bluuebunny/crossref_metadata_2025 huggingface dataset:
Cha4p of Saccharomyces cerevisiae Activates Transcription Via Serine/Threonine Response Elements
Authors: Holmberg Steen , Schjerling Peter
Published: January 2021

The CHA1 gene of Saccharomyces cereuisiae encodes the catabolic l-serine (l-threonine) deaminase responsible for the utilization of serine/threonine as nitrogen sources. Previously, we identified two serine/threonine response elements in the CHA1 promoter, UASCHA. We report isolation of a mutation, cha4-1, that impairs serine/threonine induction of CHA1 transcription. The cha4-l allele causes noninducibility of a CHA1p-lacZ translational gene fusion, indicating that Cha4p exerts its action through the CHA1 promoter. Molecular and genetic mapping positioned the cha4 locus 17 cM centromere proximal to put1 on chromosome XII. The coding region of CHA4 predicts a 648-amino acid protein with a DNA-binding motif (residues 43–70) belonging to the Cys6 zinc cluster class. Gel retardation employing a recombinant peptide, Cha4p1-174, demonstrated that the peptide in vitro specifically binds UASCHA. Binding is abolished by a G-C to T-A mutation in the middle bases of the two CEZ-elements in UASCHA. The transcriptional activating ability of UASCHA derivatives in vivo correlates with their ability to bind Cha4p1-174  in vitro. We conclude that Cha4p is a positive regulator of CHA1 transcription and that Cha4p alone, or as part of a complex, is binding UASCHA.
Premature Death in Podospora anserina: Sporadic Accumulation of the Deleted Mitochondrial Genome, Translational Parameters and Innocuity of the Mating Types
Authors: Contamine Véronique , Lecellier Gael , Belcour Leon , Picard Marguerite
Published: January 2021

The Podospora anserina premature death syndrome was described as early growth arrest caused by a site-specific deletion of the mitochondrial genome (mtDNA) and occurring in strains displaying the genotype AS1-4 mat−. The AS1-4 mutation lies in a gene encoding a cytosolic ribosomal protein, while mat− is one of the two forms (mat− and mat+) of the mating-type locus. Here we show that, depending on culture conditions, death due to the accumulation of the deleted mtDNA molecule can occur in the AS1-4 mat+ context and can be delayed in the AS1-4 mat− background. Furthermore, we show that premature death and the classical senescence process are mutually exclusive. Several approaches permit the identification of the mat-linked gene involved in the appearance of premature death. This gene, rmp, exhibits two natural alleles, rmp− linked to mat− and rmp+ linked to mat+. The first is probably functional while the second probably carries a nonsense mutation and is sporadically expressed through natural suppression. A model is proposed that emphasizes the roles played by the AS1-4 mutation, the rmp gene, and environmental conditions in the accumulation of the deleted mitochondrial genome characteristic of this syndrome.
Macrorestriction Analysis of Caenorhabditis elegans Genomic DNA
Authors: Browning Heidi , Berkowitz Laura , Madej Cynthia , Paulsen Janet E , Zolan Miriam E , Strome Susan
Published: January 2021

The usefulness of genomic physical maps is greatly enhanced by linkage of the physical map with the genetic map. We describe a “macrorestriction mapping” procedure for Caenarhabditis elegans that we have applied to this endeavor. High molecular weight, genomic DNA is digested with infrequently cutting restriction enzymes and size-fractionated by pulsed field gel electrophoresis. Southern blots of the gels are probed with clones from the C. elegans physical map. This procedure allows the construction of restriction maps covering several hundred kilobases and the detection of polymorphic restriction fragments using probes that map several hundred kilobases away. We describe several applications of this technique. (1) We determined that the amount of DNA in a previously uncloned region is <220 kb. (2) We mapped the mes-1 gene to a cosmid, by detecting polymorphic restriction fragments associated with a deletion allele of the gene. The 25-kb deletion was initially detected using as a probe sequences located ~400 kb away from the gene. (3) We mapped the molecular endpoint of the deficiency hDf6, and determined that three spontaneously derived duplications in the unc-38-dpy-5 region have very complex molecular structures, containing internal rearrangements and deletions.
Molecular Characterization of hobo-Mediated Inversions in Drosophila melanogaster
Authors: Eggleston William B , Rim Nac R , Lim Johng K
Published: January 2021

The structure of chromosomal inversions mediated by hobo transposable elements in the Uc-1 X chromosome was investigated using cytogenetic and molecular methods. Uc-1 contains a phenotypically silent hobo element inserted in an intron of the Notch locus. Cytological screening identified six independent Notch mutations resulting from chromosomal inversions with one breakpoint at cytological position 3C7, the location of Notch. In situ hybridization to salivary gland polytene chromosomes determined that both ends of each inversion contained hobo and Notch sequences. Southern blot analyses showed that both breakpoints in each inversion had hobo-Notch junction fragments indistinguishable in structure from those present in the Uc-1 X chromosome prior to the rearrangements. Polymerase chain reaction amplification of the 12 hobo-Notch junction fragments in the six inversions, followed by DNA sequence analysis, determined that each was identical to one of the two hobo-Notch junctions present in Uc-1. These results are consistent with a model in which hobo-mediated inversions result from homologous pairing and recombination between a pair of hobo elements in reverse orientation.
Dosage Compensation Regulatory Proteins and the Evolution of Sex Chromosomes in Drosophila
Authors: Bone James R , Kuroda Mitzi I
Published: January 2021

