bluuebunny/crossref_metadata_2025 huggingface dataset:
Involvement of the ArabidopsisSWI2/SNF2Chromatin Remodeling Gene Family in DNA Damage Response and Recombination
Authors: Shaked Hezi , Avivi-Ragolsky Naomi , Levy Avraham A
Published: March 2006

The genome of plants, like that of other eukaryotes, is organized into chromatin, a compact structure that reduces the accessibility of DNA to machineries such as transcription, replication, and DNA recombination and repair. Plant genes, which contain the characteristic ATPase/helicase motifs of the chromatin remodeling Swi2/Snf2 family of proteins, have been thoroughly studied, but their role in homologous recombination or DNA repair has received limited attention. We have searched for homologs of the yeast RAD54 gene, whose role in recombination and repair and in chromatin remodeling is well established. Forty Arabidopsis SWI2/SNF2 genes were identified and the function of a selected group of 14 was analyzed. Mutant analysis and/or RNAi-mediated silencing showed that 11 of the 14 genes tested played a role in response to DNA damage. Two of the 14 genes were involved in homologous recombination between inverted repeats. The putative ortholog of RAD54 and close homologs of ERCC6/RAD26 were involved in DNA damage response, suggesting functional conservation across kingdoms. In addition, genes known for their role in development, such as PICKLE/GYMNOS and PIE1, or in silencing, such as DDM1, turned out to also be involved in DNA damage response. A comparison of ddm1 and met1 mutants suggests that DNA damage response is affected essentially by chromatin structure and that cytosine methylation is less critical. These results emphasize the broad involvement of the SWI2/SNF2 family, and thus of chromatin remodeling, in genome maintenance and the link between epigenetic and genetic processes.
Effect of Varying Epistasis on the Evolution of Recombination
Authors: Kouyos Roger D , Otto Sarah P , Bonhoeffer Sebastian
Published: March 2006

Whether recombination decelerates or accelerates a population's response to selection depends, at least in part, on how fitness-determining loci interact. Realistically, all genomes likely contain fitness interactions both with positive and with negative epistasis. Therefore, it is crucial to determine the conditions under which the potential beneficial effects of recombination with negative epistasis prevail over the detrimental effects of recombination with positive epistasis. Here, we examine the simultaneous effects of diverse epistatic interactions with different strengths and signs in a simplified model system with independent pairs of interacting loci and selection acting only on the haploid phase. We find that the average form of epistasis does not predict the average amount of linkage disequilibrium generated or the impact on a recombination modifier when compared to results using the entire distribution of epistatic effects and associated single-mutant effects. Moreover, we show that epistatic interactions of a given strength can produce very different effects, having the greatest impact when selection is weak. In summary, we observe that the evolution of recombination at mutation–selection balance might be driven by a small number of interactions with weak selection rather than by the average epistasis of all interactions. We illustrate this effect with an analysis of published data of Saccharomyces cerevisiae. Thus to draw conclusions on the evolution of recombination from experimental data, it is necessary to consider the distribution of epistatic interactions together with the associated selection coefficients.
A Novel Method for Detecting Intramolecular Coevolution: Adding a Further Dimension to Selective Constraints Analyses
Authors: Fares Mario A , Travers Simon A A
Published: March 2006

Protein evolution depends on intramolecular coevolutionary networks whose complexity is proportional to the underlying functional and structural interactions among sites. Here we present a novel approach that vastly improves the sensitivity of previous methods for detecting coevolution through a weighted comparison of divergence between amino acid sites. The analysis of the HIV-1 Gag protein detected convergent adaptive coevolutionary events responsible for the selective variability emerging between subtypes. Coevolution analysis and functional data for heat-shock proteins, Hsp90 and GroEL, highlight that almost all detected coevolving sites are functionally or structurally important. The results support previous suggestions pinpointing the complex interdomain functional interactions within these proteins and we propose new amino acid sites as important for interdomain functional communication. Three-dimensional information sheds light on the functional and structural constraints governing the coevolution between sites. Our covariation analyses propose two types of coevolving sites in agreement with previous reports: pairs of sites spatially proximal, where compensatory mutations could maintain the local structure stability, and clusters of distant sites located in functional domains, suggesting a functional dependency between them. All sites detected under adaptive evolution in these proteins belong to coevolution groups, further underlining the importance of testing for coevolution in selective constraints analyses.
Construction of a Single Nucleotide Polymorphism Linkage Map for the Silkworm, Bombyx mori, Based on Bacterial Artificial Chromosome End Sequences
Authors: Yamamoto Kimiko , Narukawa Junko , Kadono-Okuda Keiko , Nohata Junko , Sasanuma Motoe , Suetsugu Yoshitaka , Banno Yutaka , Fujii Hiroshi , Goldsmith Marian R , Mita Kazuei
Published: March 2006

