PLoS Genetics

A genetic screen for dihydropyridine (DHP)-resistant worms reveals new residues required for DHP-blockage of mammalian calcium channels.

Publication Date: 2008 May PMID: 18464914
Authors: Kwok, T. C. - Hui, K. - Kostelecki, W. - Ricker, N. - Selman, G. - Feng, Z. P. - Roy, P. J.
Journal: PLoS Genet

Dihydropyridines (DHPs) are L-type calcium channel (Ca(v)1) blockers prescribed to treat several diseases including hypertension. Ca(v)1 channels normally exist in three states: a resting closed state, an open state that is triggered by membrane depolarization, followed by a non-conducting inactivated state that is triggered by the influx of calcium ions, and a rapid change in voltage. DHP binding is thought to alter the conformation of the channel, possibly by engaging a mechanism similar to voltage dependent inactivation, and locking a calcium ion in the pore, thereby blocking channel conductance. As a Ca(v)1 channel crystal structure is lacking, the current model of DHP action has largely been achieved by investigating the role of candidate Ca(v)1 residues in mediating DHP-sensitivity. To better understand DHP-block and identify additional Ca(v)1 residues important for DHP-sensitivity, we screened 440,000 randomly mutated Caenorhabditis elegans genomes for worms resistant to DHP-induced growth defects. We identified 30 missense mutations in the worm Ca(v)1 pore-forming (alpha(1)) subunit, including eleven in conserved residues known to be necessary for DHP-binding. The remaining polymorphisms are in eight conserved residues not previously associated with DHP-sensitivity. Intriguingly, all of the worm mutants that we analyzed phenotypically exhibited increased channel activity. We also created orthologous mutations in the rat alpha(1C) subunit and examined the DHP-block of current through the mutant channels in culture. Six of the seven mutant channels examined either decreased the DHP-sensitivity of the channel and/or exhibited significant residual current at DHP concentrations sufficient to block wild-type channels. Our results further support the idea that DHP-block is intimately associated with voltage dependent inactivation and underscores the utility of C. elegans as a screening tool to identify residues important for DHP interaction with mammalian Ca(v)1 channels.

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A genome-wide association study identifies protein quantitative trait loci (pQTLs).

Publication Date: 2008 May PMID: 18464913
Authors: Melzer, D. - Perry, J. R. - Hernandez, D. - Corsi, A. M. - Stevens, K. - Rafferty, I. - Lauretani, F. - Murray, A. - Gibbs, J. R. - Paolisso, G. - Rafiq, S. - Simon-Sanchez, J. - Lango, H. - Scholz, S. - Weedon, M. N. - Arepalli, S. - Rice, N. - Washecka, N. - Hurst, A. - Britton, A. - Henley, W. - van de Leemput, J. - Li, R. - Newman, A. B. - Tranah, G. - Harris, T. - Panicker, V. - Dayan, C. - Bennett, A. - McCarthy, M. I. - Ruokonen, A. - Jarvelin, M. R. - Guralnik, J. - Bandinelli, S. - Frayling, T. M. - Singleton, A. - Ferrucci, L.
Journal: PLoS Genet

