Biomedical subjects

H A Wichman

Publications and source records attributed to H A Wichman.

At least 19 recordsLinked to original sources

PubMed2002

Distribution of LINEs and other repetitive elements in the karyotype of the bat Carollia: implications for X-chromosome inactivation.

The Lyon repeat hypothesis postulates that long interspersed elements (LINEs) play a role in X-chromosome inactivation. Evidence to support this hypothesis includes the observation that the degree of inactivation of autosomes translocated to the X chromosome is correlated with LINE density on that autosome. We examined the distribution of LINEs in the fruit bat Carollia brevicauda, which has an autosomal translocation to the X that occurred at least 7 million years ago. A quantitative analysis of LINE accumulation on multiple metaphase chromosome spreads revealed a significant accumulation on the original X relative to the attached autosome, the homolog of that autosome (Y(2)), and chromosome 1. Previous replication studies indicate that for the X and attached autosome, only the original X chromosome replicates late in Carollia females, and that the attached autosome replicates in the same timeframe as other autosomes. These data are compatible with the Lyon repeat hypothesis, and the possibility that LINEs act as booster elements for X inactivation remains a viable hypothesis. We address the procedures and limitations of quantitative analysis based on in situ hybridization.

Animals↗

PubMed2001

An ancient retrovirus-like element contains hot spots for SINE insertion.

Vertebrate retrotransposons have been used extensively for phylogenetic analyses and studies of molecular evolution. Information can be obtained from specific inserts either by comparing sequence differences that have accumulated over time in orthologous copies of that insert or by determining the presence or absence of that specific element at a particular site. The presence of specific copies has been deemed to be an essentially homoplasy-free phylogenetic character because the probability of multiple independent insertions into any one site has been believed to be nil. Mys elements are a type of LTR-containing retrotransposon present in Sigmodontine rodents. In this study we have shown that one particular insert, mys-9, is an extremely old insert present in multiple species of the genus Peromyscus. We have found that different copies of this insert show a surprising range of sizes, due primarily to a continuing series of SINE (short interspersed element) insertions into this locus. We have identified two hot spots for SINE insertion within mys-9 and at each hot spot have found that two independent SINE insertions have occurred at identical sites. These results have major repercussions for phylogenetic analyses based on SINE insertions, indicating the need for caution when one concludes that the existence of a SINE at a specific locus in multiple individuals is indicative of common ancestry. Although independent insertions at the same locus may be rare, SINE insertions are not homoplasy-free phylogenetic markers.

Alleles↗

PubMed2000

Experimental evolution recapitulates natural evolution.

Genomes of the closely related bacteriophages phiX174 and S13 are 5386 bases long and differ at 114 nucleotides, affecting 28 amino acids. Both parental phages were adapted to laboratory culture conditions in replicate lineages and analysed for nucleotide changes that accumulated experimentally Of the 126 experimental substitutions, 90% encoded amino-acid changes, and 62% of the substitutions occurred in parallel in more than one experimental line. Furthermore, missense changes at 12 of the experimental sites were at residues differing between the parental phages; in ten cases the phiX174 experimental lineages were convergent with the S13 parent, or vice versa, at both the nucleotide and amino-acid levels. Convergence at a site was even obtained in both directions in three cases. These results point to a limited number of pathways taken during evolution in these viruses, and also raise the possibility that much of the amino-acid variation in the natural evolution of these viruses has been selected.

Bacteriophage phi X 174↗

PubMed2000

Evolutionary reversals during viral adaptation to alternating hosts.

Experimental adaptation of the bacteriophage phiX174 to a Salmonella host depressed its ability to grow on the traditional Escherichia host, whereas adaptation to Escherichia did not appreciably affect growth on Salmonella. Continued host switching consistently exhibited this pattern. Growth inhibition on Escherichia resulted from two to three substitutions in the major capsid gene. When these phages were forced to grow again on Escherichia, fitness recovery occurred predominantly by reversions at these same sites, rather than by second-site compensatory changes, the more frequently observed mechanism in most microbial systems. The affected residues lie on the virion surface and they alter attachment efficiency, yet they occur in a region distinct from a putative binding region previously identified from X-ray crystallography. These residues not only experienced high rates of evolution in our experiments, but also exhibited high levels of radical amino acid variation among phiX174 and its known relatives, consistent with a history of adaptation involving these sites.

Adaptation, Physiological↗

PubMed2000

The end of the LINE?: lack of recent L1 activity in a group of South American rodents.