We have developed a linkage map for the silkworm Bombyx mori based on single nucleotide polymorphisms (SNPs) between strains p50T and C108T initially found on regions corresponding to the end sequences of bacterial artificial chromosome (BAC) clones. Using 190 segregants from a backcross of a p50T female × an F1 (p50T × C108T) male, we analyzed segregation patterns of 534 SNPs between p50T and C108T, detected among 3840 PCR amplicons, each associated with a p50T BAC end sequence. This enabled us to construct a linkage map composed of 534 SNP markers spanning 1305 cM in total length distributed over the expected 28 linkage groups. Of the 534 BACs whose ends harbored the SNPs used to construct the linkage map, 89 were associated with 107 different ESTs. Since each of the SNP markers is directly linked to a specific genomic BAC clone and to whole-genome sequence data, and some of them are also linked to EST data, the SNP linkage map will be a powerful tool for investigating silkworm genome properties, mutation mapping, and map-based cloning of genes of industrial and agricultural interest.
Identification of Genetic Regions of Importance for Reproductive Performance in Female Mice
Authors: Liljander Maria , Sällström Mary-Ann , Andersson Sara , Wernhoff Patrik , Andersson Åsa , Holmdahl Rikard , Mattsson Ragnar
Published: March 2006

Both environmental and genetic factors can dramatically affect reproductive performance in mice. In this study we have focused on the identification of genetic regions, quantitative trait loci (QTL), which affect the breeding capacity of female mice. We have identified polymorphic microsatellite markers for the mouse strains used and performed a genomewide scan on 237 females from a gene-segregating backcross between a high breeder and a relatively poor breeder. The high-breeder mouse strain we used is the inbred NFR/N mouse (MHC haplotype H-2q), which has extraordinary good breeding properties. The moderate breeder chosen for F1 and N2 progeny was B10.Q, which is a genetically well-characterized MHC-congenic mouse of the H-2q haplotype. Each of the 237 females of the N2 generation was allowed to mate twice with MHC-congenic B10.RIII (H-2r) males and twice with B10.Q males. A predetermined number of phenotypes related to reproductive performance were recorded, and these included litter size, neonatal growth, and pregnancy rate. Loci controlling litter size were detected on chromosomes 1 (Fecq3) and 9 (Fecq4). The neonatal growth phenotype was affected by Fecq3 and a locus on chromosome 9 (Neogq1). On chromosome 11 two loci affecting the pregnancy rate (Pregq1 and Pregq2) were identified. Furthermore, on chromosomes 13 and 17 we found loci (Pregq3 and Pregq4) influencing the outcome of allogeneic pregnancy (allogeneic by means of MHC disparity between mother and fetuses). A locus on chromosome 1 affecting maternal body weight was also identified and has been denoted Bwq7. It is well known that reproductive performance is polygenically controlled, and the identification of the major loci in this complex process opens the possibility of investigating the natural genetic control of reproduction.
Challenges of Detecting Directional Selection After a Bottleneck: Lessons From Sorghum bicolor
Authors: Hamblin Martha T , Casa Alexandra M , Sun Hong , Murray Seth C , Paterson Andrew H , Aquadro Charles F , Kresovich Stephen
Published: March 2006

Multilocus surveys of sequence variation can be used to identify targets of directional selection, which are expected to have reduced levels of variation. Following a population bottleneck, the signal of directional selection may be hard to detect because many loci may have low variation by chance and the frequency spectrum of variation may be perturbed in ways that resemble the effects of selection. Cultivated Sorghum bicolor contains a subset of the genetic diversity found in its wild ancestor(s) due to the combined effects of a domestication bottleneck and human selection on traits associated with agriculture. As a framework for distinguishing between the effects of demography and selection, we sequenced 204 loci in a diverse panel of 17 cultivated S. bicolor accessions. Genomewide patterns of diversity depart strongly from equilibrium expectations with regard to the variance of the number of segregating sites, the site frequency spectrum, and haplotype configuration. Furthermore, gene genealogies of most loci with an excess of low frequency variants and/or an excess of segregating sites do not show the characteristic signatures of directional and diversifying selection, respectively. A simple bottleneck model provides an improved but inadequate fit to the data, suggesting the action of other population-level factors, such as population structure and migration. Despite a known history of recent selection, we find little evidence for directional selection, likely due to low statistical power and lack of an appropriate null model.
Strong Regional Heterogeneity in Base Composition Evolution on the Drosophila X Chromosome
Authors: Ko Wen-Ya , Piao Shengfu , Akashi Hiroshi
Published: March 2006

Fluctuations in base composition appear to be prevalent in Drosophila and mammal genome evolution, but their timescale, genomic breadth, and causes remain obscure. Here, we study base composition evolution within the X chromosomes of Drosophila melanogaster and five of its close relatives. Substitutions were inferred on six extant and two ancestral lineages for 14 near-telomeric and 9 nontelomeric genes. GC content evolution is highly variable both within the genome and within the phylogenetic tree. In the lineages leading to D. yakuba and D. orena, GC content at silent sites has increased rapidly near telomeres, but has decreased in more proximal (nontelomeric) regions. D. orena shows a 17-fold excess of GC-increasing vs. AT-increasing synonymous changes within a small (∼130-kb) region close to the telomeric end. Base composition changes within introns are consistent with changes in mutation patterns, but stronger GC elevation at synonymous sites suggests contributions of natural selection or biased gene conversion. The Drosophila yakuba lineage shows a less extreme elevation of GC content distributed over a wider genetic region (∼1.2 Mb). A lack of change in GC content for most introns within this region suggests a role of natural selection in localized base composition fluctuations.
Comparative Physical Mapping of the Apospory-Specific Genomic Region in Two Apomictic Grasses: Pennisetum squamulatum and Cenchrus ciliaris
Authors: Goel Shailendra , Chen Zhenbang , Akiyama Yukio , Conner Joann A , Basu Manojit , Gualtieri Gustavo , Hanna Wayne W , Ozias-Akins Peggy
Published: March 2006

