bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantitative Genetic Analysis of copia Retrotransposon Activity in Inbred Drosophila melanogaster Lines
Authors: Nuzhdin Sergey V , Pasyukova Elena G , Morozova Elena A , Flavell Andrew J
Published: April 2021

The rates of transcription andtransposition of retrotransposons vary betweenlines of Drosophila melanogaster. We have studied the genetics of differences in copia retrotransposon activity by quantitative trait loci (QTL) mapping. Ninety-eight recombinant inbred lines were constructed from two parental lines exhibiting a 10-fold difference in copia transcript level and a 100-fold difference in transposition rate. The lines were scored for 126 molecular markers, copia transcript level, and rate of copia transposition. Transcript level correlated with copia copy number, and the difference in copia copy number between parental lines accounted for 45.1% of copia transcript-level difference. Most of the remaining difference was accounted for by two transcript-level QTL mapping to cytological positions 27B-30D and 50F-57C on the second chromosome, which accounted for 11.5 and 30.4%, respectively. copia transposition rate was controlled by interacting QTL mapping to the region 27B-48D on the second and 61A-65A and 97D-100A on the third chromosome. The genes controlling copia transcript level are thus not necessarily those involved in controlling copia transposition rate. Segregation of modifying genes, rather than mutations, might explain the variability in copia retrotransposon activity between lines.
The Correlation Between Synonymous and Nonsynonymous Substitutions in Drosophila: Mutation, Selection or Relaxed Constraints?
Authors: Comeron Josep M , Kreitman Martin
Published: April 2021

Codon usage bias, the preferential use of particular codons within each codon family, is characteristic of synonymous base composition in many species, including Drosophila, yeast, and many bacteria. Preferential usage of particular codons in these species is maintained by natural selection acting largely at the level of translation. In Drosophila, as in bacteria, the rate of synonymous substitution per site is negatively correlated with the degree of codon usage bias, indicating stronger selection on codon usage in genes with high codon bias than in genes with low codon bias. Surprisingly, in these organisms, as well as in mammals, the rate of synonymous substitution is also positively correlated with the rate of nonsynonymous substitution. To investigate this correlation, we carried out a phylogenetic analysis of substitutions in 22 genes between two species of Drosophila, Drosophila pseudoobscura and D. subobscura, in codons that differ by one replacement and one synonymous change. We provide evidence for a relative excess of double substitutions in the same species lineage that cannot be explained by the simultaneous mutation of two adjacent bases. The synonymous changes in these codons also cannot be explained by a shift to a more preferred codon following a replacement substitution. We, therefore, interpret the excess of double codon substitutions within a lineage as being the result of relaxed constraints on both kinds of substitutions in particular codons.
Characterization of Deleterious Mutations in Outcrossing Populations
Authors: Deng Hong-Wen
Published: April 2021

Deng and Lynch recently proposed estimating the rate and effects of deleterious genomic mutations from changes in the mean and genetic variance of fitness upon selfing/outcrossing in outcrossing/highly selfing populations. The utility of our original estimation approach is limited in outcrossing populations, since selfing may not always be feasible. Here we extend the approach to any form of inbreeding in outcrossing populations. By simulations, the statistical properties of the estimation under a common form of inbreeding (sib mating) are investigated under a range of biologically plausible situations. The efficiencies of different degrees of inbreeding and two different experimental designs of estimation are also investigated. We found that estimation using the total genetic variation in the inbred generation is generally more efficient than employing the genetic variation among the mean of inbred families, and that higher degree of inbreeding employed in experiments yields higher power for estimation. The simulation results of the magnitude and direction of estimation bias under variable or epistatic mutation effects may provide a basis for accurate inferences of deleterious mutations. Simulations accounting for environmental variance of fitness suggest that, under full-sib mating, our extension can achieve reasonably well an estimation with sample sizes of only ∼2000-3000.
Suppressors of Cdc25p Overexpression Identify Two Pathways That Influence the G2/M Checkpoint in Fission Yeast
Authors: Forbes Kristi Chrispell , Humphrey Timothy , Enoch Tamar
Published: April 2021

Checkpoints maintain the order of cell-cycle events. At G2/M, a checkpoint blocks mitosis in response to damaged or unreplicated DNA. There are significant differences in the checkpoint responses to damaged DNA and unreplicated DNA, although many of the same genes are involved in both responses. To identify new genes that function specifically in the DNA replication checkpoint pathway, we searched for high-copy suppressors of overproducer of Cdc25p (OPcdc25+), which lacks a DNA replication checkpoint. Two classes of suppressors were isolated. One class includes a new gene encoding a putative DEAD box helicase, suppressor of uncontrolled mitosis (sum3+). This gene negatively regulates the cell-cycle response to stress when overexpressed and restores the checkpoint response by a mechanism that is independent of Cdc2p tyrosine phosphorylation. The second class includes chk1+ and the two Schizosaccharomyces pombe 14-3-3 genes, rad24+ and rad25+, which appear to suppress the checkpoint defect by inhibiting Cdc25p. We show that rad24Δ mutants are defective in the checkpoint response to the DNA replication inhibitor hydroxyurea at 37° and that cds1Δ rad24Δ mutants, like cds1Δ chk1Δ mutants, are entirely checkpoint deficient at 29°. These results suggest that chk1+ and rad24+ may function redundantly with cds1+ in the checkpoint response to unreplicated DNA.
Regulators of Pseudohyphal Differentiation in Saccharomyces cerevisiae Identified Through Multicopy Suppressor Analysis in Ammonium Permease Mutant Strains
Authors: Lorenz Michael C , Heitman Joseph
Published: April 2021