In the fruitfly Drosophila melanogaster, the four male-specific lethal (msl) genes are required to achieve dosage compensation of the male X chromosome. The MSL proteins are thought to interact with cis-acting sites that confer dosage compensation to nearby genes, as they are detected at hundreds of discrete sites along the length of the polytene X chromosome in males but not in females. The histone H4 acetylated isoform, H4Ac16, colocalizes with the MSL proteins at a majority of sites on the D. melanogaster X chromosome. Using polytene chromosome immunostaining of other species from the genus Drosophila, we found that X chromosome association of MSL proteins and H4Ac16 is conserved despite differences in the sex chromosome karyotype between species. Our results support a model in which cis-acting regulatory sites for dosage compensation evolve on a neo-X chromosome arm in response to the degeneration of its former homologue.
Quantitative Genetic Variation of Odor-Guided Behavior in a Natural Population of Drosophila melanogaster
Authors: Mackay Trudy F C , Hackett J Brant , Lyman Richard F , Wayne Marta L , Anholt Robert R H
Published: January 2021

Quantitative genetic variation in behavioral response to the odorant, benzaldehyde, was assessed among a sample of 43 X and 35 third chromosomes extracted from a natural population and substituted into a common inbred background. Significant genetic variation among chromosome lines was detected. Heritability estimates for olfactory response, however, were low, as is typical for traits under natural selection. Furthermore, the loci affecting naturally occurring variation in olfactory response to benzaldehyde were not the same in males and females, since the genetic correlation between the sexes was low and not significantly different from zero for the chromosome 3 lines. Competitive fitness, viability and fertility of the chromosome 3 lines were estimated using the balancer equilibrium technique. Genetic correlations between fitness and odor-guided behavior were not significantly different from zero, suggesting the number of loci causing variation in olfactory response is small relative to the number of loci causing variation in fitness. Since different genes affect variation in olfactory response in males and females, genetic variation for olfactory response could be maintained by genotype × sex environment interaction. This unusual genetic architecture implies that divergent evolutionary trajectories for olfactory behavior may occur in males and females.
Sequence Analysis of Ewkaryotic Developmental Proteins: Ancient and Novel Domains
Authors: Mushegian Arcady R , Koonin Eugene V
Published: January 2021

Most of the genes involved in the development of multicellular eukaryotes encode large, multidomain proteins. To decipher the major trends in the evolution of these proteins and make functional predictions for uncharacterized domains, we applied a strategy of sequence database search that includes construction of specialized data sets and iterative subsequence masking. This computational approach allowed us to detect previously unnoticed but potentially important sequence similarities. Developmental gene products are enriched in predicted nonglobular regions as compared to unbiased sets of eukaryotic and bacterial proteins. Developmental genes that act intracellularly, primarily at the level of transcription regulation, typically code for proteins containing highly conserved DNA-binding domains, most of which appear to have evolved before the radiation of bacteria and eukaryotes. We identified bacterial homologues, namely a protein family that includes the Escherichia coli universal stress protein UspA, for the MADS-box transcription regulators previously described only in eukaryotes. We also show that the FUS6 family of eukaryotic proteins contains a putative DNA-binding domain related to bacterial helix-turn-helix transcription regulators. Developmental proteins that act extracellularly are less conserved and often do not have bacterial homologues. Nevertheless, several provocative similarities between different groups of such proteins were detected.
Genetic and Physical Variability at the Mating Type Locus of the Oomycete, Phytophthora infestans
Authors: Judelson Howard S
Published: January 2021

Mating type in the oomyceteous fungus, Phytophthora infestans, is determined by a single locus. In a previous study of a few isolates, the locus segregated in a manner genetically consistent with its linkage to a system of balanced lethal loci. To determine the prevalence of this phenomenon within P. infestans, genetic analyses were performed using isolates representative of the diversity within the species that had been selected by DNA fingerprinting using probes linked to mating type. Non-Mendelian segregation of the mating type locus was observed in crosses performed with each isolate. An unusual group of isolates was identified in which the mating type determinants had been rearranged within the genome; these strains also produced an aberrantly large number of self-fertile progeny. Curiously, in all isolates, markers linked to the mating type locus appeared prone to duplication, transposition, deletion, or other rearrangement. This was not observed for loci unlinked to mating type. Data from the crosses and analyses of marker variation were used to erect models to explain the bases of mating type determination and of the unusual segregation of the chromosomal region containing the mating type locus.
Cloning and Characterization of the scute (sc) Gene of Drosophila subobscura
Authors: Botella L M , Doñoro C , Sáchez L , Segarra C , Granadino B
Published: January 2021