L1s (LINE-1: Long Interspersed Nuclear Element 1) are present in all mammals examined to date. They occur in both placental mammals and marsupials and thus are thought to have been present in the genome prior to the mammalian radiation. This unusual conservation of a transposable element family for over 100 million years has led to speculation that these elements provide an advantage to the genomes they inhabit. We have recently identified a group of South American rodents, including rice rats (Oryzomys), in which L1s appear to be quiescent or extinct. Several observations support this conclusion. First, genomic Southern blot analysis fails to reveal genus-specific bands in Oryzomys. Second, we were unable to find recently inserted elements. Procedures to enrich for young elements did not yield any with an intact open reading frame for reverse transcriptase; all elements isolated had numerous insertions, deletions, and stop codons. Phylogenetic analysis failed to yield species-specific clusters among the L1 elements isolated, and all Oryzomys sequences had numerous private mutations. Finally, in situ hybridization of L1 to Oryzomys chromosomes failed to reveal the characteristic L1 distribution in Oryzomys with either a homologous or heterologous probe. Thus, Oryzomys is a viable candidate for L1 extinction from a mammalian host.

Animals↗

PubMed2000

Big-benefit mutations in a bacteriophage inhibited with heat.

High temperature inhibits the growth of the wild-type bacteriophage phiX174. Three different point mutations were identified that each recovered growth at high temperature. Two affected the major capsid protein (residues F188 and F242), and one affected the internal scaffolding protein (B114). One of the major capsid mutations (F242) is located in a beta strand that contacts B114 in the procapsid during viral maturation, whereas the other capsid mutation (F188) is involved in subunit interactions at the threefold axis of symmetry. Selective coefficients of these mutations ranged from 13.9 to 3.8 in the inhibitory, hot environment, but all mutations reduced fitness at normal temperature. The selective effect of one of the mutations (F242) was evaluated at high temperature in four different genetic backgrounds and exhibited epistasis of diminishing returns: as log fitness of the background genotype increased from -0.1 to 4.1, the fitness boost provided by the F242 mutation decreased from 3.9 to 0. 8. These results support a model in which viral fitness is bounded by an upper limit and the benefit of a mutation is scaled according to the remaining opportunity for fitness improvement in the genome.

Bacteriophage phi X 174↗

PubMed2000

Isolation of markers from recently transposed LINE-1 retrotransposons.

Many previous techniques for the isolation of endogenous retroelements such as LINE-1 retrotransposons have produced major sampling bias or required laborious procedures. These problems led to the isolation of only older elements in some cases. In other cases, specialized systems were required for the isolation of recently transposed elements. We report here a system for the easy isolation of markers from a wide range of LINE-1 elements and the screening of recently transposed elements from that population. This is accomplished by the use of PCR with degenerate primers specific for conserved regions of the reverse transcriptase gene, a modified screening vector, and a refined blue/white colony assay that screens for amplified DNA containing open reading frames. This method should be applicable to searches for endogenous retroviruses.

Amino Acid Sequence↗

PubMed1999

Different trajectories of parallel evolution during viral adaptation.

The molecular basis of adaptation is a major focus of evolutionary biology, yet the dynamic process of adaptation has been explored only piecemeal. Experimental evolution of two bacteriophage lines under strong selection led to over a dozen nucleotide changes genomewide in each replicate. At least 96 percent of the amino acid substitutions appeared to be adaptive, and half the changes in one line also occurred in the other. However, the order of these changes differed between replicates, and parallel substitutions did not reflect the changes with the largest beneficial effects or indicate a common trajectory of adaptation.

Adaptation, Physiological↗

PubMed1998

Molecular evolution of two lineages of L1 (LINE-1) retrotransposons in the california mouse, Peromyscus californicus.

The large number of L1 [long interspersed elements (LINE)-1] sequences found in the genome is due to the insertion of copies of the retrotransposon over evolutionary time. The majority of copies appear to be replicates of a few active, or "master" templates. A continual replacement of master templates over time gives rise to lineages distinguishable by their own unique set of shared-sequence variants. A previous analysis of L1 sequences in deer mice, Peromyscus maniculatus and P. leucopus, revealed two active L1 lineages, marked by different rates of evolution, whose most recent common ancestor predates the expansion of the Peromyscus species. Here we exploit lineage-specific, shared-sequence variants to reveal a paucity of Lineage 2 sequences in at least one species, P. californicus. The dearth of Lineage 2 copies in P. californicus suggests that Lineage 2 may have been unproductive until after the most recent common ancestor of P. californicus and P. maniculatus. We also show that Lineage 1 appears to have a higher rate of evolution in P. maniculatus relative to either P. californicus or P. leucopus. As a phylogenetic tool, L1 lineage-specific variants support a close affinity between P. californicus and P. eremicus relative to the other species examined.

Animals↗

PubMed1997

Analysis of orthologous retrovirus-like elements in the white-footed mouse, Peromyscus leucopus.