In gametophytic apomicts of the aposporous type, each cell of the embryo sac is genetically identical to somatic cells of the ovule because they are products of mitosis, not of meiosis. The egg of the aposporous embryo sac follows parthenogenetic development into an embryo; therefore, uniform progeny result even from heterozygous plants, a trait that would be valuable for many crop species. Attempts to introgress apomixis from wild relatives into major crops through traditional breeding have been hindered by low or no recombination within the chromosomal region governing this trait (the apospory-specific genomic region or ASGR). The lack of recombination also has been a major obstacle to positional cloning of key genes. To further delineate and characterize the nonrecombinant ASGR, we have identified eight new ASGR-linked, AFLP-based molecular markers, only one of which showed recombination with the trait for aposporous embryo sac development. Bacterial artificial chromosome (BAC) clones identified with the ASGR-linked AFLPs or previously mapped markers, when mapped by fluorescence in situ hybridization in Pennisetum squamulatum and Cenchrus ciliaris, showed almost complete macrosynteny between the two apomictic grasses throughout the ASGR, although with an inverted order. A BAC identified with the recombinant AFLP marker mapped most proximal to the centromere of the ASGR-carrier chromosome in P. squamulatum but was not located on the ASGR-carrier chromosome in C. ciliaris. Exceptional regions where synteny was disrupted probably are nonessential for expression of the aposporous trait. The ASGR appears to be maintained as a haplotype even though its position in the genome can be variable.
Rates and Patterns of Chromosomal Evolution in Drosophila pseudoobscura and D. miranda
Authors: Bartolomé Carolina , Charlesworth Brian
Published: March 2006

Comparisons of gene orders between species permit estimation of the rate of chromosomal evolution since their divergence from a common ancestor. We have compared gene orders on three chromosomes of Drosophila pseudoobscura with its close relative, D. miranda, and the distant outgroup species, D. melanogaster, by using the public genome sequences of D. pseudoobscura and D. melanogaster and ∼50 in situ hybridizations of gene probes in D. miranda. We find no evidence for extensive transfer of genes among chromosomes in D. miranda. The rates of chromosomal rearrangements between D. miranda and D. pseudoobscura are far higher than those found before in Drosophila and approach those for nematodes, the fastest rates among higher eukaryotes. In addition, we find that the D. pseudoobscura chromosome with the highest level of inversion polymorphism (Muller's element C) does not show an unusually fast rate of evolution with respect to chromosome structure, suggesting that this classic case of inversion polymorphism reflects selection rather than mutational processes. On the basis of our results, we propose possible ancestral arrangements for the D. pseudoobscura C chromosome, which are different from those in the current literature. We also describe a new method for correcting for rearrangements that are not detected with a limited set of markers.
Determining the Effective Dimensionality of the Genetic Variance–Covariance Matrix
Authors: Hine Emma , Blows Mark W
Published: March 2006

Determining the dimensionality of G provides an important perspective on the genetic basis of a multivariate suite of traits. Since the introduction of Fisher's geometric model, the number of genetically independent traits underlying a set of functionally related phenotypic traits has been recognized as an important factor influencing the response to selection. Here, we show how the effective dimensionality of G can be established, using a method for the determination of the dimensionality of the effect space from a multivariate general linear model introduced by Amemiya (1985). We compare this approach with two other available methods, factor-analytic modeling and bootstrapping, using a half-sib experiment that estimated G for eight cuticular hydrocarbons of Drosophila serrata. In our example, eight pheromone traits were shown to be adequately represented by only two underlying genetic dimensions by Amemiya's approach and factor-analytic modeling of the covariance structure at the sire level. In contrast, bootstrapping identified four dimensions with significant genetic variance. A simulation study indicated that while the performance of Amemiya's method was more sensitive to power constraints, it performed as well or better than factor-analytic modeling in correctly identifying the original genetic dimensions at moderate to high levels of heritability. The bootstrap approach consistently overestimated the number of dimensions in all cases and performed less well than Amemiya's method at subspace recovery.
Natural Variation in a Subtelomeric Region of Arabidopsis: Implications for the Genomic Dynamics of a Chromosome End
Authors: Kuo Hui-Fen , Olsen Kenneth M , Richards Eric J
Published: March 2006

We investigated genome dynamics at a chromosome end in the model plant Arabidopsis thaliana through a study of natural variation in 35 wild accessions. We focused on the single-copy subtelomeric region of chromosome 1 north (∼3.5 kb), which represents the relatively simple organization of subtelomeric regions in this species. PCR fragment-length variation across the subtelomeric region indicated that the 1.4-kb distal region showed elevated structural variation relative to the centromere-proximal region. Examination of nucleotide sequences from this 1.4-kb region revealed diverse DNA rearrangements, including an inversion, several deletions, and an insertion of a retrotransposon LTR. The structures at the deletion and inversion breakpoints are characteristic of simple deletion-associated nonhomologous end-joining (NHEJ) events. There was strong linkage disequilibrium between the distal subtelomeric region and the proximal telomere, which contains degenerate and variant telomeric repeats. Variation in the proximal telomere was characterized by the expansion and deletion of blocks of repeats. Our sample of accessions documented two independent chromosome-healing events associated with terminal deletions of the subtelomeric region as well as the capture of a scrambled mitochondrial DNA segment in the proximal telomeric array. This natural variation study highlights the variety of genomic events that drive the fluidity of chromosome termini.
Identification of Novel Mutations in ACT1 and SLA2 That Suppress the Actin-Cable-Overproducing Phenotype Caused by Overexpression of a Dominant Active Form of Bni1p in Saccharomyces cerevisiae
Authors: Yoshiuchi Shiro , Yamamoto Takaharu , Sakane Hiroshi , Kadota Jun , Mochida Junko , Asaka Masahiro , Tanaka Kazuma
Published: March 2006