The scute (sc) gene, a member of the achaete-scute complex of Drosophila melanogaster, has dual functions: sisterless (sis-b) function required for sex determination and dosage compensation and scute function, which is involved in neurogenesis. The sc homologue of D. subobscura was cloned. It lacks introns and encodes a single 1.7-kb transcript slightly larger than that of D. melanogaster (1.6 kb). The Sc protein of D. subobscura is slightly larger than that of D. melanogaster (382 vs. 345 amino acids). Sequence comparisons between both species show the Sc protein to have a highly conserved bHLH domain. Outside this domain, amino acid replacements are not randomly distributed. Two additional conserved domains, of 20 and 36 amino acids, are present near the C-terminal end. They may represent domains confering specificity upon the Sc protein with respect to other proteins of the achaete-scute complex. In its 3′ untranslated region, Sc RNA contains uridine stretches, putative Sxl protein DNA-binding sites. The D. subobscura Sc protein can cooperate with other D. melanogaster bHLH proteins because D. subobscura sc supplies sis-b function when introduced into D. melanogaster transgenic flies mutant for sc.
Mutations in New Cell Cycle Genes That Fail to Complement a Multiply Mutant Third Chromosome of Drosophila
Authors: White-Cooper Helen , Cannena Mar , Gonzalez Cayetano , Glover David M
Published: January 2021

We have simultaneously screened for new alleles and second site mutations that fail to complement five cell cycle mutations of Drosophila carried on a single third chromosome (gnu, polo, mgr, asp, stg). Females that are either transheterozygous for scott of the antartic (scant) and polo, or homozygous for scant produce embryos that show mitotic defects. A maternal effect upon embryonic mitoses is also seen in embryos derived from females transheterozygous with helter skelter (hsk) and either mgr or asp. deopatra (cleo), fails to complement asp but is not uncovered by a deficiency for asp. The mitotic phenotype of larvae heterozygous for cleo and the multiple mutant chromosome is similar to weak alleles of asp, but there are no defects in male meiosis. Mutations that failed to complement stgfell into two complementation groups corresponding to stg and a new gene noose. Three of the new stg alleles are early zygotic lethals, whereas the fourth is a pharate adult lethal allele that affects both mitosis and meiosis. Mutations in noose fully complement a small deficiency that removes stg, but when placed in trans to certain stg alleles, result in late lethality and mitotic abnormalities in larval brains.
The Speciation history of Drosophila pseudoobscura and Close Relatives: Inferences from DNA Sequence Variation at the Period Locus
Authors: Wang Rong-Lin , Hey Jody
Published: January 2021

Thirty-five period locus sequences from Drosophila pseudoobscura and its siblings species, D. p. bogotana, D. persirnilis, and D. miranda, were studied. A large amount of variation was found within D. pseudoobscura and D. persimilis, consistent with histories of large effective population sizes. D. p. bogotana, however, has a severe reduction in diversity. Combined analysis of per with two other loci, in both D. p. bogotana and D. pseudoobscura, strongly suggest this reduction is due to recent directional selection at or near per within D. p. bogotana. Since D. p. bogotana is highly variable and shares variation with D. pseudoobscura at other loci, the low level of variation at per within D. p. bogotana can not be explained by a small effective population size or by speciation via founder event. Both D. pseudoobscura and D. persirnilis have considerable intraspecific gene flow. A large portion of one D. persirnilis sequence appears to have arisen via introgression from D. pseudoobscura. The time of this event appears to be well after the initial separation of these two species. The estimated times since speciation are one mya for D. pseudoobscura and D. persirnilis and 2 mya since the formation of D. miranda.
Discord Between the Phylogenies Inferred From Molecular Versus Functional Data: Uneven Rates of Functional Evolution or Low Levels of Gene Flow?
Authors: Palopoli Michael F , Davis Andrew W , Wu Chung-I
Published: January 2021

According to measures of molecular divergence, the three species of the Drosophila simulans clade are closely related to and essentially equidistant from each other. We introgressed 10% of the D. sechellia X chromosome into a pure D. simulans genetic background and found that males carrying this introgressed region were consistently fertile; in contrast, males carrying the same segment from D. mauritiana are sterile and suffer from incompatibilities at a minimum of four loci. Together with other recent results, these data suggest that D. simulans and D. sechellia are much more closely related to each other than either is to D. mauritiana. How can we reconcile the phylogeny inferred from the density of hybrid sterility genes with that inferred from molecular divergence? If the molecular phylogeny is correct, the discrepancy might be explained by uneven rates of functional evolution, resulting in the uneven accumulation of substitutions with corresponding negative effects in hybrids. If the functional phylogeny is correct, then low levels of gene flow across nascent species boundaries, particularly for loci not tightly linked to a hybrid sterility gene, may have erased the original pattern of lineage splitting. We propose tests that will allow us to discriminate between these hypotheses.
Genesis and Variability of [PSI] Prion Factors in Saccharomyces cerevisiae
Authors: Derkatch Irina L , Chernoff Yury O , Kushnirov Vitaly V , Inge-Vechtomov Sergey G , Liebman Susan W
Published: January 2021