There is considerable evidence that human genetic variation influences gene expression. Genome-wide studies have revealed that mRNA levels are associated with genetic variation in or close to the gene coding for those mRNA transcripts - cis effects, and elsewhere in the genome - trans effects. The role of genetic variation in determining protein levels has not been systematically assessed. Using a genome-wide association approach we show that common genetic variation influences levels of clinically relevant proteins in human serum and plasma. We evaluated the role of 496,032 polymorphisms on levels of 42 proteins measured in 1200 fasting individuals from the population based InCHIANTI study. Proteins included insulin, several interleukins, adipokines, chemokines, and liver function markers that are implicated in many common diseases including metabolic, inflammatory, and infectious conditions. We identified eight Cis effects, including variants in or near the IL6R (p = 1.8x10(-57)), CCL4L1 (p = 3.9x10(-21)), IL18 (p = 6.8x10(-13)), LPA (p = 4.4x10(-10)), GGT1 (p = 1.5x10(-7)), SHBG (p = 3.1x10(-7)), CRP (p = 6.4x10(-6)) and IL1RN (p = 7.3x10(-6)) genes, all associated with their respective protein products with effect sizes ranging from 0.19 to 0.69 standard deviations per allele. Mechanisms implicated include altered rates of cleavage of bound to unbound soluble receptor (IL6R), altered secretion rates of different sized proteins (LPA), variation in gene copy number (CCL4L1) and altered transcription (GGT1). We identified one novel trans effect that was an association between ABO blood group and tumour necrosis factor alpha (TNF-alpha) levels (p = 6.8x10(-40)), but this finding was not present when TNF-alpha was measured using a different assay , or in a second study, suggesting an assay-specific association. Our results show that protein levels share some of the features of the genetics of gene expression. These include the presence of strong genetic effects in cis locations. The identification of protein quantitative trait loci (pQTLs) may be a powerful complementary method of improving our understanding of disease pathways.

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Gene set enrichment in eQTL data identifies novel annotations and pathway regulators.

Publication Date: 2008 May PMID: 18464898
Authors: Wu, C. - Delano, D. L. - Mitro, N. - Su, S. V. - Janes, J. - McClurg, P. - Batalov, S. - Welch, G. L. - Zhang, J. - Orth, A. P. - Walker, J. R. - Glynne, R. J. - Cooke, M. P. - Takahashi, J. S. - Shimomura, K. - Kohsaka, A. - Bass, J. - Saez, E. - Wiltshire, T. - Su, A. I.
Journal: PLoS Genet

Genome-wide gene expression profiling has been extensively used to generate biological hypotheses based on differential expression. Recently, many studies have used microarrays to measure gene expression levels across genetic mapping populations. These gene expression phenotypes have been used for genome-wide association analyses, an analysis referred to as expression QTL (eQTL) mapping. Here, eQTL analysis was performed in adipose tissue from 28 inbred strains of mice. We focused our analysis on "trans-eQTL bands", defined as instances in which the expression patterns of many genes were all associated to a common genetic locus. Genes comprising trans-eQTL bands were screened for enrichments in functional gene sets representing known biological pathways, and genes located at associated trans-eQTL band loci were considered candidate transcriptional modulators. We demonstrate that these patterns were enriched for previously characterized relationships between known upstream transcriptional regulators and their downstream target genes. Moreover, we used this strategy to identify both novel regulators and novel members of known pathways. Finally, based on a putative regulatory relationship identified in our analysis, we identified and validated a previously uncharacterized role for cyclin H in the regulation of oxidative phosphorylation. We believe that the specific molecular hypotheses generated in this study will reveal many additional pathway members and regulators, and that the analysis approaches described herein will be broadly applicable to other eQTL data sets.

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An inducible and reversible mouse genetic rescue system.

Publication Date: 2008 May PMID: 18464897
Authors: Zeng, H. - Horie, K. - Madisen, L. - Pavlova, M. N. - Gragerova, G. - Rohde, A. D. - Schimpf, B. A. - Liang, Y. - Ojala, E. - Kramer, F. - Roth, P. - Slobodskaya, O. - Dolka, I. - Southon, E. A. - Tessarollo, L. - Bornfeldt, K. E. - Gragerov, A. - Pavlakis, G. N. - Gaitanaris, G. A.
Journal: PLoS Genet

Inducible and reversible regulation of gene expression is a powerful approach for uncovering gene function. We have established a general method to efficiently produce reversible and inducible gene knockout and rescue in mice. In this system, which we named iKO, the target gene can be turned on and off at will by treating the mice with doxycycline. This method combines two genetically modified mouse lines: a) a KO line with a tetracycline-dependent transactivator replacing the endogenous target gene, and b) a line with a tetracycline-inducible cDNA of the target gene inserted into a tightly regulated (TIGRE) genomic locus, which provides for low basal expression and high inducibility. Such a locus occurs infrequently in the genome and we have developed a method to easily introduce genes into the TIGRE site of mouse embryonic stem (ES) cells by recombinase-mediated insertion. Both KO and TIGRE lines have been engineered for high-throughput, large-scale and cost-effective production of iKO mice. As a proof of concept, we have created iKO mice in the apolipoprotein E (ApoE) gene, which allows for sensitive and quantitative phenotypic analyses. The results demonstrated reversible switching of ApoE transcription, plasma cholesterol levels, and atherosclerosis progression and regression. The iKO system shows stringent regulation and is a versatile genetic system that can easily incorporate other techniques and adapt to a wide range of applications.