A formin Bni1p nucleates actin to assemble actin cables, which guide the polarized transport of secretory vesicles in budding yeast. We identified mutations that suppressed both the lethality and the excessive actin cable formation caused by overexpression of a truncated Bni1p (BNI1ΔN). Two recessive mutations, act1-301 in the actin gene and sla2-82 in a gene involved in cortical actin patch assembly, were identified. The isolation of sla2-82 was unexpected, because cortical actin patches are required for the internalization step of endocytosis. Both act1-301 and sla2-82 exhibited synthetic growth defects with bni1Δ. act1-301, which resulted in an E117K substitution, interacted genetically with mutations in profilin (PFY1) and BUD6, suggesting that Act1-301p was not fully functional in formin-mediated polymerization. sla2-82 also interacted genetically with genes involved in actin cable assembly. Some experiments, however, suggested that the effects of sla2-82 were caused by depletion of actin monomers, because the temperature-sensitive growth phenotype of the bni1Δ sla2-82 mutant was suppressed by increased expression of ACT1. The isolation of suppressors of the BNI1ΔN phenotype may provide a useful system for identification of actin amino-acid residues that are important for formin-mediated actin polymerization and mutations that affect the availability of actin monomers.
A Second-Generation Integrated Map of the Silkworm Reveals Synteny and Conserved Gene Order Between Lepidopteran Insects
Authors: Yasukochi Yuji , Ashakumary Laksmikutty A , Baba Kotaro , Yoshido Atsuo , Sahara Ken
Published: March 2006

A second-generation linkage map was constructed for the silkworm, Bombyx mori, focusing on mapping Bombyx sequences appearing in public nucleotide databases and bacterial artificial chromosome (BAC) contigs. A total of 874 BAC contigs containing 5067 clones (22% of the library) were constructed by PCR-based screening with sequence-tagged sites (STSs) derived from whole-genome shotgun (WGS) sequences. A total of 523 BAC contigs, including 342 independent genes registered in public databases and 85 expressed sequence tags (ESTs), were placed onto the linkage map. We found significant synteny and conserved gene order between B. mori and a nymphalid butterfly, Heliconius melpomene, in four linkage groups (LGs), strongly suggesting that using B. mori as a reference for comparative genomics in Lepidotera is highly feasible.
Characterization of the split ends-Like Gene spenito Reveals Functional Antagonism Between SPOC Family Members During Drosophila Eye Development
Authors: Jemc Jennifer , Rebay Ilaria
Published: March 2006

The novel family of SPOC domain proteins is composed of broadly conserved nuclear factors that fall into two subclasses, termed large and small, based on protein size. Members of the large subgroup, which includes Drosophila SPEN and human SHARP, have been characterized as transcriptional corepressors acting downstream of a variety of essential cell signaling pathways, while those of the small subclass have remained largely unstudied. Since SPEN has been implicated in Drosophila eye development, and the small SPOC protein NITO is also expressed in the developing eye, we have used this context to perform a structure–function analysis of NITO and to examine the relationship between the two SPOC family subclasses. Our results demonstrate that the phenotypes obtained from overexpressing NITO share striking similarity to those associated with loss of spen. Dosage-sensitive genetic interactions further support a model of functional antagonism between NITO and SPEN during Drosophila eye development. These results suggest that large and small SPOC family proteins may have opposing functions in certain developmental contexts.
Searching for Neuronal Left/Right Asymmetry: Genomewide Analysis of Nematode Receptor-Type Guanylyl Cyclases
Authors: Ortiz Christopher O , Etchberger John F , Posy Shoshana L , Frøkjær-Jensen Christian , Lockery Shawn , Honig Barry , Hobert Oliver
Published: March 2006

Functional left/right asymmetry (“laterality”) is a fundamental feature of many nervous systems, but only very few molecular correlates to functional laterality are known. At least two classes of chemosensory neurons in the nematode Caenorhabditis elegans are functionally lateralized. The gustatory neurons ASE left (ASEL) and ASE right (ASER) are two bilaterally symmetric neurons that sense distinct chemosensory cues and express a distinct set of four known chemoreceptors of the guanylyl cyclase (gcy) gene family. To examine the extent of lateralization of gcy gene expression patterns in the ASE neurons, we have undertaken a genomewide analysis of all gcy genes. We report the existence of a total of 27 gcy genes encoding receptor-type guanylyl cyclases and of 7 gcy genes encoding soluble guanylyl cyclases in the complete genome sequence of C. elegans. We describe the expression pattern of all previously uncharacterized receptor-type guanylyl cyclases and find them to be highly biased but not exclusively restricted to the nervous system. We find that >41% (11/27) of all receptor-type guanylyl cyclases are expressed in the ASE gustatory neurons and that one-third of all gcy genes (9/27) are expressed in a lateral, left/right asymmetric manner in the ASE neurons. The expression of all laterally expressed gcy genes is under the control of a gene regulatory network composed of several transcription factors and miRNAs. The complement of gcy genes in the related nematode C. briggsae differs from C. elegans as evidenced by differences in chromosomal localization, number of gcy genes, and expression patterns. Differences in gcy expression patterns in the ASE neurons of C. briggsae arise from a difference in cis-regulatory elements and trans-acting factors that control ASE laterality. In sum, our results indicate the existence of a surprising multitude of putative chemoreceptors in the gustatory ASE neurons and suggest the existence of a substantial degree of laterality in gustatory signaling mechanisms in nematodes.
The Adaptor Protein soc-1/Gab1 Modifies Growth Factor Receptor Output in Caenorhabditis elegans
Authors: Hopper Neil A
Published: March 2006

