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PubMed2000

Rapid in vivo evolution of a beta-lactamase using phagemids.

RNA viruses are capable of undergoing extremely rapid evolution due to their high rates of reproduction, small genome size, and a high frequency of spontaneous mutagenesis. Here we demonstrate that a virus-like, evolutionary state can be created by propagating a phagemid population in a hypermutator strain of Escherichia coli in the presence of a helper phage. This enables one to subject individual phagemid-encoded genes to rapid in vivo evolution. We applied this approach to TEM-1 beta-lactamase which confers resistance to 0.05 mg/L of the antibiotic cefotaxime. After 3 weeks of in vivo evolution we were able to isolate a double mutant, E104K/G238S, of the enzyme which confers a 500-fold increased level of resistance to cefotaxime compared to the starting enzyme. In two independent experiments we obtained a triple mutant, E104K/G238S/T263M, which confers a 1000-fold increase in resistance compared to the wild type enzyme. The same three mutations have been previously observed in TEM-4 beta-lactamase which was discovered in a highly cefotaxime-resistant clinical isolate. The probability of randomly obtaining a beta-lactamase carrying three identical point mutations is less than 10(-10). This indicates that phagemid evolution can rapidly reproduce evolution occurring in nature.

Bacteriophages↗

PubMed1992

Cranial capacity evolution in Homo erectus and early Homo sapiens.

This paper investigates patterns of cranial capacity evolution in Homo erectus, early Homo sapiens, and in regional subsamples of H. erectus. Specifically, models explaining evolution of cranial capacity in these taxa are evaluated with statistical techniques developed for the analysis of time series data. Regression estimates of rates of evolution in cranial capacity are also obtained. A non-parametric test for trend suggests that cranial capacity in both H. erectus and early H. sapiens may increase significantly through time. Cranial capacity in an Asian subsample of H. erectus (comprised of Chinese and Indonesian specimens) increases significantly through time. Other subsamples of H. erectus (African, Chinese, and Indonesian) do not appear to increase significantly through time. Regression results generally corroborate results of the test for trend. Spatial and temporal variation may characterize evolution of cranial capacity in H. erectus. Different patterns of cranial capacity evolution may distinguish H. erectus from early H. sapiens.

Africa↗

PubMed2001

Directed evolution of operon of trehalose-6-phosphate synthase/phosphatase from Escherichia coli.

Trehalose is a nonspecific protective agent for biomacromolecules. Trehalose-6-phosphate synthase (OtsA)/phosphatase (OtsB), which is encoded by the gene operon otsBA located at -42 of the Escherichia coli genome, is the main enzyme system that catalyzes the synthesis of trehalose in E. coli. We cloned the operon and modified it by directed evolution. Unlike in the previously reported work, we modified the whole operon and screened the positive mutant simultaneously. Thus we believe that the gene complex solves the negative effects between two enzymes if one of them diversifies its structure or functions and finds the form most suitable for trehalose synthesis. It thus mimics the natural process, in which the functional improvement of organisms is related to alterations in coordinated enzymes. The evolution procedure was carried out in a sequence of error-prone PCR, shuffling PCR, and then strict screening of the mutants. After screening of a library of more than 4000 colonies, about 15 positive colonies were analyzed, resulting in a higher concentration of trehalose than control. One of them, E. coli TS7, shows 12.3-fold higher trehalose synthesis ability than E. coli DH5alpha. In contrast, we introduced the cDNA sequence of the tps1 gene from Saccharomyces cerevisiae, which has 54% identity with the gene otsA, as one of the templates in shuffling PCR. By hybrid evolution and screening, we obtained 10 positive colonies with higher concentrations of trehalose than control. E. coli TS22 appears to have 5.3-fold higher trehalose synthesis ability than E. coli DH5alpha and 1.6-fold more than E. coli DEF3(pOTS11). This result demonstrated that coevolution and hybrid evolution, as powerful protocols in protein engineering, are effective in modifying enzyme. It indicates that repeating the process of genomic evolution in nature is feasible.