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The impact of recombination on nucleotide substitutions in the human genome.

Publication Date: 2008 May PMID: 18464896
Authors: Duret, L. - Arndt, P. F.
Journal: PLoS Genet

Unraveling the evolutionary forces responsible for variations of neutral substitution patterns among taxa or along genomes is a major issue for detecting selection within sequences. Mammalian genomes show large-scale regional variations of GC-content (the isochores), but the substitution processes at the origin of this structure are poorly understood. We analyzed the pattern of neutral substitutions in 1 Gb of primate non-coding regions. We show that the GC-content toward which sequences are evolving is strongly negatively correlated to the distance to telomeres and positively correlated to the rate of crossovers (R2 = 47%). This demonstrates that recombination has a major impact on substitution patterns in human, driving the evolution of GC-content. The evolution of GC-content correlates much more strongly with male than with female crossover rate, which rules out selectionist models for the evolution of isochores. This effect of recombination is most probably a consequence of the neutral process of biased gene conversion (BGC) occurring within recombination hotspots. We show that the predictions of this model fit very well with the observed substitution patterns in the human genome. This model notably explains the positive correlation between substitution rate and recombination rate. Theoretical calculations indicate that variations in population size or density in recombination hotspots can have a very strong impact on the evolution of base composition. Furthermore, recombination hotspots can create strong substitution hotspots. This molecular drive affects both coding and non-coding regions. We therefore conclude that along with mutation, selection and drift, BGC is one of the major factors driving genome evolution. Our results also shed light on variations in the rate of crossover relative to non-crossover events, along chromosomes and according to sex, and also on the conservation of hotspot density between human and chimp.

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The lambda red proteins promote efficient recombination between diverged sequences: implications for bacteriophage genome mosaicism.

Publication Date: 2008 PMID: 18451987
Authors: Martinsohn, J. T. - Radman, M. - Petit, M. A.
Journal: PLoS Genet

Genome mosaicism in temperate bacterial viruses (bacteriophages) is so great that it obscures their phylogeny at the genome level. However, the precise molecular processes underlying this mosaicism are unknown. Illegitimate recombination has been proposed, but homeologous recombination could also be at play. To test this, we have measured the efficiency of homeologous recombination between diverged oxa gene pairs inserted into lambda. High yields of recombinants between 22% diverged genes have been obtained when the virus Red Gam pathway was active, and 100 fold less when the host Escherichia coli RecABCD pathway was active. The recombination editing proteins, MutS and UvrD, showed only marginal effects on lambda recombination. Thus, escape from host editing contributes to the high proficiency of virus recombination. Moreover, our bioinformatics study suggests that homeologous recombination between similar lambdoid viruses has created part of their mosaicism. We therefore propose that the remarkable propensity of the lambda-encoded Red and Gam proteins to recombine diverged DNA is effectively contributing to mosaicism, and more generally, that a correlation may exist between virus genome mosaicism and the presence of Red/Gam-like systems.

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SU(VAR)3-7 links heterochromatin and dosage compensation in Drosophila.

