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D L Hartl

Publications and source records attributed to D L Hartl.

At least 55 records · Page 3Linked to original sources
PubMed1997

Trash DNA is what gets thrown away: high rate of DNA loss in Drosophila.

We have recently described a novel method of estimating neutral rates and patterns of spontaneous mutation (Petrov et al., 1996). This method takes advantage of the propensity of non-LTR retrotransposable elements to create non-functional, 'dead-on-arrival' copies as a product of transposition. Maximum parsimony analysis is used to separate the evolution of actively transposing lineages of a non-LTR element from the fate of individual inactive insertions, and thereby allows one to assess directly the relative rates of different types of mutation, including point substitutions, deletions and insertions. Because non-LTR elements enjoy wide phylogenetic distribution, this method can be used in taxa that do not harbor a significant number of bona fide pseudogenes, as is the case in Drosophila (Jeffs and Ashburner, 1991; Weiner et al., 1986). We used this method with Helena, a non-LTR retrotransposable element present in the Drosophila virilis species group. A striking finding was the virtual absence of insertions and remarkably high incidence of large deletions, which combine to produce a high overall rate of DNA loss. On average, the rate of DNA loss in D. virilis is approximately 75 times faster than that estimated for mammalian pseudogenes (Petrov et al., 1996). The high rate of DNA loss should lead to rapid elimination of non-essential DNA and thus may explain the seemingly paradoxical dearth of pseudogenes in Drosophila. Varying rates of DNA loss may also contribute to differences in genome size (Graur et al., 1989; Petrov et al., 1996), thus explaining the celebrated 'C-value' paradox (John and Miklos, 1988). In this paper we outline the theoretical basis of our method, examine the data from this perspective, and discuss potential problems that may bias our estimates.

Animals↗
PubMed1997

Mutations in the mariner transposase: the D,D(35)E consensus sequence is nonfunctional.

Genetic analysis of eukaryote transposases and comparison with their prokaryote counterparts have been greatly hindered by difficulty in isolating mutations. We describe a simple eye-color screen that facilitates isolation and analysis of mutations in the mariner transposase in Drosophila melanogaster. Use of ethyl methanesulfonate and site-directed mutagenesis has identified 18 residues that are critical for in vivo excision of a target mariner element. When the mutations were examined in heterozygous mutant/nonmutant genotypes, more than half of the mutant transposase proteins were found to reduce the activity of the wild-type transposase, as assayed by the frequency of germline excision of a target element. Remarkably, transposase function is obliterated when the D,D(34)D acidic, ion-binding domain is replaced with the consensus sequence D,D(34)E found in the nematode Tc1 transposase and in many other transposases in the superfamily. A number of mutations strongly complement wild-type transposase in a dominant-negative manner, suggestive of subunit interactions in the excision reaction; these mutations are located in a small region that includes part of the D,D(34)D motif. Transposase function also is eliminated by a mutation in the inferred initiation codon and by a mutation in a putative nuclear localization signal.

Animals↗
PubMed1997

A framework physical map of Drosophila virilis based on P1 clones: applications in genome evolution.

The analysis of patterns of genome evolution may help to evaluate the evolutionary forces that shape the composition and organization of the genome. Comparisons between the physical maps of divergent species can be used to identify conserved blocks of closely linked genes whose synteny is possibly under selective constraint. We have used in situ hybridization to determine the genomic position of 732 randomly selected clones from a bacteriophage P1 library of Drosophila virilis. The resulting map includes at least one clone in each of 69% of the subdivisions into which the D. virilis polytene chromosomes are divided. A subset of these clones was used to carry out a comparative physical analysis of chromosome 2 from D. virilis and from Drosophila montana. A number of discrepancies with the classical scenario of chromosome evolution were noted. The D. virilis P1 clones were also used to determine the physical relations between ten genes that are located in the X chromosome of Drosophila melanogaster between the markers crn (2F1) and omb (4C5-6). In this region, which is approximately 2 Mb in length, there have been at least six breakpoints since the divergence of the species, and six of the genes are found at widely scattered locations in the D. virilis X chromosome. However, a block of four functionally unrelated genes, including white, roughest, Notch, and dunce, seems to be conserved between the two species.

Animals↗
PubMed1997

What restricts the activity of mariner-like transposable elements.

A number of mechanisms have recently been described that might be important in restricting the level of activity of mariner-like transposable elements (MLEs) in natural populations. These mechanisms include overproduction inhibition, in which increasing the dose of transposase decreases net activity. Another mechanism is mediated by certain missense mutations, in which a mutant transposase protein impairs the activity of the wild-type transposase in heterozygous mutant/nonmutant genotypes. A further mechanism is the potential for transposase titration by defective elements that retain transposase binding activity. The issue of regulation is not only of theoretical importance in understanding the molecular and evolutionary genetics of MLEs, but also of practical significance in learning how best to use MLEs in the germline transformation of insect pests and disease vectors.