Directed Molecular Evolution↗

PubMed2000

Co-evolution of proteins with their interaction partners.

The divergent evolution of proteins in cellular signaling pathways requires ligands and their receptors to co-evolve, creating new pathways when a new receptor is activated by a new ligand. However, information about the evolution of binding specificity in ligand-receptor systems is difficult to glean from sequences alone. We have used phosphoglycerate kinase (PGK), an enzyme that forms its active site between its two domains, to develop a standard for measuring the co-evolution of interacting proteins. The N-terminal and C-terminal domains of PGK form the active site at their interface and are covalently linked. Therefore, they must have co-evolved to preserve enzyme function. By building two phylogenetic trees from multiple sequence alignments of each of the two domains of PGK, we have calculated a correlation coefficient for the two trees that quantifies the co-evolution of the two domains. The correlation coefficient for the trees of the two domains of PGK is 0. 79, which establishes an upper bound for the co-evolution of a protein domain with its binding partner. The analysis is extended to ligands and their receptors, using the chemokines as a model. We show that the correlation between the chemokine ligand and receptor trees' distances is 0.57. The chemokine family of protein ligands and their G-protein coupled receptors have co-evolved so that each subgroup of chemokine ligands has a matching subgroup of chemokine receptors. The matching subfamilies of ligands and their receptors create a framework within which the ligands of orphan chemokine receptors can be more easily determined. This approach can be applied to a variety of ligand and receptor systems.

Chemokines↗

PubMed1999

The evolution of the scrotum and testicular descent in mammals: a phylogenetic view.

The adaptive significance of the scrotum and the evolution of the descent of the testicles and epididymis have been a focus of interest among biologists for a long time. In this paper we use three anatomical character states of the scrotum and descensus: (1) testicles descended and scrotal; (2) testicles descended but ascrotal; (3) testicles not descended (testicondy). These states are then mapped on an up to date phylogeny of the Mammalia. Three main points arise out of this mapping procedure: (1) the presence of a scrotum is either primitive in extant Mammalia or primitive within eutherian mammals except Insectivora; (2) evolution has generally proceeded from a scrotal condition to progressively more ascrotal; (3) loss of testicular descensus is less common in mammalian evolution than is loss of the scrotum. In the light of these findings we discuss some current hypotheses regarding the origin and evolution of the scrotum. We find that these are all incomplete in so far as it is not the presence of the scrotum in various mammal groups that requires explaining. Instead, it is the reverse process, why the scrotum has been lost in so many groups, that should be explained. We suggest that the scrotum may have evolved before the origin of mammals, in concert with the evolution of endothermy in the mammalian lineage, and that the scrotum has been lost in many groups because descensus in many respects is a costly process that will be lost in mammal lineages as soon as an alternative solution to the problem of the temperature sensitivity of spermatogenesis is available.

Animals↗

PubMed1996

The variety of human virus evolution.

There have been many wise suggestions of ways that evolution may occur but those ways seem often hard to support with good examples. Viruses have proven to be replete with some of these items. This paper reviews work that shows: (1) very fast rates of evolution; (2) positive Darwinian selection with the selective pressure specifically identified; (3) viral reassortment; (4) grossly unequal rates of evolution depending upon the host of the virus; (5) accurate dating of the cenancestor, the most recent common ancestor; (6) correspondence between the evolutionary tree and the geography of the place of isolation; (7) punctuated molecular evolution; and (8) network evolution.

Animals↗

PubMed1996

Molecular evolution and phylogenetic utility of the polyubiquitin locus in mammals and higher vertebrates.