We have previously shown that multicopy plasmids containing the complete SUP35 gene are able to induce the appearance of the non-Mendelian factor [PSI]. This result was later interpreted by others as a crucial piece of evidence for a model postulating that [PSI] is a self-modified, prion-like conformational derivative of the Sup35 protein. Here we support this interpretation by proving that it is the overproduction of Sup35 protein, and not the excess of SUP35 DNA or mRNA that causes the appearance of [PSI]. We also show that the “prion-inducing domain” of Sup35p is in the N-terminal region, which, like the “prion-inducing domain” of another yeast prion, Ure2p, was previously shown to be distinct from the functional domain of the protein. This suggests that such a chimeric organization may be a common pattern of some prion elements. Finally, we find that [PSI] factors of different efficiencies and different mitotic stabilities are induced in the same yeast strain by overproduction of the identical Sup35 protein. We suggest that the different [PSI]-containing derivatives are analogous to the mysterious mammalian prion strains and result from different conformational variants of Sup35p.
A Dominant Negative Effect of eth-1r, a Mutant Allele of the Neurospora crassa S-Adenosylmethionine Synthetase-Encoding Gene Conferring Resistance to the Methionine Toxic Analogue Ethionine
Authors: Barra José L , Mautino Mario R , Rosa Alberto L
Published: January 2021

eth-1r a thermosensitive allele of the Neurospora crassa S-adenosylmethionine (AdoMet) synthetase gene that confers ethionine resistance, has been cloned and sequenced. Replacement of an aspartic amino acid residue (D48 → N48), perfectly conserved in prokaryotic, fungal and higher eukaryotic AdoMet synthetases, was found responsible for both thermosensitivity and ethionine resistance conferred by eth-1r. Gene fusion constructs, designed to overexpress eth-1r in vivo, render transformant cells resistant to ethionine. Dominance of ethionine resistance was further demonstrated in eth-1  +/eth-1r partial diploids carrying identical gene doses of both alleles. Heterozygous eth-1  +/eth-1r cells have, at the same time, both the thermotolerance conferred by eth-1  + and the ethionine-resistant phenotype conferred by eth-1r. AdoMet levels and AdoMet synthetase activities were dramatically decreased in heterozygous eth-1  +/eth-1r cells. We propose that this negative effect exerted by eth-1r results from the in vivo formation of heteromeric eth-1  +/eth-1r AdoMet synthetase molecules.
Mapping Three Classical Isozyme Loci in Tetrahymena: Meiotic Linkage of EstA to the ChxA Linkage Group
Authors: Allen Sally Lyman , Zeilinger Dawn , Orias Eduardo
Published: January 2021

We demonstrate a reliable method for mapping conventional loci and obtaining meiotic linkage data for the ciliated protozoan Tetrahymena thermophila. By coupling nullisomic deletion mapping with meiotic linkage mapping, loci known to be located on a particular chromosome or chromosome arm can be tested for recombination. This approach has been used to map three isozyme loci, EstA (Esterase A), EstB (Esterase B), and AcpA (Acid Phosphatase A), with respect to the ChxA locus (cycloheximide resistance) and 11 RAPDs (randomly amplified polymorphic DNAs). To assign isozyme loci to chromosomes, clones of inbred strains C3 or C2 were crossed to inbred strain B nullisomics. EstA, EstB and AcpA were mapped to chromosomes 1R, 3L and 3R, respectively. To test EstA and AcpA for linkage to known RAPD loci on their respective chromosomes, a panel of Round II (genomic exclusion) segregants from a B/C3 heterozygote was used. Using the MAPMAKER program, EstA was assigned to the ChxA linkage group on chromosome IR, and a detailed map was constructed that includes 10 RAPDs. AcpA (on 3R), while unlinked to all the RAPDs assigned to chromosome 3 by nullisomic mapping, does show linkage to a RAPD not yet assignable to chromosomes by nullisomic mapping.
Genetic Interactions Between Naturally Occuning Alleles at Quantitative Trait Loci and Mutant Alleles at Candidate Loci Affecting Bristle Number in Drosophila melanogaster
Authors: Long Anthony D , Mullaney Susan L , Mackay Trudy F C , Langley Charles H
Published: January 2021

Previously, we mapped quantitative trait loci (QTL) affecting response to short-term selection for abdominal bristle number to seven suggestive regions that contain loci involved in bristle development and/or that have adult bristle number mutant phenotypes, and are thus candidates for bristle number QTL in natural populations. To test the hypothesis that the factors contributing to selection response genetically interact with these candidate loci, high and low chromosomes from selection lines were crossed to chromosomes containing wild-type or mutant alleles at the candidate loci, and the numbers of bristles were recorded in trans heterozygotes. Quantitative failure to complement, detected as a significant selection line*cross effect by analysis of variance, can be interpreted as evidence for allelism or epistasis between the factors on selected chromosomes and the candidate loci. Mutations at some candidate loci (bb, emc, h, Dl, Hairless) showed strong interactions with selected chromosomes, whereas others interacted weakly (ASC, abd, Scr) or not at all (N, mab, E(spl)). These results support the hypothesis that some candidate loci, initially identified through mutations of large effect on bristle number, either harbor or are close members in the same genetic pathway as variants that contribute to standing variation in bristle number.
Reduced Variation in Drosophila simulans Mitochondrial DNA
Authors: Ballad J William O , Hatzidakis Joy , Karr Timothy L , Kreitman Martin
Published: January 2021

We investigated the evolutionary dynamics of infection of a Drosophila simulans population by a maternally inherited insect bacterial parasite, Wolbachia, by analyzing nucleotide variability in three regions of the mitochondrial genome in four infected and 35 uninfected lines. Mitochondrial variability is significantly reduced compared to a noncoding region of a nuclear-encoded gene in both uninfected and pooled samples of flies, indicating a sweep of genetic variation. The selective sweep of mitochondrial DNA may have been generated by the fixation of an advantageous mitochondrial gene mutation in the mitochondrial genome. Alternatively, the dramatic reduction in mitochondrial diversity may be related to Wolbachia.
Functional and Genetic Characterization of the Oligomerization and DNA Binding Properties of the Drosophila Doublesex Proteins
Authors: Erdman Scott E , Chen Hui-Ju , Burtis Kenneth C
Published: January 2021