Nitrogen-starved diploid cells of the yeast Saccharomyces cerevisiae differentiate into a filamentous, pseudohyphal growth form. Recognition of nitrogen starvation is mediated, at least in part, by the ammonium permease Mep2p and the Gα subunit Gpa2p. Genetic activation of the pheromone-responsive MAP kinase cascade, which is also required for filamentous growth, only weakly suppresses the filamentation defect of Δmep2/Δmep2 and Δgpa2/Δgpa2 strain. Surprisingly, deletion of Mep1p, an ammonium permease not previously thought to regulate differentiation, significantly enhances the potency of MAP kinase activation, such that the STE11-4 allele induces filamentation to near wild-type levels in Δmep1/Δmep1 Δmep2/Δmep2 and Δmep1/Δmep1 Δgpa2/Δgpa2 strains. To identify additional regulatory components, we isolated high-copy suppressors of the filamentation defect of the Δmep1/Δmep1 Δmep2/Δmep2 mutant. Multicopy expression of TEC1, PHD1, PHD2 (MSS10/MSN1/FUP4), MSN5, CDC6, MSS11, MGA1, SKN7, DOT6, HMS1, HMS2, or MEP2 each restored filamentation in a Δmep1/Δmep1 Δmep2/Δmep2 strain. Overexpression of SRK1 (SSD1), URE2, DAL80, MEP1, or MEP3 suppressed only the growth defect of the Δmep1/Δmep1 Δmep2/Δmep2 mutant strain. Characterization of these genes through deletion analysis and epistasis underscores the complexity of this developmental pathway and suggests that stress conditions other than nitrogen deprivation may also promote filamentous growth.
Transfer of a Supernumerary Chromosome Between Vegetatively Incompatible Biotypes of the Fungus Colletotrichum gloeosporioides
Authors: He Chaozu , Rusu Anca G , Poplawski Agnieszka M , Irwin John A G , Manners John M
Published: April 2021

Two biotypes (A and B) of Colletotrichum gloeosporioides infect the tropical legumes Stylosanthes spp. in Australia. These biotypes are asexual and vegetatively incompatible. However, field isolates of biotype B carrying a supernumerary 2-Mb chromosome, thought to originate from biotype A, have been reported previously. We tested the hypothesis that the 2-Mb chromosome could be transferred from biotype A to biotype B under laboratory conditions. Selectable marker genes conferring resistance to hygromycin and phleomycin were introduced into isolates of biotypes A and B, respectively. A transformant of biotype A, with the hygromycin resistance gene integrated on the 2-Mb chromosome, was cocultivated with phleomycin-resistant transformants of biotype B. Double antibiotic-resistant colonies were obtained from conidia of these mixed cultures at a frequency of approximately 10-7. Molecular analysis using RFLPs, RAPDs, and electrophoretic karyotypes showed that these colonies contained the 2-Mb chromosome in a biotype B genetic background. In contrast, no double antibiotic colonies developed from conidia obtained from mixed cultures of phleomycin-resistant transformants of biotype B with biotype A transformants carrying the hygromycin resistance gene integrated in chromosomes >2 Mb in size. The results demonstrated that the 2-Mb chromosome was selectively transferred from biotype A to biotype B. The horizontal transfer of specific chromosomes across vegetative incompatibility barriers may explain the origin of supernumerary chromosomes in fungi.
Detailed Alignment of Saccharum and Sorghum Chromosomes: Comparative Organization of Closely Related Diploid and Polyploid Genomes
Authors: Ming R , Liu S-C , Lin Y-R , da Silva J , Wilson W , Braga D , van Deynze A , Wenslaff T F , Wu K K , Moore P H , Burnquist W , Sorrells M E , Irvine J E , Paterson A H
Published: April 2021

The complex polyploid genomes of three Saccharum species have been aligned with the compact diploid genome of Sorghum (2n = 2x = 20). A set of 428 DNA probes from different Poaceae (grasses) detected 2460 loci in F1 progeny of the crosses Saccharum officinarum Green German × S. spontaneum IND 81-146, and S. spontaneum PIN 84-1 × S. officinarum Muntok Java. Thirty-one DNA probes detected 226 loci in S. officinarum LA Purple × S. robustum Molokai 5829. Genetic maps of the six Saccharum genotypes, including up to 72 linkage groups, were assembled into “homologous groups” based on parallel arrangements of duplicated loci. About 84% of the loci mapped by 242 common probes were homologous between Saccharum and Sorghum. Only one interchromosomal and two intrachromosomal rearrangements differentiated both S. officinarum and S. spontaneum from Sorghum, but 11 additional cases of chromosome structural polymorphism were found within Saccharum. Diploidization was advanced in S. robustum, incipient in S. officinarum, and absent in S. spontaneum, consistent with biogeographic data suggesting that S. robustum is the ancestor of S. officinarum, but raising new questions about the antiquity of S. spontaneum. The densely mapped Sorghum genome will be a valuable tool in ongoing molecular analysis of the complex Saccharum genome.
Meiotic Transmission Rates Correlate With Physical Features of Rearranged Centromeres in Maize
Authors: Kaszás Étienne , Birchler James A
Published: April 2021

