fitness of a number of lineages changed with the environment, demonstrating that the fitness of some genotypes was environmentally dependent. These results present an unprecedented picture of the fitness effects of spontaneous mutations in a bacterium with multiple chromosomes and provide greater quantitative support for the theory that the vast majority of spontaneous mutations are neutral or deleterious. Thursday, November 10 2016 08:40:34 AM Selection on Position of Nonsense Codons in Introns [Population and Evolutionary Genetics] Behringer, M. G., Hall, D. W. Introns occasionally remain in mature messenger RNAs (mRNAs) due to splicing errors and the translated, aberrant proteins that result represent a metabolic cost and may have other deleterious consequences. The nonsense-mediated decay (NMD) pathway degrades aberrant mRNAs, which it recognizes by the presence of an in-frame premature termination codon (PTC). We investigated whether selection has shaped the location of PTCs in introns to reduce waste and facilitate NMD. We found across seven model organisms, that in both first and last introns, PTCs occur earlier in introns than expected by chance, suggesting that selection favors earlier position. This pattern is more pronounced in species with larger effective population sizes. The pattern does not hold for last introns in the two mammal species, however, perhaps because in these species NMD is not initiated from 3'-terminal introns. We conclude that there is compelling evidence that the location of PTCs is shaped by selection for reduced waste and efficient degradation of aberrant mRNAs. Thursday, November 10 2016 08:40:34 AM Evolution of Mutation Rates in Rapidly Adapting Asexual Populations [Population and Evolutionary Genetics] Good, B. H., Desai, M. M. Mutator and antimutator alleles often arise and spread in both natural microbial populations and laboratory evolution experiments. The evolutionary dynamics of these mutation rate modifiers are determined by indirect selection on linked beneficial and deleterious mutations. These indirect selection pressures have been the focus of much earlier theoretical and empirical work, but we still have a limited analytical understanding of how the interplay between hitchhiking and deleterious load influences the fates of modifier alleles. Our understanding is particularly limited when clonal interference is common, which is the regime of primary interest in laboratory microbial evolution experiments. Here, we calculate the fixation probability of a mutator or antimutator allele in a rapidly adapting asexual population, and we show how this quantity depends on the population size, the beneficial and deleterious mutation rates, and the strength of a typical driver mutation. In the absence of deleterious mutations, we find that clonal interference enhances the fixation probability of mutators, even as they provide a diminishing benefit to the overall rate of adaptation. When deleterious mutations are included, natural selection pushes the population toward a stable mutation rate that can be suboptimal for the adaptation of the population as a whole. The approach to this stable mutation rate is not necessarily monotonic: even in the absence of epistasis, selection can favor mutator and antimutator alleles that "overshoot" the stable mutation rate by substantial amounts. Thursday, November 10 2016 08:40:35 AM Joint Prediction of the Effective Population Size and the Rate of Fixation of Deleterious Mutations [Population and Evolutionary Genetics] Santiago, E., Caballero, A. Mutation, genetic drift, and selection are considered the main factors shaping genetic variation in nature. There is a lack, however, of general predictions accounting for the mutual interrelation between these factors. In the context of the background selection model, we provide a set of equations for the joint prediction of the effective population size and the rate of fixation of deleterious mutations, which are applicable both to sexual and asexual species. For a population of N haploid individuals and a model of deleterious mutations with effect s appearing with rate U in a genome L Morgans long, the asymptotic effective population size (Ne) and the average number of generations (T) between consecutive fixations can be approximated by $${\mathit{N}}_{\hbox{ e }}\approx \mathit{N}\hspace{0.17em}\hbox{ exp }\hspace{0.17em}[-2\mathit{U}/(2\mathit{s}\hspace{0.17em}+\mathit{L})\hspace{0.17em}{(1-1/\mathit{UT})}^{3}]$$ and $$\mathit{T}\approx [\hbox{ exp }\left(2\mathit{s}{\mathit{N}}_{\hbox{ e }}\right)\hspace{0.17em}-1]/\left[2\mathit{Us}{\mathit{N}}_{\hbox{ e }}\right]$$. The solution is applicable to Muller’s ratchet, providing satisfactory approximations to the rate of accumulation of mutations for a wide range of parameters. We also obtain predictions of the effective size accounting for the expected nucleotide diversity. Predictions for sexual populations allow for outlining the general conditions where mutational meltdown occurs. The equations can be extended to any distribution of mutational effects and the consideration of hotspots of recombination, showing that Ne is rather insensitive and not proportional to changes in N for many combinations of parameters. This could contribute to explain the observed small differences in levels of polymorphism between species with very different