The doublesex (dsx) gene of Drosophila melanogaster encodes both male-specific (DSXM) and female-specific (DSXF) polypeptides, which are required for normal differentiation of numerous sexually dimorphic somatic traits. The DSX polypeptides are transcription factors and have been shown previously to bind through a zinc finger-like domain to specific sites in an enhancer regulating sex-specific expression of yolk protein genes. We have determined the consensus target sequence for this DNA binding domain to be a palindromic sequence NNACTAAGAATGTNNTC composed of two half-sites around a central (A/T) base pair. As predicted by the symmetric nature of this site, we have found that the DSX proteins exist as dimers in vivo and have mapped two independent dimerization domains by the yeast two-hybrid method; one in the non-sex-specific amino-terminal region of the protein and one that includes the partially sex-specific carboxy-terminal domains of both the male and female polypeptides. We have further identified a missense mutation that eliminates dsx function in female flies, and shown that the same mutation prevents dimerization of DSXF in the yeast two-hybrid system, indicating a critical role for dimerization in dsx function in vivo.
A Transgene-Induced Mitotic Arrest Mutation in the Mouse Allelic With Oligosyndactylism
Authors: Pravtcheva Dimitrina D , Wise Thomas L
Published: January 2021

Oligosyndactylism (Os) is a radiation-induced mutation on mouse chromosome 8 associated with early postimplantation lethality in homozygotes and abnormal development of the limbs and kidneys in heterozygotes. The recessive lethal effect of Os is due to a mitotic block of the embryonic cells that becomes apparent at the blastocyst stage, but it is not known if the heterozygous effect of Os is due to haploinsufficiency of the gene responsible for the mitotic arrest, or is due to mutation(s) of other gene(s). We have recently described a transgene-induced recessive mutation, 94-A/K, that results in early postimplantation death of the embryos, and we have mapped this mutation to the same region of chromosome 8 where Os has been assigned. On the basis of complementation tests between transgenic and Os/+ mice, in vitro growth characteristics and increased mitotic index of 94-A/K embryos, and molecular structural analysis of 94-A and 94-K transgenic and Os/+ mice, we conclude that the 94-A/K mutation represents a new allele of Os. This insertional mutation should facilitate the isolation of a mammalian gene essential for normal progression of the cell cycle beyond metaphase.
Low Frequency of Mouse t Haplotypes in Wild Populations Is Not Explained by Modifiers of Meiotic Drive
Authors: Ardlie Kristin G , Silver Lee M
Published: January 2021

t haplotypes are naturally occurring forms of mouse chromosome 17 that show non-Mendelian transmission from heterozygous +/t males. In laboratory studies, transmission ratios of ≥0.90 or higher are typically observed. With transmission ratios of this level, theoretical analyses predict high frequencies of t haplotypes (∼75%) in wild populations. In contrast, empirical frequencies of only 15–25% are typically found. This has led to the suggestion that modifiers of drive may play a role in reducing t frequencies. We have measured transmission ratio distortion (TRD) levels in wild +/t mice to examine this hypothesis. TRD was very high in both litters collected from wild-caught pregnant females, and in wild litters bred in the laboratory (mean = 0.9). Contrary to the results of other studies, we found no difference in TRD levels between semilethal and lethal t haplotypes nor between litters conceived from cycling or postpartum estrus. We found three litters with aberrantly low TRDs that were all multiply sired, although the role this might play in natural populations is unknown. These findings show a general absence of modifiers of drive in natural populations and suggest that other factors are responsible for the low observed frequencies of wild t haplotypes.
Marker-Assisted Introgression in Backcross Breeding Programs
Authors: Visscher Peter M , Haley Chris S , Thompson Robin
Published: January 2021

The efficiency of marker-assisted introgression in backcross populations derived from inbred lines was investigated by simulation. Background genotypes were simulated assuming that a genetic model of many genes of small effects in coupling phase explains the observed breed difference and variance in backcross populations. Markers were efficient in introgression backcross programs for simultaneously introgressing an allele and selecting for the desired genomic background. Using a marker spacing of 10–20 cM gave an advantage of one to two backcross generations selection relative to random or phenotypic selection. When the position of the gene to be introgressed is uncertain, for example because its position was estimated from a trait gene mapping experiment, a chromosome segment should be introgressed that is likely to include the allele of interest. Even for relatively precisely mapped quantitative trait loci, flanking markers or marker haplotypes should cover ∼10–20 cM around the estimated position of the gene, to ensure that the allele frequency does not decline in later backcross generations.
An Allelic Series of Blue Fluorescent trp1 Mutants of Arabidopsis thaliana
Authors: Rose Alan B , Li Jiayang , Last Robert L
Published: January 2021