The centromere of the maize B chromosome was used as a model to study the physical features of a functional centromere. Pulsed-field gel electrophoresis was previously used to determine the organization of a repetitive sequence (referred to as the B-specific repeat) localized in the centromeric region of the maize B chromosome. The centromere is composed mostly of this repeat. In this report, a collection of 25 B chromosome derivatives that suffered from misdivision of the centromere was examined for the content and organization of the B repeat. Meiotic transmission of these derivatives was also determined and compared with rearrangements within the centromere. This analysis revealed that there is a strong correlation between the size of the centromere and meiotic transmission. In addition, the loss of a particular PmeI fragment of 370 kb considerably reduced meiotic transmission. This sequence contains a 55-kb EcoRI fragment that is also present in all but four derivatives. Because the centromere of the maize B chromosome can be divided by successive misdivisions to derivatives with centromeres of <300 kb, it should be possible for artificial chromosomes to be produced in maize.
A New Approach to the Problem of Multiple Comparisons in the Genetic Dissection of Complex Traits
Authors: Weller Joel Ira , Song Jiu Zhou , Heyen David W , Lewin Harris A , Ron Micha
Published: April 2021

Saturated genetic marker maps are being used to map individual genes affecting quantitative traits. Controlling the “experimentwise” type-I error severely lowers power to detect segregating loci. For preliminary genome scans, we propose controlling the “false discovery rate,” that is, the expected proportion of true null hypotheses within the class of rejected null hypotheses. Examples are given based on a granddaughter design analysis of dairy cattle and simulated backcross populations. By controlling the false discovery rate, power to detect true effects is not dependent on the number of tests performed. If no detectable genes are segregating, controlling the false discovery rate is equivalent to controlling the experimentwise error rate. If quantitative loci are segregating in the population, statistical power is increased as compared to control of the experimentwise type-I error. The difference between the two criteria increases with the increase in the number of false null hypotheses. The false discovery rate can be controlled at the same level whether the complete genome or only part of it has been analyzed. Additional levels of contrasts, such as multiple traits or pedigrees, can be handled without the necessity of a proportional decrease in the critical test probability.
Natural Selection on Synonymous Sites Is Correlated With Gene Length and Recombination in Drosophila
Authors: Comeron Josep M , Kreitman Martin , Aguadé Montserrat
Published: April 2021

Evolutionary analysis of codon bias in Drosophila indicates that synonymous mutations are not neutral, but rather are subject to weak selection at the translation level. Here we show that the effectiveness of natural selection on synonymous sites is strongly correlated with the rate of recombination, in accord with the nearly neutral hypothesis. This correlation, however, is apparent only in genes encoding short proteins. Long coding regions have both a lower codon bias and higher synonymous substitution rates, suggesting that they are affected less efficiently by selection. Therefore, both the length of the coding region and the recombination rate modulate codon bias. In addition, the data indicate that selection coefficients for synonymous mutations must vary by a minimum of one or two orders of magnitude. Two hypotheses are proposed to explain the relationship among the coding region length, the codon bias, and the synonymous divergence and polymorphism levels across the range of recombination rates in Drosophila. The first hypothesis is that selection coefficients on synonymous mutations are inversely related to the total length of the coding region. The second hypothesis proposes that interference among synonymous mutations reduces the efficacy of selection on these mutations. We investigated this second hypothesis by carrying out forward simulations of weakly selected mutations in model populations. These simulations show that even with realistic recombination rates, this interference, which we call the “small-scale” Hill-Robertson effect, can have a moderately strong influence on codon bias.
A Consensus Map for Loblolly Pine (Pinus taeda L.). I. Construction and Integration of Individual Linkage Maps From Two Outbred Three-Generation Pedigrees
Authors: Sewell Mitchell M , Sherman Bradley K , Neale David B
Published: April 2021

A consensus map for loblolly pine (Pinus taeda L.) was constructed from the integration of linkage data from two unrelated three-generation outbred pedigrees. The progeny segregation data from restriction fragment length polymorphism, random amplified polymorphic DNA, and isozyme genetic markers from each pedigree were recoded to reflect the two independent populations of parental meioses, and genetic maps were constructed to represent each parent. The rate of meiotic recombination was significantly greater for males than females, as was the average estimate of genome length for males {1983.7 cM [Kosambi mapping function (K)]} and females [1339.5 cM(K)]. The integration of individual maps allows for the synthesis of genetic information from independent sources onto a single consensus map and facilitates the consolidation of linkage groups to represent the chromosomes (n = 12) of loblolly pine. The resulting consensus map consists of 357 unique molecular markers and covers ∼1300 cM(K).
Escherichia coli MutM Suppresses Illegitimate Recombination Induced by Oxidative Stress
Authors: Onda Masaaki , Hanada Katsuhiro , Kawachi Hirokazu , Ikeda Hideo
Published: April 2021

DNA damage by oxidative stress is one of the causes of mutagenesis. However, whether or not DNA damage induces illegitimate recombination has not been determined. To study the effect of oxidative stress on illegitimate recombination, we examined the frequency of λbio transducing phage in the presence of hydrogen peroxide and found that this reagent enhances illegitimate recombination. To clarify the types of illegitimate recombination, we examined the effect of mutations in mutM and related genes on the process. The frequency of λbio transducing phage was 5- to 12-fold higher in the mutM mutant than in the wild type, while the frequency in the mutY and mutT mutants was comparable to that of the wild type. Because 7,8-dihydro-8-oxoguanine (8-oxoG) and formamido pyrimidine (Fapy) lesions can be removed from DNA by MutM protein, these lesions are thought to induce illegitimate recombination. Analysis of recombination junctions showed that the recombination at Hotspot I accounts for 22 or 4% of total λbio transducing phages in the wild type or in the mutM mutant, respectively. The preferential increase of recombination at nonhotspot sites with hydrogen peroxide in the mutM mutant was discussed on the basis of a new model, in which 8-oxoG and/or Fapy residues may introduce double-strand breaks into DNA.
An Allele of RFA1 Suppresses RAD52-Dependent Double-Strand Break Repair in Saccharomyces cerevisiae
Authors: Smith Julianne , Rothstein Rodney
Published: April 2021