Animals↗
PubMed1997

Discordant rates of chromosome evolution in the Drosophila virilis species group.

In Drosophila, the availability of polytene chromosome maps and of sets of probes covering most regions of the chromosomes allows a direct comparison of the organization of the genome in different species. In this work, we report the localization, in Drosophila virilis, D. montana, and D. novamexicana, of > 100 bacteriophage PI clones containing approximately 65 kilobase inserts of genomic DNA from D. virilis. Each clone hybridizes with a single euchromatic site in either chromosome 1 or chromosome 3 in D. virilis. From these data, it is possible to estimate the minimum number of inversions required to transform the map positions of the probes in one species into the map positions of the same probes in a related species. The data indicate that, in the D. virilis species group, the X chromosome has up to four times the number of inversions as are observed in chromosome 3. The first photographic polytene chromosome maps for D. montana and D. novamexicana are also presented.

Animals↗
PubMed1997

The evolution of small gene clusters: evidence for an independent origin of the maltase gene cluster in Drosophila virilis and Drosophila melanogaster.

We analyzed a 5,770-bp genomic region of Drosophila virilis that contains a cluster of two maltase genes showing sequence similarity with genes in a cluster of three maltase genes previously identified in Drosophila melanogaster. The D. virilis maltase genes are designated Mav1 and Mav2. In addition to being different in gene number, the cluster of genes in D. virilis differs dramatically in intron-exon structure from the maltase genes in D. melanogaster, the transcriptional orientation of the genes in the cluster also differs between the species. Our findings support a model in which the maltase gene cluster in D. virilis and D. melanogaster evolved independently. Furthermore, while in D. melanogaster the maltase gene cluster lies only 10 kb distant from the larval cuticle gene cluster, the maltase and larval cuticle gene clusters in D. virilis are located very far apart and on a different chromosome than that expected from the known chromosome arm homologies between D. virilis and D. melanogaster. A region of the genome containing the maltase and larval cuticle gene clusters appears to have been relocated between nonhomologous chromosomes.

Aedes↗
PubMed1997

Nonrandom location of IS1 elements in the genomes of natural isolates of Escherichia coli.

We have studied the spatial distribution of IS1 elements in the genomes of natural isolates comprising the ECOR reference collection of Escherichia coli. We find evidence for nonrandomness at three levels. Many pairs of IS1 elements are in much closer proximity (< 10 kb) than can be accounted for by chance. IS1 elements in close proximity were identified by long-range PCR amplification of the genomic sequence between them. Each amplified region was sequenced and its map location determined by database screening of DNA hybridization. Among the ECOR strains with at least two IS1 elements, 54% had one or more pairs of elements separated by < 10 kb. We propose that this type of clustering is a result of "local hopping," in which we assume that a significant proportion of tranposition events leads to the insertion of a daughter IS element in the vicinity of the parental element. A second level of nonrandomness is found in strains with a modest number of IS1 elements that are mapped through the use of inverse PCR to amplify flanking genomic sequences: in these strains, the insertion sites tend to be clustered over a smaller region of chromosome than would be expected by chance. A third level of nonrandomness is observed in the composite distribution of IS elements across strains: among 20 mapped IS1 elements, none were found in the region of 48-77 minutes, a significant gap. One region of the E. coli chromosome, at 98 min, had a cluster of IS1 elements in seven ECOR strains of diverse phylogenetic origin. We deduce from sequence analysis that this pattern of distribution is a result of initial insertion in the most recent common ancestor of these strains and therefore not a hot spot of insertion. Analysis using long-range PCR with primers for IS2 and IS3 also yielded pairs of elements in close proximity, suggesting that these elements may also occasionally transpose by local hopping.

Chromosome Mapping↗
PubMed1997

Heterogeneity of the internal transcribed spacer (ITS-2) region within individual deer ticks.