Previous genetic analysis has shown that dos/soc-1/Gab1 functions positively in receptor tyrosine kinase (RTK)-stimulated Ras/Map kinase signaling through the recruitment of csw/ptp-2/Shp2. Using sensitized assays in Caenorhabditis elegans for let-23/Egfr and daf-2/InsR (insulin receptor-like) signaling, it is shown that soc-1/Gab1 inhibits phospholipase C-γ (PLCγ) and phosphatidylinositol 3′-kinase (PI3K)-mediated signaling. Furthermore, as well as stimulating Ras/Map kinase signaling, soc-1/Gab1 stimulates a poorly defined signaling pathway that represses class 2 daf-2 phenotypes. In addition, it is shown that SOC-1 binds the C-terminal SH3 domain of SEM-5. This binding is likely to be functional as the sem-5(n2195)G201R mutation, which disrupts SOC-1 binding, behaves in a qualitatively similar manner to a soc-1 null allele in all assays for let-23/Egfr and daf-2/InsR signaling that were examined. Further genetic analysis suggests that ptp-2/Shp2 mediates the negative function of soc-1/Gab1 in PI3K-mediated signaling, as well as the positive function in Ras/Map kinase signaling. Other effectors of soc-1/Gab1 are likely to inhibit PLCγ-mediated signaling and stimulate the poorly defined signaling pathway that represses class 2 daf-2 phenotypes. Thus, the recruitment of soc-1/Gab1, and its effectors, into the RTK-signaling complex modifies the cellular response by enhancing Ras/Map kinase signaling while inhibiting PI3K and PLCγ-mediated signaling.
Increase of the Spontaneous Mutation Rate in a Long-Term Experiment With Drosophila melanogaster
Authors: Ávila Victoria , Chavarrías David , Sánchez Enrique , Manrique Antonio , López-Fanjul Carlos , García-Dorado Aurora
Published: March 2006

In a previous experiment, the effect of 255 generations of mutation accumulation (MA) on the second chromosome viability of Drosophila melanogaster was studied using 200 full-sib MA1 lines and a large C1 control, both derived from a genetically homogeneous base population. At generation 265, one of those MA1 lines was expanded to start 150 new full-sib MA2 lines and a new C2 large control. After 46 generations, the rate of decline in mean viability in MA2 was ∼2.5 times that estimated in MA1, while the average degree of dominance of mutations was small and nonsignificant by generation 40 and moderate by generation 80. In parallel, the inbreeding depression rate for viability and the amount of additive variance for two bristle traits in C2 were 2–3 times larger than those in C1. The results are consistent with a mutation rate in the line from which MA2 and C2 were derived about 2.5 times larger than that in MA1. The mean viability of C2 remained roughly similar to that of C1, but the rate of MA2 line extinction increased progressively, leading to mutational collapse, which can be ascribed to accelerated mutation and/or synergy after important deleterious accumulation.
Selection Under Domestication: Evidence for a Sweep in the Rice Waxy Genomic Region
Authors: Olsen Kenneth M , Caicedo Ana L , Polato Nicholas , McClung Anna , McCouch Susan , Purugganan Michael D
Published: March 2006

Rice (Oryza sativa) was cultivated by Asian Neolithic farmers >11,000 years ago, and different cultures have selected for divergent starch qualities in the rice grain during and after the domestication process. An intron 1 splice donor site mutation of the Waxy gene is responsible for the absence of amylose in glutinous rice varieties. This mutation appears to have also played an important role in the origin of low amylose, nonglutinous temperate japonica rice varieties, which form a primary component of Northeast Asian cuisines. Waxy DNA sequence analyses indicate that the splice donor mutation is prevalent in temperate japonica rice varieties, but rare or absent in tropical japonica, indica, aus, and aromatic varieties. Sequence analysis across a 500-kb genomic region centered on Waxy reveals patterns consistent with a selective sweep in the temperate japonicas associated with the mutation. The size of the selective sweep (>250 kb) indicates very strong selection in this region, with an inferred selection coefficient that is higher than similar estimates from maize domestication genes or wild species. These findings demonstrate that selection pressures associated with crop domestication regimes can exceed by one to two orders of magnitude those observed for genes under even strong selection in natural systems.
Nonadditive Regulation of FRI and FLC Loci Mediates Flowering-Time Variation in Arabidopsis Allopolyploids
Authors: Wang Jianlin , Tian Lu , Lee Hyeon-Se , Chen Z Jeffrey
Published: March 2006