Three loci in the genome of the white-footed mouse, Peromyscus leucopus, were examined for the presence or absence of orthologous copies of the retrovirus-like element mys using polymerase chain reaction. We examined these loci in 28 mice collected throughout the P. leucopus species range. Mys insertions were present in only one of the individuals examined at the mys-1 and mys-7 loci. Conversely, the mys-6 element was found in several individuals, but the presence of this element was limited to northern latitudes. Because the long terminal repeats (LTRs) of a given element are expected to be identical at the time of retrotransposition into the genome, and to accumulate changes over evolutionary time, within-element LTR sequence comparisons can be used to estimate the relative age of insertions. Within-element LTR differences are greater in mys-6 than in mys-1 or mys-7. The LTRs from orthologous mys-6 elements of six mice were sequenced. The alignment revealed 13 of the 22 differences between the right and left LTRs that were shared by all orthologous mys-6 sites, suggesting that relative to its time of insertion into the genome, mys-6 has only recently spread across the northern part of the species range.

Animals↗

PubMed1997

Exceptional convergent evolution in a virus.

Replicate lineages of the bacteriophage phiX 174 adapted to growth at high temperature on either of two hosts exhibited high rates of identical, independent substitutions. Typically, a dozen or more substitutions accumulated in the 5.4-kilobase genome during propagation. Across the entire data set of nine lineages, 119 independent substitutions occurred at 68 nucleotide sites. Over half of these substitutions, accounting for one third of the sites, were identical with substitutions in other lineages. Some convergent substitutions were specific to the host used for phage propagation, but others occurred across both hosts. Continued adaptation of an evolved phage at high temperature, but on the other host, led to additional changes that included reversions of previous substitutions. Phylogenetic reconstruction using the complete genome sequence not only failed to recover the correct evolutionary history because of these convergent changes, but the true history was rejected as being a significantly inferior fit to the data. Replicate lineages subjected to similar environmental challenges showed similar rates of substitution and similar rates of fitness improvement across corresponding times of adaptation. Substitution rates and fitness improvements were higher during the initial period of adaptation than during a later period, except when the host was changed.

Bacteriophage phi X 174↗

PubMed1996

Retrotransposon Mys was active during evolution of the Peromyscus leucopus-maniculatus complex.

Mys is a retrovirus-like transposable element found throughout the genus Peromyscus. Several mys subfamilies identified on the basis of restriction site variation occur in more than one species. The distribution of these subfamilies is consistent with the accepted species phylogeny, suggesting that mys was present in the ancestor of Peromyscus and has been active through much of the evolution of this genus. Quantitative Southern blot analysis was used to examine the variability of subfamilies in P. leucopus and maniculatus. We found that subfamilies with phylogenetically narrow distributions were more variable in copy number both within and between species than subfamilies with a broader distribution. Taken together, our data suggest that mys has undergone multiple rounds of transposition since the peromyscine radiation, and that five subfamilies have been amplified during the evolution of the leucopus-maniculatus species complex.

Animals↗

PubMed1996

Computer simulation of transposable element evolution: random template and strict master models.

It has been proposed that the most extensively studied mammalian retrotransposons replicate by some form of a master template model. This conclusion has been drawn largely from DNA sequence analysis and is based on phylogenetic tree topology, the presence and ordering of shared variants, the degree of divergence between elements within a subfamily, and the shape of the distribution of pairwise differences between elements. To investigate how robust these parameters are as predictors of the model of transposition, computer simulations of the two most extreme transposition models, the Random Template Model and the Strict Master Model, were carried out. A prototype of a computer simulator for studying retrotransposition is presented. The simulator is a versatile digital workbench that maintains DNA sequence data and allows manipulation of a range of factors including reverse transcriptase and in situ mutation rates, transposition template, and transposition rate. All parameters previously used as predictors of the model of transposition were markedly different for the two extreme models when evaluated using large sample sizes of sequences from experiments simulating up to 15 million years of evolution.

Animals↗

PubMed1996

Two persistent LINE-1 lineages in Peromyscus have unequal rates of evolution.

LINE-1, the major family of long, interspersed repeats in the mammalian genome, moves via an RNA intermediate and encodes its own reverse transcriptase. Comparative sequence analysis was used to reconstruct the phylogenetic history of LINE-1 dynamics in the deer mouse, Peromyscus. As is the case in Mus and Rattus, a very small number of active templates produce the majority of LINE-1 copies in Peromyscus. However, in contrast to the single LINE-1 lineage seen in the muroid rodents, Peromyscus has at least two LINE-1 lineages whose most recent common ancestor probably existed before the peromyscine radiation. Species-specific variants of Lineage 1, and intact open reading frames in the youngest elements of both Lineages 1 and 2, suggest that both lineages have remained active within the same genome. The higher number of shared-sequence variants in Lineage 1 relative to Lineage 2 suggests that Lineage 1 has replaced its master template much more frequently than Lineage 2 or that the reverse transcriptase Lineage 1 is more error prone. The implications of the method used to acquire LINE-1 sequences for analysis are discussed.