To determine whether nuclear rDNA sequences provide a useful means for assessing the structure of populations of Ixodes ticks, we compared variability among copies of an internal transcribed spacer (ITS-2) sequence within individual ticks to the variability between ticks. At least 4% of the nucleotides comprising this sequence vary among the copies present within individual ticks. ITS-2 diversity in each of two ticks is nearly half as great as that reported between ticks from geographically disparate populations. Because individual ticks retain ancestral polymorphism, ITS-2 variation does not accurately reflect descent relationships among these ticks. Sequencing single copies of PCR-amplified ITS-2 therefore does not permit assessment of the phylogenetic relationships among the I. ricinus-like ticks in eastern North America. We recommend caution in future analyses, and emphasize the importance of procedures designed to ensure that the many paralogous copies of the rDNA cistron have been sufficiently homogenized by concerted evolutionary processes. Such precautionary measures will make certain that phylogenetic trees based on these gene sequences reflect the phyletic relatedness of the biological species.

Animals↗
PubMed1997

Recent horizontal transmission of plasmids between natural populations of Escherichia coli and Salmonella enterica.

Seventy-one natural isolates obtained from a Salmonella reference collection were examined for the presence of plasmids closely related to the Escherichia coli F plasmid. The collection consists of several serovars of the S. enterica Typhimurium complex, subspecies I, to which 99% of pathogenic salmonellae belong. Molecular genetic techniques of DNA hybridization, along with PCR and DNA sequencing, were used to examine the occurrence, distribution, and genetic diversity of F-like plasmids among Salmonella strains. The F plasmid genes examined were finO, traD, traY, and repA, which map at dispersed positions on the F plasmid of E. coli. Comparative sequence analysis of each of the four genes in Salmonella plasmids showed them to be homologous (in some cases, virtually identical) to those found in F plasmids of E. coli natural isolates. Furthermore, the frequency of F-like plasmids in Salmonella strains was approximately the same as that observed in the E. coli Reference Collection. However, in Salmonella, the distribution was confined predominately to the serovars Typhimurium and Muenchen. The unexpected finding of a shared pool of F-like plasmids between S. enterica and E. coli demonstrates the significant role of conjugation in the histories of these important bacterial species.

Amino Acid Sequence↗
PubMed1997

Modern thoughts on an ancyent marinere: function, evolution, regulation.

The mariner/Tc1 superfamily of transposable elements is one of the most diverse and widespread Class II transposable elements. Within the larger assemblage, the mariner-like elements (MLEs) and the Tc1-like elements (TLEs) are distinct families differing characteristically in the composition of the "D,D(35)E" cation-binding domain. Based on levels of sequence similarity, the elements in each family can be subdivided further into several smaller subfamilies. MLEs and TLEs both have an extraordinarily wide host range. They are abundant in insect genomes and other invertebrates and are found even in some vertebrate species including, in the case of mariner, humans, in which one element on chromosome 17p has been implicated as a hotspot of recombination. In spite of the extraordinary evolutionary success of the elements, virtually nothing is known about their mode of regulation within genomes. There is abundant evidence that the elements are disseminated to naive host genomes by horizontal transmission, and there is a substantial base of evidence for inference about the subsequent population dynamics. Studies of engineered mariner elements and induced mutations in the transposase have identified two mechanisms that may be operative in mariner regulation. One mechanism is overproduction inhibition, in which excessive wild-type transposase reduces the rate of excision of a target element. A second mechanism is dominant-negative complementation, in which certain mutant transposase proteins antagonize the activity of the wild-type transposase. The latter process may help explain why the vast majority of MLEs in nature undergo "vertical inactivation" by multiple mutations and, eventually, stochastic loss. There is also evidence that mariner/Tc1 elements can be mobilized in hybrid dysgenesis; in particular, certain dysgenic crosses in Drosophila virilis result in mobilization of a TLE designated Paris as well as the mobilization of other unrelated transposable elements.

Animals↗
PubMed1997

Regulation of the transposable element mariner.

The mariner/Tcl superfamily of transposable elements is widely distributed in animal genomes and is especially prevalent in insects. Their wide distribution results from their ability to be disseminated among hosts by horizontal transmission and also by their ability to persist in genomes through multiple speciation events. Although a great deal is known about the molecular mechanisms of transposition and excision, very little is known about the mechanisms by which transposition is controlled within genomes. The issue of mariner/Tcl regulation is critical in view of the great interest in these elements as vectors for germline transformation of insect pests and vectors of human disease. Several potentially important regulatory mechanisms have been identified in studies of genetically engineered mariner elements. One mechanism is overproduction inhibition, in which excessive wild-type transposase reduces the rate of excision of a target element. A second mechanism is mediated by certain mutant transposase proteins, which antagonize the activity of the wild-type transposase. The latter process may help explain why the vast majority of MLEs in nature undergo 'vertical inactivation' by multiple mutations and, eventually, stochastic loss. Another potential mechanism of regulation may result from transposase titration by defective elements that retain their DNA binding sites and ability to transpose. There is also evidence that some mariner/Tcl elements can be mobilized in a type of hybrid dysgenesis.