Publication Date: 2008 PMID: 18451980
Authors: Spierer, A. - Begeot, F. - Spierer, P. - Delattre, M.
Journal: PLoS Genet

In Drosophila, dosage compensation augments X chromosome-linked transcription in males relative to females. This process is achieved by the Dosage Compensation Complex (DCC), which associates specifically with the male X chromosome. We previously found that the morphology of this chromosome is sensitive to the amounts of the heterochromatin-associated protein SU(VAR)3-7. In this study, we examine the impact of change in levels of SU(VAR)3-7 on dosage compensation. We first demonstrate that the DCC makes the X chromosome a preferential target for heterochromatic markers. In addition, reduced or increased amounts of SU(VAR)3-7 result in redistribution of the DCC proteins MSL1 and MSL2, and of Histone 4 acetylation of lysine 16, indicating that a wild-type dose of SU(VAR)3-7 is required for X-restricted DCC targeting. SU(VAR)3-7 is also involved in the dosage compensated expression of the X-linked white gene. Finally, we show that absence of maternally provided SU(VAR)3-7 renders dosage compensation toxic in males, and that global amounts of heterochromatin affect viability of ectopic MSL2-expressing females. Taken together, these results bring to light a link between heterochromatin and dosage compensation.

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How to perform meaningful estimates of genetic effects.

Publication Date: 2008 PMID: 18451979
Authors: Alvarez-Castro, J. M. - Le Rouzic, A. - Carlborg, O.
Journal: PLoS Genet

Although the genotype-phenotype map plays a central role both in Quantitative and Evolutionary Genetics, the formalization of a completely general and satisfactory model of genetic effects, particularly accounting for epistasis, remains a theoretical challenge. Here, we use a two-locus genetic system in simulated populations with epistasis to show the convenience of using a recently developed model, NOIA, to perform estimates of genetic effects and the decomposition of the genetic variance that are orthogonal even under deviations from the Hardy-Weinberg proportions. We develop the theory for how to use this model in interval mapping of quantitative trait loci using Halley-Knott regressions, and we analyze a real data set to illustrate the advantage of using this approach in practice. In this example, we show that departures from the Hardy-Weinberg proportions that are expected by sampling alone substantially alter the orthogonal estimates of genetic effects when other statistical models, like F2 or G2A, are used instead of NOIA. Finally, for the first time from real data, we provide estimates of functional genetic effects as sets of effects of natural allele substitutions in a particular genotype, which enriches the debate on the interpretation of genetic effects as implemented both in functional and in statistical models. We also discuss further implementations leading to a completely general genotype-phenotype map.

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SNPs meet CNVs in genome-wide association studies: HGV2007 meeting report.

Publication Date: 2008 Apr PMID: 18437244
Authors: Estivill, X. - Cox, N. J. - Chanock, S. J. - Kwok, P. Y. - Scherer, S. W. - Brookes, A. J.
Journal: PLoS Genet

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Proofreading activity of DNA polymerase Pol2 mediates 3'-end processing during nonhomologous end joining in yeast.

Publication Date: 2008 Apr PMID: 18437220
Authors: Tseng, S. F. - Gabriel, A. - Teng, S. C.
Journal: PLoS Genet

Genotoxic agents that cause double-strand breaks (DSBs) often generate damage at the break termini. Processing enzymes, including nucleases and polymerases, must remove damaged bases and/or add new bases before completion of repair. Artemis is a nuclease involved in mammalian nonhomologous end joining (NHEJ), but in Saccharomyces cerevisiae the nucleases and polymerases involved in NHEJ pathways are poorly understood. Only Pol4 has been shown to fill the gap that may form by imprecise pairing of overhanging 3' DNA ends. We previously developed a chromosomal DSB assay in yeast to study factors involved in NHEJ. Here, we use this system to examine DNA polymerases required for NHEJ in yeast. We demonstrate that Pol2 is another major DNA polymerase involved in imprecise end joining. Pol1 modulates both imprecise end joining and more complex chromosomal rearrangements, and Pol3 is primarily involved in NHEJ-mediated chromosomal rearrangements. While Pol4 is the major polymerase to fill the gap that may form by imprecise pairing of overhanging 3' DNA ends, Pol2 is important for the recession of 3' flaps that can form during imprecise pairing. Indeed, a mutation in the 3'-5' exonuclease domain of Pol2 dramatically reduces the frequency of end joins formed with initial 3' flaps. Thus, Pol2 performs a key 3' end-processing step in NHEJ.

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Coordinated regulation of intestinal functions in C. elegans by LIN-35/Rb and SLR-2.