An allele of RFA1, the largest subunit of the single-stranded DNA-binding complex RP-A, was identified as a suppressor of decreased direct-repeat recombination in rad1 rad52 double mutants. In this study, we used two LEU2 direct-repeat assays to investigate the mechanism by which the rfa1-D228Y allele increases recombination. We found that both intrachromatid and sister chromatid recombination are stimulated in rfa1-D228Y strains. In a rad1 rad52 background, however, the majority of the increased recombination is caused by stimulation of deletion events by an intrachromatid recombination mechanism that is likely to be single-strand annealing. Studies in which an HO endonuclease cut was introduced between the two leu2 copies indicate that the rfa1-D228Y mutation partially suppresses the rad52 defect in recovering recombination products. Furthermore, molecular analysis of processing and product formation kinetics reveals that, in a rad52 background, the rfa1-D228Y mutation results in increased levels of recombinant products and the disappearance of large single-stranded intermediates characteristic of rad52 strains. On the basis of these results, we propose that in the absence of wild-type Rad52, the interaction of RP-A with single-stranded DNA inhibits strand annealing, and that this inhibition is overcome by the rfa1-D228Y mutation.
Molecular Characterization of tol, a Mediator of Mating-Type-Associated Vegetative Incompatibility in Neurospora crassa
Authors: Shiu Patrick Ka Tai , Glass N Louise
Published: April 2021

The mating-type locus in the haploid filamentous fungus, Neurospora crassa, controls mating and sexual development. The fusion of reproductive structures of opposite mating type, A and a, is required to initiate sexual reproduction. However, the fusion of hyphae of opposite mating type during vegetative growth results in growth inhibition and cell death, a process that is mediated by the tol locus. Mutations in tol are recessive and suppress mating-type-associated heterokaryon incompatibility. In this study, we describe the cloning and characterization of tol. The tol gene encodes a putative 1011-amino-acid polypeptide with a coiled-coil domain and a leucine-rich repeat. Both regions are required for tol activity. Repeat-induced point mutations in tol result in mutants that are wild type during vegetative growth and sexual reproduction, but that allow opposite mating-type individuals to form a vigorous heterokaryon. Transcript analyses show that tol mRNA is present during vegetative growth but absent during a cross. These data suggest that tol transcription is repressed to allow the coexistence of opposite mating-type nuclei during the sexual reproductive phase. tol is expressed in a mat A, mat a, A/a partial diploid and in a mating-type deletion strain, indicating that MAT A-1 and MAT a-1 are not absolutely required for transcription or repression of tol. These data suggest that TOL may rather interact with MAT A-1 and/or MAT a-1 (or downstream products) to form a death-triggering complex.
Genetic and Molecular Analysis of fox-1, a Numerator Element Involved in Caenorhabditis elegans Primary Sex Determination
Authors: Skipper Magdalena , Milne Catherine A , Hodgkin Jonathan
Published: April 2021

fox-1 was previously identified as a candidate numerator element based on its overexpression phenotype. FOX-1 is an RRM-type RNA-binding protein, which can bind RNAs in vitro. Western analysis detects FOX-1 throughout development. fox-1::lacZ comes on ubiquitously early during embryogenesis. Postembryonically, fox-1::lacZ is expressed sex specifically in a subset of cells in the head and tail. We describe a Tc1derived deletion allele [fox-1(Δ)] that removes the RRM domain. fox-1(Δ) confers no phenotype in XXs, but can rescue XO-specific lethality and feminization caused by duplications of the left end of the X. fox-1(Δ) synergizes with putative numerators, resulting in abnormal XX development. Genetic analysis indicated that fox-1(Δ) leads to a slight increase in xol-1 activity, while fox-1(gf) leads to partial loss of xol-1 activity, and xol-1 is epistatic to fox-1. RNase protection experiments revealed increased levels of the 2.2-kb xol-1 message in fox-1(Δ) animals, and reduced levels in fox-1(gf) animals. Additionally, fox-1(Δ) impairs male mating efficiency, which, we propose, represents another function of fox-1, independent of xol-1 and its role in sex determination.
An Analysis of Transvection at the yellow Locus of Drosophila melanogaster
Authors: Morris James R , Chen Ji-long , Filandrinos Stephen T , Dunn Rebecca C , Fisk Ridgely , Geyer Pamela K , Wu Chao-ting
Published: April 2021

Studies of a wide variety of organisms have shown that homologous sequences can exert a significant impact on each other, resulting in changes in gene sequence, gene expression, chromatin structure, and global chromosome architecture. Our work has focused on transvection, a process that can cause genes to be sensitive to the proximity of a homologue. Transvection is seen at the yellow gene of Drosophila, where it mediates numerous cases of intragenic complementation. In this article, we describe two approaches that have characterized the process of transvection at yellow. The first entailed a screen for mutations that support intragenic complementation at yellow. The second involved the analysis of 53 yellow alleles, obtained from a variety of sources, with respect to complementation, molecular structure, and transcriptional competence. Our data suggest two ways in which transvection may be regulated at yellow: (1) a transcriptional mechanism, whereby the ability of an allele to support transvection is influenced by its transcriptional competency, and (2) a structural mechanism, whereby the pairing of structurally dissimilar homologues results in conformational changes that affect gene expression.
Identification of Genes Controlling Malpighian Tubule and Other Epithelial Morphogenesis in Drosophila melanogaster
Authors: Liu Xuejun , Kiss István , Lengyel Judith A
Published: April 2021