Animals↗
PubMed1996

High intrinsic rate of DNA loss in Drosophila.

Pseudogenes are common in mammals but virtually absent in Drosophila. All putative Drosophila pseudogenes show patterns of molecular evolution that are inconsistent with the lack of functional constraints. The absence of bona fide pseudogenes is not only puzzling, it also hampers attempts to estimate rates and patterns of neutral DNA change. The estimation problem is especially acute in the case of deletions and insertions, which are likely to have large effects when they occur in functional genes and are therefore subject to strong purifying selection. We propose a solution to this problem by taking advantage of the propensity of retrotransposable elements without long terminal repeats (non-LTR) to create non-functional, 'dead-on-arrival' copies of themselves as a common by-product of their transpositional cycle. Phylogenetic analysis of a non-LTR element, Helena, demonstrates that copies lose DNA at an unusually high rate, suggesting that lack of pseudogenes in Drosophila is the product of rampant deletion of DNA in unconstrained regions. This finding has important implications for the study of genome evolution in general and the 'C-value paradox' in particular.

Animals↗
PubMed1996

Compensatory nearly neutral mutations: selection without adaptation.

One implication of Kacser's analysis of complex metabolic systems is that mutations with small effects exist as a consequence of the typically small flux control coefficient relating enzyme activity to the rate of a metabolic process. Although a slightly detrimental mutation is somewhat less likely to become fixed by chance than a slightly favorable mutation, mutations that are slightly detrimental might be expected to be more numerous than favorable mutations owing to the previous incorporation of favorable mutations by a long history of natural selection. The result is that, as Ohta has pointed out, a significant fraction of mutations that are fixed in evolution are slightly detrimental. In the long run, the fixation of detrimental mutations in a gene increases the opportunity for the occurrence of a compensatory favorable mutation, either in the same gene or in an interacting gene. On a suitably long timescale, therefore, every gene incorporates favorable mutations that compensate for detrimental mutations previously fixed. This form of evolution is driven primarily by natural selection, but it results in no change or permanent improvement in enzymatic function.

Adaptation, Physiological↗
PubMed1996

Sequence scanning: A method for rapid sequence acquisition from large-fragment DNA clones.

A strategy of "sequence scanning" is proposed for rapid acquisition of sequence from clones such as bacteriophage P1 clones, cosmids, or yeast artificial chromosomes. The approach makes use of a special vector, called LambdaScan, that reliably yields subclones with inserts in the size range 8-12 kb. A number of subclones, typically 96 or 192, are chosen at random, and the ends of the inserts are sequenced using vector-specific primers. Then long-range spectrum PCR is used to order and orient the clones. This combination of shotgun and directed sequencing results in a high-resolution physical map suitable for the identification of coding regions or for comparison of sequence organization among genomes. Computer simulations indicate that, for a target clone of 100 kb, the scanning of 192 subclones with sequencing reads as short as 350 bp results in an approximate ratio of 1:2:1 of regions of double-stranded sequence, single-stranded sequence, and gaps. Longer sequencing reads tip the ratio strongly toward increased double-stranded sequence.

Animals↗
PubMed1996

EST! EST!! EST!!!

A recently published study has identified a set of candidate genes for human diseases based on findings from Drosophila. Each human expressed sequence tag (EST) in a large database was compared with all known Drosophila genes. After eliminating matches between genes of already known function, the remaining sequences were mapped in the human genome. In each region, the phenotypes of all known human diseases were compared with the phenotypes of known Drosophila mutations in order to identify candidate genes for the human diseases. Are the correspondences real or coincidental?

Animals↗
PubMed1996

Molecular phylogeny of Azteca ants (Hymenoptera:Formicidae) and the colonization of Cecropia trees.

Despite the prominence of the Azteca-Cecropia interaction as the focus of extensive ecological investigation, a reliable phylogeny of the Azteca ants has been lacking, primarily because many of the morphological and behavioral characters are phylogenetically uninformative or conflicting. A phylogenetic analysis of a select set of Azteca ants, including six Cecropia inhabitants and two non-Cecropia inhabitants, plus an outgroup taxon, is presented on the basis of mitochondrial DNA sequences. The evolutionary relationships deduced from the molecular data are analyzed with reference to ecological and morphological studies, specifically addressing the phylogenetic relationship of structurally an behaviorally ambiguous taxa, species complex groupings, and the colonization of Cecropia trees. According to the molecular phylogeny, the Cecropia-inhabiting Azteca do not form a monophyletic clade, indicating multiple independent colonization or abandonment of Cecropia trees by the Azteca.

Animals↗