census sizes. Thursday, November 10 2016 08:40:35 AM General Methods for Evolutionary Quantitative Genetic Inference from Generalized Mixed Models [Genetics of Complex Traits] de Villemereuil, P., Schielzeth, H., Nakagawa, S., Morrissey, M. Methods for inference and interpretation of evolutionary quantitative genetic parameters, and for prediction of the response to selection, are best developed for traits with normal distributions. Many traits of evolutionary interest, including many life history and behavioral traits, have inherently nonnormal distributions. The generalized linear mixed model (GLMM) framework has become a widely used tool for estimating quantitative genetic parameters for nonnormal traits. However, whereas GLMMs provide inference on a statistically convenient latent scale, it is often desirable to express quantitative genetic parameters on the scale upon which traits are measured. The parameters of fitted GLMMs, despite being on a latent scale, fully determine all quantities of potential interest on the scale on which traits are expressed. We provide expressions for deriving each of such quantities, including population means, phenotypic (co)variances, variance components including additive genetic (co)variances, and parameters such as heritability. We demonstrate that fixed effects have a strong impact on those parameters and show how to deal with this by averaging or integrating over fixed effects. The expressions require integration of quantities determined by the link function, over distributions of latent values. In general cases, the required integrals must be solved numerically, but efficient methods are available and we provide an implementation in an R package, QGglmm. We show that known formulas for quantities such as heritability of traits with binomial and Poisson distributions are special cases of our expressions. Additionally, we show how fitted GLMM can be incorporated into existing methods for predicting evolutionary trajectories. We demonstrate the accuracy of the resulting method for evolutionary prediction by simulation and apply our approach to data from a wild pedigreed vertebrate population. Thursday, November 10 2016 08:40:35 AM Genetic Mapping by Bulk Segregant Analysis in Drosophila: Experimental Design and Simulation-Based Inference [Genetics of Complex Traits] Pool, J. E. Identifying the genomic regions that underlie complex phenotypic variation is a key challenge in modern biology. Many approaches to quantitative trait locus mapping in animal and plant species suffer from limited power and genomic resolution. Here, I investigate whether bulk segregant analysis (BSA), which has been successfully applied for yeast, may have utility in the genomic era for trait mapping in Drosophila (and other organisms that can be experimentally bred in similar numbers). I perform simulations to investigate the statistical signal of a quantitative trait locus (QTL) in a wide range of BSA and introgression mapping (IM) experiments. BSA consistently provides more accurate mapping signals than IM (in addition to allowing the mapping of multiple traits from the same experimental population). The performance of BSA and IM is maximized by having multiple independent crosses, more generations of interbreeding, larger numbers of breeding individuals, and greater genotyping effort, but is less affected by the proportion of individuals selected for phenotypic extreme pools. I also introduce a prototype analysis method for simulation-based inference for BSA mapping (SIBSAM). This method identifies significant QTL and estimates their genomic confidence intervals and relative effect sizes. Importantly, it also tests whether overlapping peaks should be considered as two distinct QTL. This approach will facilitate improved trait mapping in Drosophila and other species for which hundreds or thousands of offspring (but not millions) can be studied. Thursday, November 10 2016 08:40:35 AM A Variable Genetic Architecture of Melanic Evolution in Drosophila melanogaster [Genetics of Complex Traits] Bastide, H., Lange, J. D., Lack, J. B., Yassin, A., Pool, J. E. Unraveling the genetic architecture of adaptive phenotypic divergence is a fundamental quest in evolutionary biology. In Drosophila melanogaster, high-altitude melanism has evolved in separate mountain ranges in sub-Saharan Africa, potentially as an adaptation to UV intensity. We investigated the genetic basis of this melanism in three populations using a new bulk segregant analysis mapping method. We identified 19 distinct QTL regions from nine mapping crosses, with several QTL peaks overlapping between two or all populations, and yet different crosses involving the same melanic population commonly yielded distinct QTL. The strongest QTL often overlapped well-known pigmentation genes, but we typically did not find wide signals of genetic differentiation (FST) between lightly and darkly pigmented populations at these genes. Instead, we found small numbers of highly differentiated SNPs at the probable causative genes. A simulation analysis showed that these patterns of polymorphism were consistent with selection on standing genetic variation. Overall, our results suggest that, even for potentially simpler traits like pigmentation, the complexity of adaptive trait evolution poses important challenges for QTL mapping and population genetic analysis.