Publication Date: 2008 Apr PMID: 18437219
Authors: Kirienko, N. V. - McEnerney, J. D. - Fay, D. S.
Journal: PLoS Genet

LIN-35 is the sole C. elegans representative of the pocket protein family, which includes the mammalian Retinoblastoma protein pRb and its paralogs p107 and p130. In addition to having a well-established and central role in cell cycle regulation, pocket proteins have been increasingly implicated in the control of critical and diverse developmental and cellular processes. To gain a greater understanding of the roles of pocket proteins during development, we have characterized a synthetic genetic interaction between lin-35 and slr-2, which we show encodes a C2H2-type Zn-finger protein. Whereas animals harboring single mutations in lin-35 or slr-2 are viable and fertile, lin-35; slr-2 double mutants arrest uniformly in early larval development without obvious morphological defects. Using a combination of approaches including transcriptome profiling, mosaic analysis, starvation assays, and expression analysis, we demonstrate that both LIN-35 and SLR-2 act in the intestine to regulate the expression of many genes required for normal nutrient utilization. These findings represent a novel role for pRb family members in the maintenance of organ function. Our studies also shed light on the mechanistic basis of genetic redundancy among transcriptional regulators and suggest that synthetic interactions may result from the synergistic misregulation of one or more common targets.

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Defining the role of the MHC in autoimmunity: a review and pooled analysis.

Publication Date: 2008 Apr PMID: 18437207
Authors: Fernando, M. M. - Stevens, C. R. - Walsh, E. C. - De Jager, P. L. - Goyette, P. - Plenge, R. M. - Vyse, T. J. - Rioux, J. D.
Journal: PLoS Genet

The major histocompatibility complex (MHC) is one of the most extensively studied regions in the human genome because of the association of variants at this locus with autoimmune, infectious, and inflammatory diseases. However, identification of causal variants within the MHC for the majority of these diseases has remained difficult due to the great variability and extensive linkage disequilibrium (LD) that exists among alleles throughout this locus, coupled with inadequate study design whereby only a limited subset of about 20 from a total of approximately 250 genes have been studied in small cohorts of predominantly European origin. We have performed a review and pooled analysis of the past 30 years of research on the role of the MHC in six genetically complex disease traits - multiple sclerosis (MS), type 1 diabetes (T1D), systemic lupus erythematosus (SLE), ulcerative colitis (UC), Crohn's disease (CD), and rheumatoid arthritis (RA) - in order to consolidate and evaluate the current literature regarding MHC genetics in these common autoimmune and inflammatory diseases. We corroborate established MHC disease associations and identify predisposing variants that previously have not been appreciated. Furthermore, we find a number of interesting commonalities and differences across diseases that implicate both general and disease-specific pathogenetic mechanisms in autoimmunity.

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Primary coenzyme Q deficiency in Pdss2 mutant mice causes isolated renal disease.

Publication Date: 2008 Apr PMID: 18437205
Authors: Peng, M. - Falk, M. J. - Haase, V. H. - King, R. - Polyak, E. - Selak, M. - Yudkoff, M. - Hancock, W. W. - Meade, R. - Saiki, R. - Lunceford, A. L. - Clarke, C. F. - L Gasser, D.
Journal: PLoS Genet

Coenzyme Q (CoQ) is an essential electron carrier in the respiratory chain whose deficiency has been implicated in a wide variety of human mitochondrial disease manifestations. Its multi-step biosynthesis involves production of polyisoprenoid diphosphate in a reaction that requires the enzymes be encoded by PDSS1 and PDSS2. Homozygous mutations in either of these genes, in humans, lead to severe neuromuscular disease, with nephrotic syndrome seen in PDSS2 deficiency. We now show that a presumed autoimmune kidney disease in mice with the missense Pdss2(kd/kd) genotype can be attributed to a mitochondrial CoQ biosynthetic defect. Levels of CoQ9 and CoQ10 in kidney homogenates from B6.Pdss2(kd/kd) mutants were significantly lower than those in B6 control mice. Disease manifestations originate specifically in glomerular podocytes, as renal disease is seen in Podocin/cre,Pdss2(loxP/loxP) knockout mice but not in conditional knockouts targeted to renal tubular epithelium, monocytes, or hepatocytes. Liver-conditional B6.Alb/cre,Pdss2(loxP/loxP) knockout mice have no overt disease despite demonstration that their livers have undetectable CoQ9 levels, impaired respiratory capacity, and significantly altered intermediary metabolism as evidenced by transcriptional profiling and amino acid quantitation. These data suggest that disease manifestations of CoQ deficiency relate to tissue-specific respiratory capacity thresholds, with glomerular podocytes displaying the greatest sensitivity to Pdss2 impairment.