The Drosophila Malpighian tubule is a model system for studying genetic mechanisms that control epithelial morphogenesis. From a screen of 1800 second chromosome lethal lines, by observing uric acid deposits in unfixed inviable embryos, we identified five previously described genes (barr, fas, flb, raw, and thr) and one novel gene, walrus (wal), that affect Malpighian tubule morphogenesis. Phenotypic analysis of these mutant embryos allows us to place these genes, along with other previously described genes, into a genetic pathway that controls Malpighian tubule development. Specifically, wal affects evagination of the Malpighian tubule buds, fas and thr affect bud extension, and barr, flb, raw, and thr affect tubule elongation. In addition, these genes were found to have different effects on development of other epithelial structures, such as foregut and hindgut morphogenesis. Finally, from the same screen, we identified a second novel gene, drumstick, that affects only foregut and hindgut morphogenesis.
A Genetic Screen for Modifiers of Drosophila Src42A Identifies Mutations in Egfr, rolled and a Novel Signaling Gene
Authors: Zhang Qian , Zheng Qingxia , Lu Xiangyi
Published: April 2021

Drosophila Src42A, a close relative of the vertebrate c-Src, has been implicated in the Ras-Mapk signaling cascade. An allele of Src42A, Su(Raf)1, dominantly suppresses the lethality of partial loss-of-function Raf mutations. To isolate genes involved in the same pathway where Src42A functions, we carried out genetic screens for dominant suppressor mutations that prevented Su(Raf)1 from suppressing Raf. Thirty-six mutations representing at least five genetic loci were recovered from the second chromosome. These are Drosophila EGF Receptor (Egfr), rolled, Src42A, and two other new loci, one of which was named semang (sag). During embryogenesis, sag affects the development of the head, tail, and tracheal branches, suggesting that it participates in the pathways of Torso and DFGF-R1 receptor tyrosine kinases. sag also disrupts the embryonic peripheral nervous system. During the development of imaginal discs, sag affects two processes known to require Egfr signaling: the recruitment of photoreceptor cells and wing vein formation. Thus sag functions in several receptor tyrosine kinase (RTK)-mediated processes. In addition, sag dominantly enhances the phenotypes associated with loss-of-function Raf and rl, but suppresses those of activated Ras1V12 mutation. This work provides the first genetic evidence that both Src42A and sag are modulators of RTK signaling.
Intragenic Sex-Chromosomal Crossovers of Xmrk Oncogene Alleles Affect Pigment Pattern Formation and the Severity of Melanoma in Xiphophorus
Authors: Gutbrod Heidrun , Schartl Manfred
Published: April 2021

The X and Y chromosomes of the platyfish (Xiphophorus maculatus) contain a region that encodes several important traits, including the determination of sex, pigment pattern formation, and predisposition to develop malignant melanoma. Several sex-chromosomal crossovers were identified in this region. As the melanoma-inducing oncogene Xmrk is the only molecularly identified constituent, its genomic organization on both sex chromosomes was analyzed in detail. Using X and Y allele-specific sequence differences a high proportion of the crossovers was found to be intragenic in the oncogene Xmrk, concentrating in the extracellular domain-encoding region. The genetic and molecular data allowed establishment of an order of loci over ∼0.6 cM. It further revealed a sequence located within several kilobases of the extracellular domain-encoding region of Xmrk that regulates overexpression of the oncogene.
A Second-Generation Genetic Linkage Map of the Domestic Dog, Canis familiaris
Authors: Neff Mark W , Broman Karl W , Mellersh Cathryn S , Ray Kunal , Acland Gregory M , Aguirre Gustavo D , Ziegle Janet S , Ostrander Elaine A , Rine Jasper
Published: April 2021

Purebred strains, pronounced phenotypic variation, and a high incidence of heritable disease make the domestic dog uniquely suited to complement genetic analyses in humans and mice. A comprehensive genetic linkage map would afford many opportunities in dogs, ranging from the positional cloning of disease genes to the dissection of quantitative differences in size, shape, and behavior. Here we report a canine linkage map with the number of mapped loci expanded to 276 and 10-cM coverage extended to 75–90% of the genome. Most of the 38 canine autosomes are likely represented in the collection of 39 autosomal linkage groups. Eight markers were sufficiently informative to detect linkage at distances of 10–13 cM, yet remained unlinked to any other marker. Taken together, the results suggested a genome size of about 27 M. As in other species, the genetic length varied between sexes, with the female autosomal distance being ∼1.4-fold greater than that of male meioses. Fifteen markers anchored well-described genes on the map, thereby serving as landmarks for comparative mapping in dogs. We discuss the utility of the current map and outline steps necessary for future map improvement.
Transposition of the Autonomous Fot1 Element in the Filamentous Fungus Fusarium oxysporum
Authors: Migheli Quirico , Laugé Richard , Davière Jean-Michel , Gerlinger Catherine , Kaper Fiona , Langin Thierry , Daboussi Marie-Josée
Published: April 2021