Nine blue fluorescent mutants of the flowering plant Arabidopsis thaliana were isolated by genetic selections and fluorescence screens. Each was shown to contain a recessive allele of trp1, a previously described locus that encodes the tryptophan biosynthetic enzyme phosphoribosylanthranilate transferase (PAT, called trpD in bacteria). The trp1 mutants consist of two groups, tryptophan auxotrophs and prototrophs, that differ significantly in growth rate, morphology, and fertility. The trp1 alleles cause plants to accumulate varying amounts of blue fluorescent anthranilate compounds, and only the two least severely affected of the prototrophs have any detectable PAT enzyme activity. All four of the trp1 mutations that were sequenced are G to A or C to T transitions that cause an amino acid change, but in only three of these is the affected residue phylogenetically conserved. There is an unusually high degree of sequence divergence in the single-copy gene encoding PAT from the wild-type Columbia and Landsberg erecta ecotypes of Arabidopsis.
On the Specificity of Adaptive Mutations
Authors: Hall Barry G
Published: January 2021

Adaptive mutations are mutations that occur in nondividing or slowly dividing cells during prolonged nonlethal selection, and that appear to be specific to the challenge of the selection in the sense that the only mutations that arise are those that provide a growth advantage to the cell. The issue of the specificity has been controversial because it violates our most basic assumptions about the randomness of mutations with respect to their effect on the cell. Although a variety of experiments in several systems in both bacteria and yeast have claimed to demonstrate that specificity, those experiments have been subjected to a variety of technical criticisms suggesting that the specificity may not be real. Here I use the ebg system to provide evidence that when selection is applied to one specific nucleotide site within a gene, mutation occurs at that site but not at an alternative and equally mutable site within the same gene.
Molecular Evolution at the decapentaplegic Locus in Drosophila
Authors: Newfeld Stuart J , Padgett Richard W , Findley Seth D , Richter Brent G , Sanicola Michele , de Cuevas Margaret , Gelbart William M
Published: January 2021

Using an elaborate set of cis-regulatory sequences, the decapentaplegic (dpp) gene displays a dynamic pattern of gene expression during development. The C-terminal portion of the DPP protein is processed to generate a secreted signaling molecule belonging to the transforming growth factor-β (TGF-β) family. This signal, the DPP ligand, is able to influence the developmental fates of responsive cells in a concentration-dependent fashion. Here we examine the sequence level organization of a significant portion of the dpp locus in Drosophila melanogaster and use interspecific comparisons with D. simulans, D. pseudoobscura and D.virilis to explore the molecular evolution of the gene. Our interspecific analysis identified significant selective constraint on both the nucleotide and amino acid sequences. As expected, interspecific comparison of protein coding sequences shows that the C-terminal ligand region is highly conserved. However, the central portion of the protein is also conserved, while the N-terminal third is quite variable. Comparison of noncoding regions reveals significant stretches of nucleotide identity in the 3′ untranslated portion of exon 3 and in the intron between exons 2 and 3. An examination of cDNA sequences representing five classes of dpp transcripts indicates that these transcripts encode the same polypeptide.
Beaded of Goldschmidt, an Antimorphic Allele of Serrate, Encodes a Protein Lacking Transmembrane and Intracellular Domains
Authors: Hukriede Neil A , Fleming Robert J
Published: January 2021

Serrate (Ser) is an essential gene in Drosophila melanogaster best known for the Ser dominant (SerD) allele and its effects on wing development. Animals heterozygous or homozygous for (SerD) are viable and exhibit loss of wing margin tissue and associated bristles and hairs. The Beaded of Goldschmidt (BdG) allele of Ser, when heterozygous to wild type, will also produce animals exhibiting loss of wing margin material. However, animals homozygous for BdG exhibit a larval lethal phenotype comparable to animals homozygous for loss-of-function Ser alleles. BdG is a partial duplication of the Ser locus with a single 5′ Ser-homologous region and two distinct 3′ regions. Meiotic recombination between BdG and a wild-type Ser chromosome demonstrated that only one DNA lesion, caused by the insertion of a transposable roo element into the coding regions of the Ser transcript, appears capable of generating BdG phenotypes. Due to the roo insertion, the protein product is predicted to be prematurely truncated and lack an extracellular cysteine-rich region along with the transmembrane and intracellular domains found within the normal SERRATE (SER) protein. The loss of these protein domains apparently contributes to the antimorphic nature of this mutation.
Origin and Evolution of a New Gene Descended From alcohol dehydrogenase in Drosophila
Authors: Begun David J
Published: January 2021

Drosophila alcohol dehydrogenase (Adh) is highly conserved in size, organization, and amino acid sequence. Adh-ψ was hypothesized to be a pseudogene derived from an Adh duplication in the repleta group of Drosophila; however, several results from molecular analyses of this gene conflict with currently held notions of molecular evolution. Perhaps the most difficult observations to reconcile with the pseudogene hypothesis are that the hypothetical replacement sites of Adh-ψ evolve only slightly more quickly than replacement sites of closely related, functional Adh genes, and that the replacement sites of the pseudogenes evolve considerably more slowly than neighboring silent sites. The data have been presented as a paradox that challenges our understanding of the mechanisms underlying DNA sequence divergence. Here I show that Adh-ψ is actually a new, functional gene recently descended from an Adh duplication. This descendant recruited ∼60 new N-terminal amino acids, is considerably more basic than ADH, and is evolving at a faster rate than Adh. Furthermore, though the descendant is clearly functional, as inferred from molecular evolution and population genetic data, it retains no obvious ADH activity. This probably reflects functional divergence from its Adh ancestor.
Multinomial-Sampling Models for Random Genetic Drift
Authors: Nagylaki Thomas
Published: January 2021