Animals↗

PubMed1996

Remote ergonomic research in space: spacelab findings and a proposal.

This paper discusses ergonomics research using remotely situated video cameras in spacecraft. Two prototype studies of crewmembers working in the micro-G environments aboard the first two flights of Spacelab are described. Various aspects of crew restraint, stabilization, manipulation of controls, and mobilization were observed, operationally defined, and quantified by observing videotaped scenes of Spacelab crewmembers. In the first study, four performance behaviors were quantified to provide estimates of their frequency of occurrence and variation over the course of each of the flights. The behaviors and their mean percent of observed times were: Hand-Hold 32.2%, Foot Restraint 35.3%, Translation 9.4%, and Struggle 3.7%. Because we observed that nearly a third of a crewmember's time was spent inefficiently holding on with one hand while trying to work with the other, a second study was conducted exploring the use of foot restraints and hand stabilization. During 18 episodes of single-foot restraint, for example, there were 52 instances of hand stabilization and 135 instances of stabilization attempts with the other foot. The paper concludes with some defining characteristics of adequate foot restraints, and a proposal for extending this research model to future spacecraft studies.

Astronauts↗

PubMed1994

Rapidly evolving repetitive DNAs in a conservative genome: a test of factors that affect chromosomal evolution.

The hypothesis that tandemly repeated DNA sequences may facilitate chromosomal rearrangements was tested by comparing a conservatively evolving karyotype of a bat species (Macrotus waterhousii) with data published for a rapidly evolving karyotype of an equid species (Equus zebra). Empirical data generated from the phylogenetic screening of rapidly evolving repetitive DNAs from approximately 0.1% of the M. waterhousii genome showed only one sequence that was repetitive in M. waterhousii but low in copy number or absent from the outgroup Artibeus jamaicensis. This compares to 34 such clones containing sequences which were repetitive in E. zebra but were low in copy number or absent from the outgroup Ceratotherium simum. The bat sequence represents a single family of repeated sequences, whereas six families of sequences were identified in E. zebra. Southern blot analysis suggested that the sequence from M. waterhousii is interspersed rather than tandemly repeated, as are the sequences in E. zebra. These data support the above hypothesis and suggest that species with conservatively evolving karyotypes have fewer numbers and families of rapidly evolving DNA sequences than do species such as the equids that possess a karyotype that is considered to have undergone rapid karyotypic evolution.

Animals↗

PubMed1994

Unusual pattern of bacterial ice nucleation gene evolution.

Bacterial ice nucleation activity (INA+ phenotype) can be traced to the product of a single gene, ina. A remarkably sparse distribution of this phenotype within three bacterial genera indicates that the ina gene may have followed an unusual evolutionary path. Southern blot analyses, coupled with assays for ice-nucleating ability, revealed that within four bacterial species an ina gene is present in some strains but absent from others. Results of hybridization experiments using DNA fragments that flank the ina gene suggested that the genotypic dimorphism of ina may be anomalous. A phylogenetic analysis of 16S ribosomal RNA gene sequences from a total of 14 ina+ and ina- bacterial strains indicated that the ina+ bacteria are not monophyletic but instead phylogenetically interspersed among ina- bacteria. The relationships of ina+ bacteria inferred from ina sequence did not coincide with those inferred from the 16S data. These results suggest the possibility of horizontal transfer in the evolution of bacterial ina genes.

Amino Acid Sequence↗

PubMed1993

Genome organization of repetitive elements in the rodent, Peromyscus leucopus.

To document the frequency and distribution of repetitive elements in Peromyscus leucopus, the white-footed mouse, a cosmid genomic library was examined. Two thousand thirteen randomly chosen recombinants, with an average insert size of 35 kb and representing 2.35% of the haploid genome of P. leucopus, were screened with probes representing microsatellites, tandem repeats, and transposable elements. Of the four dinucleotides, (GT)n was present in 87% of the clones, (CT)n was present in 59% of the clones, and (AT)n and (GC)n each was represented in our sample by a single clone (0.05%). (TCC)n was present in 8% of the clones. Of the tandem repeats, the 28S ribosomal probe and the (TTAGGG)n telomere probe were not represented in the library, whereas a heterochromatic fragment was present in 9% of the clones. A transposable element, mys, was estimated to occur in 4700 copies, whereas a long interspersed element (LINE) was estimated to occur in about 41,000 copies per haploid genome. LINE and mys occurred together in the same clones more frequently than expected on the basis of chance. Hybridizing the library to genomic DNA from P. leucopus, Reithrodontomys fulvescens, Mus musculus, and human produced general agreement between phylogenetic relatedness and intensity of hybridization. However, dinucleotide repeats appeared to account for a disproportionately high number of positive clones in the more distantly related taxa.

Animals↗