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Heterogeneity of breast cancer associations with five susceptibility Loci by clinical and pathological characteristics.

Publication Date: 2008 Apr PMID: 18437204
Authors: Garcia-Closas, M. - Hall, P. - Nevanlinna, H. - Pooley, K. - Morrison, J. - Richesson, D. A. - Bojesen, S. E. - Nordestgaard, B. G. - Axelsson, C. K. - Arias, J. I. - Milne, R. L. - Ribas, G. - Gonzalez-Neira, A. - Benitez, J. - Zamora, P. - Brauch, H. - Justenhoven, C. - Hamann, U. - Ko, Y. D. - Bruening, T. - Haas, S. - Dork, T. - Schurmann, P. - Hillemanns, P. - Bogdanova, N. - Bremer, M. - Karstens, J. H. - Fagerholm, R. - Aaltonen, K. - Aittomaki, K. - von Smitten, K. - Blomqvist, C. - Mannermaa, A. - Uusitupa, M. - Eskelinen, M. - Tengstrom, M. - Kosma, V. M. - Kataja, V. - Chenevix-Trench, G. - Spurdle, A. B. - Beesley, J. - Chen, X. - Australian Ovarian Cancer Management, G. r. o. u. p. - Kathleen Cuningham Foundation Consortium For Research Into Familial, B. r. e. a. s. t. - Cancer - Devilee, P. - van Asperen, C. J. - Jacobi, C. E. - Tollenaar, R. A. - Huijts, P. E. - Klijn, J. G. - Chang-Claude, J. - Kropp, S. - Slanger, T. - Flesch-Janys, D. - Mutschelknauss, E. - Salazar, R. - Wang-Gohrke, S. - Couch, F. - Goode, E. L. - Olson, J. E. - Vachon, C. - Fredericksen, Z. S. - Giles, G. G. - Baglietto, L. - Severi, G. - Hopper, J. L. - English, D. R. - Southey, M. C. - Haiman, C. A. - Henderson, B. E. - Kolonel, L. N. - Le Marchand, L. - Stram, D. O. - Hunter, D. J. - Hankinson, S. E. - Cox, D. G. - Tamimi, R. - Kraft, P. - Sherman, M. E. - Chanock, S. J. - Lissowska, J. - Brinton, L. A. - Peplonska, B. - Klijn, J. G. - Hooning, M. J. - Meijers-Heijboer, H. - Collee, J. M. - van den Ouweland, A. - Uitterlinden, A. G. - Liu, J. - Lin, L. Y. - Yuqing, L. - Humphreys, K. - Czene, K. - Cox, A. - Balasubramanian, S. P. - Cross, S. S. - Reed, M. W. - Blows, F. - Driver, K. - Dunning, A. - Tyrer, J. - Ponder, B. A. - Sangrajrang, S. - Brennan, P. - McKay, J. - Odefrey, F. - Gabrieau, V. - Sigurdson, A. - Doody, M. - Struewing, J. P. - Alexander, B. - Easton, D. F. - Pharoah, P. D.
Journal: PLoS Genet