Three different derivations of models with multinomial sampling of genotypes in a finite population are presented. The three derivations correspond to the operation of random drift through population regulation, conditioning on the total number of progeny, and culling, respectively. Generations are discrete and nonoverlapping; the diploid population mates at random. Each derivation applies to a single multiallelic locus in a monoecious or dioecious population; in the latter case, the locus may be autosomal or X-linked. Mutation and viability selection are arbitrary; there are no fertility differences. In a monoecious population, the model yields the Wright-Fisher model (i.e., multinomial sampling of genes) if and only if the viabilities are multiplicative. In a dioecious population, the analogous reduction does not occur even for pure random drift. Thus, multinomial sampling of genotypes generally does not lead to multinomial sampling of genes. Although the Wright-Fisher model probably lacks a sound biological basis and may be inaccurate for small populations, it is usually (perhaps always) a good approximation for genotypic multinomial sampling in large populations.
The Spread of an Advantageous Allele Across a Barrier: The Effects of Random Drift and Selection Against Heterozygotes
Authors: Piálek Jaroslav , Barton Nick H
Published: January 2021

A local barrier to gene flow will delay the spread of an advantageous allele. Exact calculations for the deterministic case show that an allele that is favorable when rare is delayed very little even by a strong barrier: its spread is slowed by a time proportional to log((B/σ)√2S)/S, where B is the barrier strength, σ the dispersal range, and fitnesses are 1:1 + S:1 + 2S. However,when there is selection against heterozygotes, such that the allele cannot increase from low frequency, a barrier can cause a much greater delay. If gene flow is reduced below a critical value, spread is entirely prevented. Stochastic simulations show that with additive selection, random drift slows down the spread of the allele, below the deterministic speed of σ√2S. The delay to the advance of an advantageous allele caused by a strong barrier can be substantially increased by random drift and increases with B/(2Sρσ2) in a one-dimensional habitat of density ρ. However, with selection against heterozygotes, drift can facilitate the spread and can free an allele that would otherwise be trapped indefinitely by a strong barrier. We discuss the implications of these results for the evolution of chromosome rearrangements.
Epigenetic Inheritance of Transcriptional Silencing and Switching Competence in Fission Yeast
Authors: Thon Geneviève , Friis Tove
Published: January 2021

Epigenetic events allow the inheritance of phenotypic changes that are not caused by an alteration in DNA sequence. Here we characterize an epigenetic phenomenon occuring in the mating-type region of fission yeast. Cells of fission yeast switch between the P and M mating-type by interconverting their expressed mating-type cassette between two allelic forms, mat1-P and mat1-M. The switch results from gene conversions of mat1 by two silent cassettes, mat2-P and mat3-M, which are linked to each other and to mat1. Grewal and Klar observed that the ability to both switch mat1 and repress transcription near mat2-P and mat3-M was maintained epigenetically in a strain with an 8-kb deletion between mat2 and mat3. Using a strain very similar to theirs, we determined that interconversions between the switching- and silencing-proficient state and the switching and silencing-deficient state occurred less frequently than once per 1000 cell divisions. Although transcriptional silencing was alleviated by the 8-kb deletion, it was not abolished. We performed a mutant search and obtained a class of trans-acting mutations that displayed a strong cumulative effect with the 8-kb deletion. These mutations allow to assess the extent to which silencing is affected by the deletion and provide new insights on the redundancy of the silencing mechanism.
Cytosolic Ribosomal Mutations That Abolish Accumulation of Circular Intron in the Mitochondria Without Preventing Senescence of Podospora anserina
Authors: Silar Philippe , Koll France , Rossignol Michèle
Published: January 2021

The filamentous fungus Podospora anserina presents a degeneration syndrome called Senescence associated with mitochondrial DNA modifications. We show that mutations affecting the two different and interacting cytosolic ribosomal proteins (S7 and S19) systematically and specifically prevent the accumulation of senDNAα (a circular double-stranded DNA plasmid derived from the first intron of the mitochondrial cox1 gene or intron α) without abolishing Senescence nor affecting the accumulation of other usually observed mitochondrial DNA rearrangements. One of the mutant proteins is homologous to the Escherichia coli S4 and Saccharomyces cerevisiae S13 ribosomal proteins, known to be involved in accuracy control of cytosolic translation. The lack of accumulation of senDNAα seems to result from a nontrivial ribosomal alteration unrelated to accuracy control, indicating that S7 and S19 proteins have an additional function. The results strongly suggest that modified expression of nucleus-encoded proteins contributes to Senescence in P. anserina. These data do not fit well with some current models, which propose that intron α plays the role of the cytoplasmic and infectious Determinant of Senescence that was defined in early studies.
On the Mode of Gene-Dosage Compensation in Drosophila
Authors: Arkhipova Irina , Li Jingjing , Meselson Matthew
Published: January 2021