The product of ubiquitin genes is a small protein involved in intracellular sorting of other proteins. The locus consists of tandemly arrayed, uninterrupted copies of the gene. As several studies have noted, the Polyubiquitin locus is a model system for studying concerted evolution. While the protein is among the most conserved known, individual copies within an organism show variation in nucleotide sequence despite clear evidence of concerted evolution. When treated as individuals, repeats from a given locus form a monophyletic group. Furthermore adjacent copies often cluster, suggestive of the mechanism of concerted evolution. Due to this concerted evolution of repeats (and loci in organisms with multiple polyubiquitins), sequencing of heterogeneous PCR products consisting of all the repeats in a given organism may yield phylogenetic signal, as with other multicopy genes. We test this possibility through 22 original sequences using primers designed so that only tandem copies are amplified. Using these and previously published data, we further explore these phenomena in higher vertebrates and mammals in particular. We suggest that multiple locus duplications have occurred within mammals. Positional codon bias is strongly evident. We also find substitutional bias with regard to codon type. GC content of the locus appears to be generally high across vertebrates. Intraorganismal variation is tallied as an indication of frequency of change in codon position and transition/transversion ratios to further elucidate the tempo and mode of molecular evolution. Using these data, a weighting scheme for ubiquitin is also presented. Despite the gene's high GC content, transitional changes still appear more frequent. While the phylogenetic utility of ubiquitin does not appear great, its mechanistic insights seem far from exhausted.

Amino Acid Sequence↗

PubMed1999

Mosaic evolution of ruminant stomach lysozyme genes.

The genomes of ruminant artiodactyls, such as cow and sheep, have approximately 10 lysozyme genes, 4 of which are expressed in the stomach. Most of the duplications of the lysozyme genes occurred 40-50 million years ago, before the divergence of cow and sheep. Despite this, the coding regions of stomach lysozyme genes within a species (e.g., cow, sheep, or deer) are more similar to each other than to lysozyme genes in other ruminants. This observation suggests that the coding regions of the stomach lysozyme genes have evolved in a concerted fashion. Our previous characterization of 3 cow stomach lysozyme genes suggested that it was only the coding exons that had participated in concerted evolution. To determine whether the introns and flanking regions of ruminant stomach lysozyme genes are evolving in a concerted or a divergent fashion, we have isolated and characterized 2 sheep stomach lysozyme genes. Comparison of the sequences of the sheep and cow stomach lysozyme genes clearly shows that the introns and flanking regions have evolved, like the 3' untranslated region of the mRNAs, in a divergent manner. Thus, if the four coding exons are evolving by concerted evolution, then a mosaic pattern of concerted and divergent evolution is occurring in these genes. The independent concerted evolution of coding exons of the ruminant stomach lysozyme gene may have assisted in the accelerated adaptive evolution of the lysozyme to new function in the early ruminant.

3' Untranslated Regions↗

PubMed2000

Evolution of Hoxa-11 in lineages phylogenetically positioned along the fin-limb transition.

HOXA11 is a transcription factor implicated in paired appendage development. To identify signatures of evolutionary change in the structural, and putative functional, domains of HOXA11, we studied its evolution in tetrapod and nontetrapod lineages that represent approximately 1.5 billion years of evolutionary time. Here, Hoxa-11 gene proper sequences were determined for frog (Xenopus tropicalis), coelacanth (Latimeria chalumnae), common zebrafish (Danio rerio; Hoxa-11a and Hoxa-11b paralogs), and giant zebrafish (D. aequipinnatus; Hoxa-11b) and aligned against previously published Hoxa-11 sequences of human, mouse, chick, and newt. Based on aligned Hoxa-11 amino acid sequences, the protein was demarcated into three segments: Domains I (N-terminal) and III (homeobox + C-terminal), which varied slightly in rates and patterns of evolution, and a variable, overall hydrophilic region (HyD), which partially overlaps with Domain I. As judged by character reconstructions of HOXA11 Domains I and III, no significant changes in rates of coding sequence evolution occurred in tetrapods (frog and chick), relative to coelacanth (a lobe-finned fish), i.e., across the fin-limb transition. Accelerated rates of Hoxa-11 coding sequence evolution were observed for the mammalian and newt lineages. This was shown to be a gene-specific phenomenon. The duplicated Hoxa-11a and Hoxa-11b genes of zebrafish exhibited accelerated rates of evolution and accumulated substitutions at sites that are conserved among coelacanth and all tetrapods examined. Amino acid sequence comparisons of the HyD of HOXA11 suggested that a putative repressor subdomain, containing stretches of consecutive alanine residues, emerged within the tetrapods. A high degree of nucleotide conservation in the 5' half of the Hoxa-11 intron was observed for tetrapod and nontetrapod lineages. Using electrophoretic mobility shift assays, a 35-bp intron sequence, which is 100% conserved in all Hoxa-11 loci except for the zebrafish Hoxa-11a paralog, was found to bind protein(s) in HeLa and chick whole-cell extracts.