Autonomous mobility of different copies of the Fot1 element was determined for several strains of the fungal plant pathogen Fusarium oxysporum to develop a transposon tagging system. Two Fot1 copies inserted into the third intron of the nitrate reductase structural gene (niaD) were separately introduced into two genetic backgrounds devoid of endogenous Fot1 elements. Mobility of these copies was observed through a phenotypic assay for excision based on the restoration of nitrate reductase activity. Inactivation of the Fot1 transposase open reading frame (frameshift, deletion, or disruption) prevented excision in strains free of Fot1 elements. Molecular analysis of the Nia+ revertant strains showed that the Fot1 element reintegrated frequently into new genomic sites after excision and that it can transpose from the introduced niaD gene into a different chromosome. Sequence analysis of several Fot1 excision sites revealed the socalled footprint left by this transposable element. Three reinserted Fot1 elements were cloned and the DNA sequences flanking the transposon were determined using inverse polymerase chain reaction. In all cases, the transposon was inserted into a TA dinucleotide and created the characteristic TA target site duplication. The availability of autonomous Fot1 copies will now permit the development of an efficient two-component transposon tagging system comprising a trans-activator element supplying transposase and a cis-responsive marked element.
Major Histocompatibility Complex Differentiation in Sacramento River Chinook Salmon
Authors: Kim Timothy J , Parker Karen M , Hedrick Philip W
Published: April 2021

The chinook salmon of the Sacramento River, California, have been reduced to a fraction of their former abundance because of human impact and use of the river system. Here we examine the genetic variation at a major histocompatibility complex class II exon in the four Sacramento chinook salmon runs. Examination of the alleles found in these and other chinook salmon revealed nucleotide patterns consistent with selection for amino acid replacement at the putative antigen-binding sites. We found a significant amount of variation in each of the runs, including the federally endangered winter run. All of the samples were in Hardy-Weinberg proportions. A significant amount of genetic differentiation between runs was revealed by several measures of differentiation. Winter run was the most genetically divergent, while the spring, late-fall, and fall runs were less differentiated.
Suppression of Intrachromosomal Gene Conversion in Mammalian Cells by Small Degrees of Sequence Divergence
Authors: Lukacsovich Tamas , Waldman Alan S
Published: April 2021

Pairs of closely linked defective herpes simplex virus (HSV) thymidine kinase (tk) gene sequences exhibiting various nucleotide heterologies were introduced into the genome of mouse Ltk– cells. Recombination events were recovered by selecting for the correction of a 16-bp insertion mutation in one of the tk sequences. We had previously shown that when two tk sequences shared a region of 232 bp of homology, interruption of the homology by two single nucleotide heterologies placed 19 bp apart reduced recombination nearly 20-fold. We now report that either one of the nucleotide heterologies alone reduces recombination only about 2.5-fold, indicating that the original pair of single nucleotide heterologies acted synergistically to inhibit recombination. We tested a variety of pairs of single nucleotide heterologies and determined that they reduced recombination from 7- to 175-fold. Substrates potentially leading to G-G or C-C mispairs in presumptive heteroduplex DNA (hDNA) intermediates displayed a particularly low rate of recombination. Additional experiments suggested that increased sequence divergence causes a shortening of gene conversion tracts. Collectively, our results suggest that suppression of recombination between diverged sequences is mediated via processing of a mispaired hDNA intermediate.
Molecular Characterization of Mutant Alleles of the DNA Repair/Basal Transcription Factor haywire/ERCC3 in Drosophila
Authors: Mounkes Leslie C , Fuller Margaret T
Published: April 2021

The haywire gene of Drosophila encodes a putative helicase essential for transcription and nucleotide excision repair. A haywire allele encoding a dominant acting poison product, lethal alleles, and viable but UV-sensitive alleles isolated as revertants of the dominant acting poison allele were molecularly characterized. Sequence analysis of lethal haywire alleles revealed the importance of the nucleotide-binding domain, suggesting an essential role for ATPase activity. The viable haync2 allele, which encodes a poison product, has a single amino acid change in conserved helicase domain VI. This mutation results in accumulation of a 68-kD polypeptide that is much more abundant than the wild-type haywire protein.
Two Loci, Tmevp2 and Tmevp3, Located on the Telomeric Region of Chromosome 10, Control the Persistence of Theiler’s Virus in the Central Nervous System of Mice
Authors: Bihl F , Brahic M , Bureau J F
Published: April 2021

Theiler’s virus persistently infects the white matter of the spinal cord in susceptible strains of mice. This infection is associated with inflammation and primary demyelination and is studied as a model of multiple sclerosis. The H-2D gene is the major gene controlling viral persistence. However, the SJL/J strain is more susceptible than predicted by its H-2s haplotype. An (SJL/J × B10.S)F1 × B10.S backcross was analyzed, and one quantitative trait locus (QTL) was located in the telomeric region of chromosome 10 close to the Ifng locus. Another one was tentatively mapped to the telomeric region of chromosome 18, close to the Mbp locus. We now report the study of 14 congenic lines that carry different segments of these two chromosomes. Although the presence of a QTL on chromosome 18 was not confirmed, two loci controlling viral persistence were identified on chromosome 10 and named Tmevp2 and Tmevp3. Furthermore, the Ifng gene was excluded from the regions containing Tmevp2 and Tmevp3. Analysis of the mode of inheritance of Tmevp2 and Tmevp3 identified an effect of sex, with males being more infected than females.
Identification of Three Putative Signal Transduction Genes Involved in R Gene-Specified Disease Resistance in Arabidopsis
Authors: Warren Randall F , Merritt Peter M , Holub Eric , Innes Roger W
Published: April 2021