A procedure is described for determining the mode and magnitude of gene-dosage compensation of transformed genes. It involves measurement of the ratio of the activity of a gene inserted at X-linked sites to the activity of the same gene inserted at autosomal sites. Applying the procedure to the Drosophila pseudoobscura Hsp82 gene inserted at ectopic sites in D. melanogaster and taking gene activity as proportional to the amount of transcript per gene copy, we conclude that (1) in both adults and larvae the gene is not compensated at autosomal sites or at a site in β-heterochromatin at the base of the X chromosome and is fully compensated at euchromatic X-chromosomal sites; (2) inappropriate normalization is responsible for a claim that the gene is compensated at autosomal sites; and (3) the observed compensation operates mainly or entirely by heightened activity of X-linked genes in males, rather than by reduced activity in females.
Identification of Trans-Acting Genes Necessary for Centromere Function in Drosophila melanogaster Using Centromere-Defective Minichromosomes
Authors: Cook Kevin R , Murphy Terence D , Nguyen Thai C , Karpen Gary H
Published: January 2021

Deletions in the Drosophila minichromosome Dp1187 were used to investigate the genetic interactions of trans-acting genes with the centromere. Mutations in several genes known to have a role in chromosome inheritance were shown to have dominant effects on the stability of minichromosomes with partially defective centromeres. Heterozygous mutations in the ncd and klp3A kinesin-like protein genes strongly reduced the transmission of minichromosomes missing portions of the genetically defined centromere, but had little effect on the transmission of minichromosomes with intact centromeres. Using this approach, ncd and klp3A were shown to require only the centromeric region of the chromosome for their roles in chromosome segregation. Increased gene dosage also affected minichromosome transmission and was used to demonstrate that the nod kinesin-like protein gene interacts genetically with the centromere, in addition to interacting with extracentromeric regions as demonstrated previously. The results presented in this study strongly suggest that dominant genetic interactions between mutations and centromere-defective minichromosomes could be used effectively to identify novel genes necessary for centromere function.
FAR1, a Negative Regulatory Locus Required for the Repression of the Nitrate Reductase Gene in Chlamydomonas reinhardtii
Authors: Zhang Donghong , Lefebvre Paul A
Published: January 2021

In Chlamydomonas reinhardtii, the genes required for nitrate assimilation, including the gene encoding nitrate reductase (NIT1), are subject to repression by ammonia. To study the mechanism of ammonia repression, we employed two approaches to search for mutants with defective repression of NIT1 gene expression. (1) PF14, a gene required for flagellar function, was used as a reporter gene for expression from the NIT1 promoter. When introduced into a pf14 mutant host, the NZTl:PF14 chimeric construct produced a transformant (T10-10B) with a conditional swimming phenotype. Spontaneous mutants with defective ammonia repression of the NIT1 promoter were screened for by isolating cells that gained constitutive motility. (2) Insertional mutagenesis was performed, followed by screening for chlorate sensitivity in the presence of ammonia ion. One insertional mutant and six spontaneous mutants were allelic and defined a new gene, FAR1 (free from ammonia repression). FAR1 was mapped to Linkage Group I, 7.7 cM to the right of the centromere. The far1-1 mutant strain was used to clone DNA adjacent to the site of plasmid insertion, which was then used as a hybridization probe to clone the FAR1 gene from wild type.
A Deficiency Screen for Zygotic Loci Required for Establishment and Patterning of the Epidermis in Caenorhabditis elegans
Authors: Terns Rebecca M , Kroll-Conner Peggy , Zhu Jiangwen , Chung Sooyoun , Rothman Joel H
Published: January 2021

To identify genomic regions required for establishment and patterning of the epidermis, we screened 58 deficiencies that collectively delete at least ∼67% of the Caenorhabditis elegans genome. The epidermal pattern of deficiency homozygous embryos was analyzed by examining expression of a marker specific for one of the three major epidermal cell types, the seam cells. The organization of the epidermis and internal organs was also analyzed using a monoclonal antibody specific for epithelial adherens junctions. While seven deficiencies had no apparent effect on seam cell production, 21 were found to result in subnormal, and five in excess numbers of these cells. An additional 23 deficiencies blocked expression of the seam cell marker, in some cases without preventing cell proliferation. Two deficiencies result in multinucleate seam cells. Deficiencies were also identified that result in subnormal numbers of epidermal cells, hyperfusion of epidermal cells into a large syncytium, or aberrant epidermal differentiation. Finally, analysis of internal epithelia revealed deficiencies that cause defects in formation of internal organs, including circularization of the intestine and bifurcation of the pharynx lumen. This study reveals that many regions of the C. elegans genome are required zygotically for patterning of the epidermis and other epithelia.
The Genetics and Development of an Eyespot Pattern in the Butterfly Bicyclus anynana: Response to Selection for Eyespot Shape
Authors: Monteiro Antónia , Brakefield Paul M , French Vernon
Published: January 2021

The normally circular eyespots on the wing of the butterfly Bicyclus anynana were selected to become elliptical in two divergent lines, with antero-posterior elongation of the eyespots in one line and proximodistal elongation in the other. Selection was continued for nine generations, and symmetrical realized heritabilities of ∼15% were achieved initially. The elliptical eyespot shapes characteristic of each line were still produced when the signaling center of the eyespot (the focus) was surgically rotated by 90 or 180° or when an eyespot was induced ectopically by localized damage. We conclude that selection changed general properties of the epidermis that responds to signals emanating from the eyespot focus but did not affect the mechanism of focal signaling.