Allopolyploidy is formed by combining two or more divergent genomes and occurs throughout the evolutionary history of many plants and some animals. Transcriptome analysis indicates that many genes in various biological pathways, including flowering time, are expressed nonadditively (different from the midparent value). However, the mechanisms for nonadditive gene regulation in a biological pathway are unknown. Natural variation of flowering time is largely controlled by two epistatically acting loci, namely FRIGIDA (FRI) and FLOWERING LOCUS C (FLC). FRI upregulates FLC expression that represses flowering in Arabidopsis. Synthetic Arabidopsis allotetraploids contain two sets of FLC and FRI genes originating from Arabidopsis thaliana and A. arenosa, respectively, and flower late. Inhibition of early flowering is caused by upregulation of A. thaliana FLC (AtFLC) that is trans-activated by A. arenosa FRI (AaFRI). Two duplicate FLCs (AaFLC1 and AaFLC2) originating from A. arenosa are expressed in some allotetraploids but silenced in other lines. The expression variation in the allotetraploids is associated with deletions in the promoter regions and first introns of A. arenosa FLCs. The strong AtFLC and AaFLC loci are maintained in natural Arabidopsis allotetraploids, leading to extremely late flowering. Furthermore, FLC expression correlates positively with histone H3-Lys4 methylation and H3-Lys9 acetylation and negatively with H3-Lys9 methylation, epigenetic marks for gene activation and silencing. We provide evidence for interactive roles of regulatory sequence changes, chromatin modification, and trans-acting effects in natural selection of orthologous FLC loci, which determines the fate of duplicate genes and adaptation of allopolyploids during evolution.
Characterization of Drosophila mini-me, a Gene Required for Cell Proliferation and Survival
Authors: Jones Chonnettia , Reifegerste Rita , Moses Kevin
Published: March 2006

In the developing Drosophila eye, the morphogenetic furrow is a developmental organizing center for patterning and cell proliferation. The furrow acts both to limit eye size and to coordinate the number of cells to the number of facets. Here we report the molecular and functional characterization of Drosophila mini-me (mnm), a potential regulator of cell proliferation and survival in the developing eye. We first identified mnm as a dominant modifier of hedgehog loss-of-function in the developing eye. We report that mnm encodes a conserved protein with zinc knuckle and RING finger domains. We show that mnm is dispensable for patterning of the eye disc, but required in the eye for normal cell proliferation and survival. We also show that mnm null mutant cells exhibit altered cell cycle profiles and contain excess nucleic acid. Moreover, mnm overexpression can induce cells to proliferate and incorporate BrdU. Thus, our data implicate mnm as a regulator of mitotic progression during the proliferative phase of eye development, possibly through the control of nucleic acid metabolism.
Homologous Recombination Is Required for Genome Stability in the Absence of DOG-1 inCaenorhabditis elegans
Authors: Youds Jillian L , O'Neil Nigel J , Rose Ann M
Published: March 2006

In C. elegans, DOG-1 prevents deletions that initiate in polyG/polyC tracts (G/C tracts), most likely by unwinding secondary structures that can form in G/C tracts during lagging-strand DNA synthesis. We have used the dog-1 mutant to assay the in vivo contribution of various repair genes to the maintenance of G/C tracts. Here we show that DOG-1 and the BLM ortholog, HIM-6, act synergistically during replication; simultaneous loss of function of both genes results in replicative stress and an increase in the formation of small deletions that initiate in G/C tracts. Similarly, we demonstrate that the C. elegans orthologs of the homologous recombination repair genes BARD1, RAD51, and XPF and the trans-lesion synthesis polymerases polη and polκ contribute to the prevention of deletions in dog-1 mutants. Finally, we provide evidence that the small deletions generated in the dog-1 background are not formed through homologous recombination, nucleotide excision repair, or nonhomologous end-joining mechanisms, but appear to result from a mutagenic repair mechanism acting at G/C tracts. Our data support the hypothesis that absence of DOG-1 leads to replication fork stalling that can be repaired by deletion-free or deletion-prone mechanisms.
Site-Specific Transformation of Drosophila via ϕC31 Integrase-Mediated Cassette Exchange
Authors: Bateman Jack R , Lee Anne M , Wu C-ting
Published: March 2006

Position effects can complicate transgene analyses. This is especially true when comparing transgenes that have inserted randomly into different genomic positions and are therefore subject to varying position effects. Here, we introduce a method for the precise targeting of transgenic constructs to predetermined genomic sites in Drosophila using the ϕC31 integrase system in conjunction with recombinase-mediated cassette exchange (RMCE). We demonstrate the feasibility of this system using two donor cassettes, one carrying the yellow gene and the other carrying GFP. At all four genomic sites tested, we observed exchange of donor cassettes with an integrated target cassette carrying the mini-white gene. Furthermore, because RMCE-mediated integration of the donor cassette is necessarily accompanied by loss of the target cassette, we were able to identify integrants simply by the loss of mini-white eye color. Importantly, this feature of the technology will permit integration of unmarked constructs into Drosophila, even those lacking functional genes. Thus, ϕC31 integrase-mediated RMCE should greatly facilitate transgene analysis as well as permit new experimental designs.
A Novel Gain-of-Function Mutant of the Cyclic GMP-Dependent Protein Kinase egl-4 Affects Multiple Physiological Processes in Caenorhabditis elegans
Authors: Raizen David M , Cullison Kevin M , Pack Allan I , Sundaram Meera V
Published: March 2006