The RPS5 disease resistance gene of Arabidopsis mediates recognition of Pseudomonas syringae strains that possess the avirulence gene avrPphB. By screening for loss of RPS5-specified resistance, we identified five pbs (avrPphB susceptible) mutants that represent three different genes. Mutations in PBS1 completely blocked RPS5-mediated resistance, but had little to no effect on resistance specified by other disease resistance genes, suggesting that PBS1 facilitates recognition of the avrPphB protein. The pbs2 mutation dramatically reduced resistance mediated by the RPS5 and RPM1 resistance genes, but had no detectable effect on resistance mediated by RPS4 and had an intermediate effect on RPS2-mediated resistance. The pbs2 mutation also had varying effects on resistance mediated by seven different RPP (recognition of Peronospora parasitica) genes. These data indicate that the PBS2 protein functions in a pathway that is important only to a subset of disease-resistance genes. The pbs3 mutation partially suppressed all four P. syringae-resistance genes (RPS5, RPM1, RPS2, and RPS4), and it had weak-to-intermediate effects on the RPP genes. In addition, the pbs3 mutant allowed higher bacterial growth in response to a virulent strain of P. syringae, indicating that the PBS3 gene product functions in a pathway involved in restricting the spread of both virulent and avirulent pathogens. The pbs mutations are recessive and have been mapped to chromosomes I (pbs2) and V (pbs1 and pbs3).
Evidence for the Involvement of the Glc7-Reg1 Phosphatase and the Snf1-Snf4 Kinase in the Regulation of INO1 Transcription in Saccharomyces cerevisiae
Authors: Shirra Margaret K , Arndt Karen M
Published: April 2021

Binding of the TATA-binding protein (TBP) to the promoter is a pivotal step in RNA polymerase II transcription. To identify factors that regulate TBP, we selected for suppressors of a TBP mutant that exhibits promoter-specific defects in activated transcription in vivo and severely reduced affinity for TATA boxes in vitro. Dominant mutations in SNF4 and recessive mutations in REG1, OPI1, and RTF2 were isolated that specifically suppress the inositol auxotrophy of the TBP mutant strains. OPI1 encodes a repressor of INO1 transcription. REG1 and SNF4 encode regulators of the Glc7 phosphatase and Snf1 kinase, respectively, and have well-studied roles in glucose repression. In two-hybrid assays, one SNF4 mutation enhances the interaction between Snf4 and Snf1. Suppression of the TBP mutant by our reg1 and SNF4 mutations appears unrelated to glucose repression, since these mutations do not alleviate repression of SUC2, and glucose levels have little effect on INO1 transcription. Moreover, mutations in TUP1, SSN6, and GLC7, but not HXK2 and MIG1, can cause suppression. Our data suggest that association of TBP with the TATA box may be regulated, directly or indirectly, by a substrate of Snf1. Analysis of INO1 transcription in various mutant strains suggests that this substrate is distinct from Opi1.
Pairing-Dependent Mislocalization of a Drosophila brown Gene Reporter to a Heterochromatic Environment
Authors: Sass Georgette L , Henikoff Steven
Published: April 2021

We describe the precise positioning of a reporter gene within heterochromatin where it may be silenced. A transposition of the 59E-60A region into pericentric heterochromatin ensnares distal 59E-60A via somatic pairing. The frequency with which a brown (bw) reporter gene in 59E is silenced is influenced by chromosomal configurations. Silencing occurs only when the bw+ reporter is unpaired due to heterozygosity with a deficiency, where the frequency of bw+ reporter expression is correlated with the extent of bw gene and flanking sequence present. Surprisingly, the frequency of pairing between the transposition in heterochromatin and distal 59E observed cytologically is indistinguishable from the frequency of pairing of homologous chromosomes at 59E in wild-type larval brains, regardless of configuration. Therefore, bringing a susceptible reporter gene into close proximity with heterochromatin does not necessarily affect its expression, but local pairing changes resulting from altered chromosomal configurations can lead to silencing. We also find that an ensnared distal copy of bw that is interrupted by a heterochromatic insertion enhances silencing. This demonstrates that bw can be simultaneously acted upon by pericentric and distal blocks of heterochromatin.
A Mutational Analysis of Leaf Morphogenesis in Arabidopsis thaliana
Authors: Berná Genoveva , Robles Pedro , Micol José Luis
Published: April 2021

As a contribution to a better understanding of the developmental processes that are specific to plants, we have begun a genetic analysis of leaf ontogeny in the model system Arabidopsis thaliana by performing a large-scale screening for mutants with abnormal leaves. After screening 46,159 M2 individuals, arising from 5770 M1 parental seeds exposed to EMS, we isolated 1926 M2 putative leaf mutants, 853 of which yielded viable M3 inbred progeny. Mutant phenotypes were transmitted with complete penetrance and small variations in expressivity in 255 lines. Most of them were inherited as recessive monogenic traits, belonging to 94 complementation groups, which suggests that we did not reach saturation of the genome. We discuss the nature of the processes presumably perturbed in the phenotypic classes defined among our mutants.
A Maize Map Standard With Sequenced Core Markers, Grass Genome Reference Points and 932 Expressed Sequence Tagged Sites (ESTs) in a 1736-Locus Map
Authors: Davis G L , McMullen M D , Baysdorfer C , Musket T , Grant D , Staebell M , Xu G , Polacco M , Koster L , Melia-Hancock S , Houchins K , Chao S , Coe E H
Published: April 2021