A three-stage genome-wide association study recently identified single nucleotide polymorphisms (SNPs) in five loci (fibroblast growth receptor 2 (FGFR2), trinucleotide repeat containing 9 (TNRC9), mitogen-activated protein kinase 3 K1 (MAP3K1), 8q24, and lymphocyte-specific protein 1 (LSP1)) associated with breast cancer risk. We investigated whether the associations between these SNPs and breast cancer risk varied by clinically important tumor characteristics in up to 23,039 invasive breast cancer cases and 26,273 controls from 20 studies. We also evaluated their influence on overall survival in 13,527 cases from 13 studies. All participants were of European or Asian origin. rs2981582 in FGFR2 was more strongly related to ER-positive (per-allele OR (95%CI) = 1.31 (1.27-1.36)) than ER-negative (1.08 (1.03-1.14)) disease (P for heterogeneity = 10(-13)). This SNP was also more strongly related to PR-positive, low grade and node positive tumors (P = 10(-5), 10(-8), 0.013, respectively). The association for rs13281615 in 8q24 was stronger for ER-positive, PR-positive, and low grade tumors (P = 0.001, 0.011 and 10(-4), respectively). The differences in the associations between SNPs in FGFR2 and 8q24 and risk by ER and grade remained significant after permutation adjustment for multiple comparisons and after adjustment for other tumor characteristics. Three SNPs (rs2981582, rs3803662, and rs889312) showed weak but significant associations with ER-negative disease, the strongest association being for rs3803662 in TNRC9 (1.14 (1.09-1.21)). rs13281615 in 8q24 was associated with an improvement in survival after diagnosis (per-allele HR = 0.90 (0.83-0.97). The association was attenuated and non-significant after adjusting for known prognostic factors. Our findings show that common genetic variants influence the pathological subtype of breast cancer and provide further support for the hypothesis that ER-positive and ER-negative disease are biologically distinct. Understanding the etiologic heterogeneity of breast cancer may ultimately result in improvements in prevention, early detection, and treatment.

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Small RNA-directed epigenetic natural variation in Arabidopsis thaliana.

Publication Date: 2008 Apr PMID: 18437202
Authors: Zhai, J. - Liu, J. - Liu, B. - Li, P. - Meyers, B. C. - Chen, X. - Cao, X.
Journal: PLoS Genet

Progress in epigenetics has revealed mechanisms that can heritably regulate gene function independent of genetic alterations. Nevertheless, little is known about the role of epigenetics in evolution. This is due in part to scant data on epigenetic variation among natural populations. In plants, small interfering RNA (siRNA) is involved in both the initiation and maintenance of gene silencing by directing DNA methylation and/or histone methylation. Here, we report that, in the model plant Arabidopsis thaliana, a cluster of approximately 24 nt siRNAs found at high levels in the ecotype Landsberg erecta (Ler) could direct DNA methylation and heterochromatinization at a hAT element adjacent to the promoter of FLOWERING LOCUS C (FLC), a major repressor of flowering, whereas the same hAT element in ecotype Columbia (Col) with almost identical DNA sequence, generates a set of low abundance siRNAs that do not direct these activities. We have called this hAT element MPF for Methylated region near Promoter of FLC, although de novo methylation triggered by an inverted repeat transgene at this region in Col does not alter its FLC expression. DNA methylation of the Ler allele MPF is dependent on genes in known silencing pathways, and such methylation is transmissible to Col by genetic crosses, although with varying degrees of penetrance. A genome-wide comparison of Ler and Col small RNAs identified at least 68 loci matched by a significant level of approximately 24 nt siRNAs present specifically in Ler but not Col, where nearly half of the loci are related to repeat or TE sequences. Methylation analysis revealed that 88% of the examined loci (37 out of 42) were specifically methylated in Ler but not Col, suggesting that small RNA can direct epigenetic differences between two closely related Arabidopsis ecotypes.

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Identification of CIITA regulated genetic module dedicated for antigen presentation.