cGMP-dependent protein kinases are key intracellular transducers of cell signaling. We identified a novel dominant mutation in the C. elegans egl-4 cGMP-dependent protein kinase (PKG) and show that this mutation causes increased normal gene activity although it is associated with a reduced EGL-4 protein level. Prior phenotypic analyses of this gain-of-function mutant demonstrated a reduced longevity and a reduced feeding behavior when the animals were left unperturbed. We characterize several additional phenotypes caused by increased gene activity of egl-4. These phenotypes include a small body size, reduced locomotion in the presence of food, a pale intestine, increased intestinal fat storage, and a decreased propensity to form dauer larvae. The multiple phenotypes of egl-4 dominant mutants are consistent with an instructive signaling role of PKG to control many aspects of animal physiology. This is among the first reported gain-of-function mutations in this enzyme of central physiological importance. In a genetic screen we have identified extragenic suppressors of this gain-of-function mutant. Thus, this mutant promises to be a useful tool for identifying downstream targets of PKG.
A Dbf4p BRCA1 C-Terminal-Like Domain Required for the Response to Replication Fork Arrest in Budding Yeast
Authors: Gabrielse Carrie , Miller Charles T , McConnell Kristopher H , DeWard Aaron , Fox Catherine A , Weinreich Michael
Published: March 2006

Dbf4p is an essential regulatory subunit of the Cdc7p kinase required for the initiation of DNA replication. Cdc7p and Dbf4p orthologs have also been shown to function in the response to DNA damage. A previous Dbf4p multiple sequence alignment identified a conserved ∼40-residue N-terminal region with similarity to the BRCA1 C-terminal (BRCT) motif called “motif N.” BRCT motifs encode ∼100-amino-acid domains involved in the DNA damage response. We have identified an expanded and conserved ∼100-residue N-terminal region of Dbf4p that includes motif N but is capable of encoding a single BRCT-like domain. Dbf4p orthologs diverge from the BRCT motif at the C terminus but may encode a similar secondary structure in this region. We have therefore called this the BRCT and DBF4 similarity (BRDF) motif. The principal role of this Dbf4p motif was in the response to replication fork (RF) arrest; however, it was not required for cell cycle progression, activation of Cdc7p kinase activity, or interaction with the origin recognition complex (ORC) postulated to recruit Cdc7p–Dbf4p to origins. Rad53p likely directly phosphorylated Dbf4p in response to RF arrest and Dbf4p was required for Rad53p abundance. Rad53p and Dbf4p therefore cooperated to coordinate a robust cellular response to RF arrest.
The Distribution of Fitness Effects of New Deleterious Amino Acid Mutations in Humans
Authors: Eyre-Walker Adam , Woolfit Megan , Phelps Ted
Published: March 2006

The distribution of fitness effects of new mutations is a fundamental parameter in genetics. Here we present a new method by which the distribution can be estimated. The method is fairly robust to changes in population size and admixture, and it can be corrected for any residual effects if a model of the demography is available. We apply the method to extensively sampled single-nucleotide polymorphism data from humans and estimate the distribution of fitness effects for amino acid changing mutations. We show that a gamma distribution with a shape parameter of 0.23 provides a good fit to the data and we estimate that >50% of mutations are likely to have mild effects, such that they reduce fitness by between one one-thousandth and one-tenth. We also infer that <15% of new mutations are likely to have strongly deleterious effects. We estimate that on average a nonsynonymous mutation reduces fitness by a few percent and that the average strength of selection acting against a nonsynonymous polymorphism is ∼9 × 10−5. We argue that the relaxation of natural selection due to modern medicine and reduced variance in family size is not likely to lead to a rapid decline in genetic quality, but that it will be very difficult to locate most of the genes involved in complex genetic diseases.
Nongastric Marginal-Zone B-Cell MALT Lymphoma: Prognostic Value of Disease Dissemination
Authors: Arcaini Luca , Burcheri Sara , Rossi Andrea , Passamonti Francesco , Paulli Marco , Boveri Emanuela , Brusamolino Ercole , Orlandi Ester , Molteni Alfredo , Pulsoni Alessandro , Cox Maria Christina , Orsucci Lorella , Fabbri Alberto , Frezzato Maurizio , Voso Maria Teresa , Zaja Francesco , Montanari Francesca , Pascutto Cristiana , Morra Enrica , Cortelazzo Sergio , Lazzarino Mario
Published: March 2006

Learning Objectives

After completing this course, the reader will be able to: Discuss the different modalities of disease dissemination in nongastric marginal-zone B-cell MALT lymphoma.Explain the rationale for using extensive staging in nongastric marginal-zone B-cell MALT lymphoma.Assess the clinical features helpful to detect patients with nongastric marginal-zone B-cell MALT lymphoma who have a poor prognosis.

Access and take the CME test online and receive 1 AMA PRA category 1 credit at CME.TheOncologist.com
The Effect of Cyclic Aerobic–Anoxic Conditions on Biodegradation of Benzoate
Authors: Deniz Timur , Çınar Özer , Marques Ana C. , Grady C. P. Leslie
Published: March 2006