We have constructed a 1736-locus maize genome map containing1156 loci probed by cDNAs, 545 probed by random genomic clones, 16 by simple sequence repeats (SSRs), 14 by isozymes, and 5 by anonymous clones. Sequence information is available for 56% of the loci with 66% of the sequenced loci assigned functions. A total of 596 new ESTs were mapped from a B73 library of 5-wk-old shoots. The map contains 237 loci probed by barley, oat, wheat, rice, or tripsacum clones, which serve as grass genome reference points in comparisons between maize and other grass maps. Ninety core markers selected for low copy number, high polymorphism, and even spacing along the chromosome delineate the 100 bins on the map. The average bin size is 17 cM. Use of bin assignments enables comparison among different maize mapping populations and experiments including those involving cytogenetic stocks, mutants, or quantitative trait loci. Integration of nonmaize markers in the map extends the resources available for gene discovery beyond the boundaries of maize mapping information into the expanse of map, sequence, and phenotype information from other grass species. This map provides a foundation for numerous basic and applied investigations including studies of gene organization, gene and genome evolution, targeted cloning, and dissection of complex traits.
Archaeal DNA Replication: Identifying the Pieces to Solve a Puzzle
Authors: Cann Isaac K O , Ishino Yoshizumi
Published: April 2021

Archaeal organisms are currently recognized as very exciting and useful experimental materials. A major challenge to molecular biologists studying the biology of Archaea is their DNA replication mechanism. Undoubtedly, a full understanding of DNA replication in Archaea requires the identification of all the proteins involved. In each of four completely sequenced genomes, only one DNA polymerase (Pol BI proposed in this review from family B enzyme) was reported. This observation suggested that either a single DNA polymerase performs the task of replicating the genome and repairing the mutations or these genomes contain other DNA polymerases that cannot be identified by amino acid sequence. Recently, a heterodimeric DNA polymerase (Pol II, or Pol D as proposed in this review) was discovered in the hyperthermophilic archaeon, Pyrococcus furiosus. The genes coding for DP1 and DP2, the subunits of this DNA polymerase, are highly conserved in the Euryarchaeota. Euryarchaeotic DP1, the small subunit of Pol II (Pol D), has sequence similarity with the small subunit of eukaryotic DNA polymerase δ. DP2 protein, the large subunit of Pol II (Pol D), seems to be a catalytic subunit. Despite possessing an excellent primer extension ability in vitro, Pol II (Pol D) may yet require accessory proteins to perform all of its functions in euryarchaeotic cells. This review summarizes our present knowledge about archaeal DNA polymerases and their relationship with those accessory proteins, which were predicted from the genome sequences.
Archaeal Aminoacyl-tRNA Synthesis: Diversity Replaces Dogma
Authors: Tumbula Debra , Vothknecht Ute C , Kim Hyun-soo , Ibba Michael , Min Bokkee , Li Tong , Pelaschier Joanne , Stathopoulos Constantinos , Becker Hubert , Söll Dieter
Published: April 2021

Accurate aminoacyl-tRNA synthesis is essential for faithful translation of the genetic code and consequently has been intensively studied for over three decades. Until recently, the study of aminoacyl-tRNA synthesis in archaea had received little attention. However, as in so many areas of molecular biology, the advent of archaeal genome sequencing has now drawn researchers to this field. Investigations with archaea have already led to the discovery of novel pathways and enzymes for the synthesis of numerous aminoacyl-tRNAs. The most surprising of these findings has been a transamidation pathway for the synthesis of asparaginyl-tRNA and a novel lysyl-tRNA synthetase. In addition, seryl- and phenylalanyl-tRNA synthetases that are only marginally related to known examples outside the archaea have been characterized, and the mechanism of cysteinyl-tRNA formation in Methanococcus jannaschii and Methanobacterium thermoautotrophicum is still unknown. These results have revealed completely unexpected levels of complexity and diversity, questioning the notion that aminoacyl-tRNA synthesis is one of the most conserved functions in gene expression. It has now become clear that the distribution of the various mechanisms of aminoacyl-tRNA synthesis in extant organisms has been determined by numerous gene transfer events, indicating that, while the process of protein biosynthesis is orthologous, its constituents are not.
Genetic Diversity of Archaea in Deep-Sea Hydrothermal Vent Environments
Authors: Takai Ken , Horikoshi Koki
Published: April 2021

Molecular phylogenetic analysis of naturally occurring archaeal communities in deep-sea hydrothermal vent environments was carried out by PCR-mediated small subunit rRNA gene (SSU rDNA) sequencing. As determined through partial sequencing of rDNA clones amplified with archaea-specific primers, the archaeal populations in deep-sea hydrothermal vent environments showed a great genetic diversity, and most members of these populations appeared to be uncultivated and unidentified organisms. In the phylogenetic analysis, a number of rDNA sequences obtained from deep-sea hydrothermal vents were placed in deep lineages of the crenarchaeotic phylum prior to the divergence of cultivated thermophilic members of the crenarchaeota or between thermophilic members of the euryarchaeota and members of the methanogen-halophile clade. Whole cell in situ hybridization analysis suggested that some microorganisms of novel phylotypes predicted by molecular phylogenetic analysis were likely present in deep-sea hydrothermal vent environments. These findings expand our view of the genetic diversity of archaea in deep-sea hydrothermal vent environments and of the phylogenetic organization of archaea.