Publication Date: 2008 Apr PMID: 18437201
Authors: Krawczyk, M. - Seguin-Estevez, Q. - Leimgruber, E. - Sperisen, P. - Schmid, C. - Bucher, P. - Reith, W.
Journal: PLoS Genet

The class II trans-activator CIITA is a transcriptional co-activator required for the expression of Major Histocompatibility Complex (MHC) genes. Although the latter function is well established, the global target-gene specificity of CIITA had not been defined. We therefore generated a comprehensive list of its target genes by performing genome-wide scans employing four different approaches designed to identify promoters that are occupied by CIITA in two key antigen presenting cells, B cells and dendritic cells. Surprisingly, in addition to MHC genes, only nine new targets were identified and validated by extensive functional and expression analysis. Seven of these genes are known or likely to function in processes contributing to MHC-mediated antigen presentation. The remaining two are of unknown function. CIITA is thus uniquely dedicated for genes implicated in antigen presentation. The finding that CIITA regulates such a highly focused gene expression module sets it apart from all other transcription factors, for which large-scale binding-site mapping has indicated that they exert pleiotropic functions and regulate large numbers of genes.

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Global transcriptome and deletome profiles of yeast exposed to transition metals.

Publication Date: 2008 Apr PMID: 18437200
Authors: Jin, Y. H. - Dunlap, P. E. - McBride, S. J. - Al-Refai, H. - Bushel, P. R. - Freedman, J. H.
Journal: PLoS Genet

A variety of pathologies are associated with exposure to supraphysiological concentrations of essential metals and to non-essential metals and metalloids. The molecular mechanisms linking metal exposure to human pathologies have not been clearly defined. To address these gaps in our understanding of the molecular biology of transition metals, the genomic effects of exposure to Group IB (copper, silver), IIB (zinc, cadmium, mercury), VIA (chromium), and VB (arsenic) elements on the yeast Saccharomyces cerevisiae were examined. Two comprehensive sets of metal-responsive genomic profiles were generated following exposure to equi-toxic concentrations of metal: one that provides information on the transcriptional changes associated with metal exposure (transcriptome), and a second that provides information on the relationship between the expression of approximately 4,700 non-essential genes and sensitivity to metal exposure (deletome). Approximately 22% of the genome was affected by exposure to at least one metal. Principal component and cluster analyses suggest that the chemical properties of the metal are major determinants in defining the expression profile. Furthermore, cells may have developed common or convergent regulatory mechanisms to accommodate metal exposure. The transcriptome and deletome had 22 genes in common, however, comparison between Gene Ontology biological processes for the two gene sets revealed that metal stress adaptation and detoxification categories were commonly enriched. Analysis of the transcriptome and deletome identified several evolutionarily conserved, signal transduction pathways that may be involved in regulating the responses to metal exposure. In this study, we identified genes and cognate signaling pathways that respond to exposure to essential and non-essential metals. In addition, genes that are essential for survival in the presence of these metals were identified. This information will contribute to our understanding of the molecular mechanism by which organisms respond to metal stress, and could lead to an understanding of the connection between environmental stress and signal transduction pathways.

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Analysis of chimpanzee history based on genome sequence alignments.

Publication Date: 2008 Apr PMID: 18421364
Authors: Caswell, J. L. - Mallick, S. - Richter, D. J. - Neubauer, J. - Schirmer, C. - Gnerre, S. - Reich, D.
Journal: PLoS Genet

Population geneticists often study small numbers of carefully chosen loci, but it has become possible to obtain orders of magnitude for more data from overlaps of genome sequences. Here, we generate tens of millions of base pairs of multiple sequence alignments from combinations of three western chimpanzees, three central chimpanzees, an eastern chimpanzee, a bonobo, a human, an orangutan, and a macaque. Analysis provides a more precise understanding of demographic history than was previously available. We show that bonobos and common chimpanzees were separated approximately 1,290,000 years ago, western and other common chimpanzees approximately 510,000 years ago, and eastern and central chimpanzees at least 50,000 years ago. We infer that the central chimpanzee population size increased by at least a factor of 4 since its separation from western chimpanzees, while the western chimpanzee effective population size decreased. Surprisingly, in about one percent of the genome, the genetic relationships between humans, chimpanzees, and bonobos appear to be different from the species relationships. We used PCR-based resequencing to confirm 11 regions where chimpanzees and bonobos are not most closely related. Study of such loci should provide information about the period of time 5-7 million years ago when the ancestors of humans separated from those of the chimpanzees.

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