The response of a mixed microbial culture to cyclic aerobic and anoxic (denitrifying) conditions was studied in a chemostat with a 48‐hour hydraulic residence time receiving a feed containing benzoate and pyruvate. When the cyclic conditions were 3‐hour aerobic and 9‐hour anoxic, the bacteria‐degraded benzoate aerobically via the catechol 2,3‐dioxygenase (C23DO) pathway. The quantity of C23DO remained constant throughout the anoxic period but decreased during the initial portion of the aerobic period before returning to the level present in the anoxic period. Anoxic biodegradation of benzoate was via benzoyl‐CoA reductase, which remained constant regardless of the redox condition. The aerobic benzoate uptake capability (AeBUC) of the culture increased during the aerobic period but decreased during the anoxic period. The anoxic benzoate uptake capability (AnBUC) exhibited the opposite response. When the cycle was 6‐hour aerobic and 6‐hour anoxic, aerobic biodegradation of benzoate proceeded via the protocatechuate 4,5‐dioxygenase (P45DO) pathway. The P45DO activity decreased early in the aerobic period, but then increased to the level present during the anoxic period. The level of benzoyl‐CoA reductase was constant throughout the cycle. Furthermore, AeBUC and AnBUC responded in much the same way as in the 3/9‐hour chemostat. During a 9‐hour aerobic and 3‐hour anoxic cycle, the culture synthesized both P45DO and C23DO, with the former having significantly higher activity. Unlike the other two cycles, AeBUC changed little during the aerobic period, although AnBUC decreased. The culture was well‐adapted to the cyclic conditions as evidenced by the lack of accumulation of either substrate during any cycle tested. This suggests that cyclic aerobic–anoxic processes can be used in industrial wastewater‐treatment facilities receiving significant quantities of simple aromatic compounds like benzoate. However, the results showed that the kinetics of benzoate degradation were different under aerobic and anoxic conditions, a situation that must be considered when modeling cyclic bioreactors receiving aromatic compounds.
Removal of Low‐Concentration Phosphorus Using a Fluidized Raw Dolomite Bed
Authors: Ayoub George M. , Kalinian Houri
Published: March 2006

Raw dolomite powder, obtained from dolomitic rocks, was evaluated for its efficiency in removing low concentrations of phosphate present in various water and wastewater matrices. A variety of experimental setups and process variables were tested, in an attempt to determine the optimal scheme (fluidized bed) that will be used to accomplish the objectives of the study. Test influents, including distilled water (DW), synthetic groundwater (SGW), tap water (TW), and sodium‐hydroxide (NaOH)‐alkalized wastewater treated with liquid bittern (STSE‐B) and wastewater treated with lime (STSE‐L) were used to assess the effect of influent parameters on the quality of the effluent generated. The adsorptive behavior of dolomite was described by fitting data generated from the study into the Freundlich and Langmuir isotherm models. Regeneration of the dolomite was attempted by the use of an acid (hydrochloric acid), DW, and an alkaline (NaOH). Test results indicate remarkable phosphate removal levels for DW and SGW. Removal levels of 100% were attained, for an average of 307 and 314 bed volumes at inflow concentration levels of 0.28 and 0.34 mg phosphate (PO 4)/L, respectively. Relative adsorption capacities were calculated to be 0.06 and 0.072 mg PO 4/g of dolomite, respectively. For TW, STSE‐L, and STSE‐B, 100% removal before the start of the breakthrough was sustained for averages of 205, 94, and 28 bed volumes at phosphate dosages of 0.34, 0.56, and 0.6 mg PO 4/L, respectively. The calculated adsorptive capacities were 0.05, 0.051, and 0.025, respectively. Zeta potential measurement resulted in values of −16.0 mV before treatment and +3.0 mV after complete exhaustion of the bed with PO 4‐jacked SGW influent, indicating an ionic adsorption of ions of opposite charge to that of the particle surface. Although both the Freundlich and Langmuir isotherms were found to fit the sorption curves, the Langmuir seems to better describe the sorption process in dolomite. Among the three attempted dolomite regeneration processes, the use of NaOH has shown to be the most successful.
Regrowth of Fecal Coliforms in Class A Biosolids
Authors: Jolis Domènec
Published: March 2006

Eight types of Class A biosolids were tested for fecal coliform (FC) reactivation and/or regrowth at 20, 35, and 50°C for 21 days. Growth of FC did not occur at 20 or 50°C, but it was observed in two samples incubated at 35°C after a lag period of 48 hours. In undigested biosolids, final FC concentration exceeded 10 4 MPN/g, whereas in thermophilically digested biosolids, the final FC concentration remained below 10 3 MPN/g, as FC regrowth may have been affected by the presence of the anaerobic bacterial consortium responsible for the digestion process. Fecal‐coliform reactivation and regrowth within treatment plant operations seem unlikely but can occur in land application of biosolids.
Investigating Activated Sludge Flocs using Microanalytical Techniques: Demonstration of Environmental Scanning Electron Microscopy and Time‐of‐Flight Secondary Ion Mass Spectrometry for Wastewater Applications
Authors: Holbrook R. David , Wagner Matthew S. , Mahoney Christine M. , Wight Scott A.
Published: March 2006

Environmental scanning electron microscopy (ESEM) with an energy‐dispersive X‐ray spectrometer (EDS) and time‐of‐flight secondary ion mass spectrometry (ToF‐SIMS) were demonstrated to be useful analytical tools for investigating surface and bulk components of individual floc particles from both full‐ and bench‐scale activated sludge systems. Detailed surface imaging of various hydrated biological floc particles by ESEM revealed substantial differences in surface features between treatment systems, while EDS identified spatial differences in the iron and the aluminum distributions. The ToF‐SIMS spectra had signature fragments of protein and polysaccharide material from the floc surface, suggesting that this technique is capable of surface profiling extracellular polymeric substances. Principal‐component analysis of the positive ion ToF‐SIMS spectra from the mixed‐liquor‐suspended solid (MLSS) samples and reference aquatic organic materials found slight differences between the full‐ and bench‐scale MLSS surface properties but substantial differences among MLSS and treated effluent from the same facility.