Alanine↗

PubMed1994

Genome evolution: between the nucleosome and the chromosome.

Intermediate between DNA sequences and broad patterns of karyotypic change there is a major gap in understanding genome structure and evolution. The gap is at the megabase level between genes and chromosomes. New methods for analyzing large DNA fragments cloned in yeast or bacterial vectors provide experimental access to genome evolution at the megabase level by enabling the assembly of megabase-size contiguous regions. Genome evolution at the megabase level can also be studied using high-resolution genetic maps. Rates and patterns of genome evolution in mammals (mouse versus humans) and Drosophila (D. virilis versus D. melanogaster) are compared and contrasted. Opportunities for research in genome evolution using the new technologies are enumerated and discussed.

Animals↗

PubMed1994

Molecular evolution of interleukin-3.

Chimpanzee, tamarin, and marmoset interleukin-3 (IL-3) genes were cloned, sequenced, and expressed. Western blot analysis demonstrated that functional genes were isolated. IL-3 sequences were compared with those of mouse, rat, rhesus monkey, gibbon, and man. Multiple alignment of the IL-3 coding regions showed that only a few regions had been conserved during mammalian evolution, which are likely associated with functional domains of the IL-3 protein. Substitution rates for the various lineages were calculated and the numbers of synonymous and nonsynonymous substitutions were estimated separately. Distance matrices of the IL-3 coding regions were used to construct phylogenetic trees which revealed large differences in IL-3 evolution rate as well as a more rapid substitution rate for rodents and a rate slowdown during hominoid evolution. Extremes were rhesus monkey IL-3, which accumulated few synonymous substitutions, and gibbon IL-3, which had almost exclusively synonymous substitutions. In rhesus monkey IL-3, nonsynonymous substitutions outnumbered synonymous substitutions, which could not be readily explained by a random process of substitutions. We assume that during evolution of IL-3, the majority of the amino acid replacements and the impaired interspecies functional cross-reactivity originate from selection mechanisms with the most likely selective force being the structure of the heterodimeric IL.3 cell-surface receptor. Insight into IL-3 architecture and structural analysis of the IL-3 receptor are needed to analyze the unusually fast evolution of IL-3 in more detail.

Animals↗

PubMed1994

Evolution of murine alpha 1-proteinase inhibitors: gene amplification and reactive center divergence.

The organization and sequence of genes encoding the alpha 1-proteinase inhibitor (alpha 1PI), a major serine proteinase inhibitor of the mammalian bloodstream, have been compared in several species, including murine rodents (genus Mus). Analysis of gene copy number indicates that amplification of alpha 1PI genes occurred at some time during evolution of the Mus genus, leading to fixation of a family of about three to five genes in several existing species (e.g., M. domesticus and M. saxicola), and only a single gene in others (e.g., M. caroli). A phylogeny for the various mammalian alpha 1PI mRNAs was constructed based upon synonymous substitutions within coding regions. The mRNAs in different murine species diverged from a common ancestor before the formation of the first species lineages of the Mus genus, i.e., about 10-13 million years ago. Thus, alpha 1PI gene amplification must have occurred prior to Mus speciation; gene families were retained in some, but not all, murine species. The reactive center region of the alpha 1PI polypeptide, which determines target protease specificity, has diverged rapidly during evolution of the Mus species, but not during evolution of other mammalian species included in the analysis. It is likely that this accelerated evolution of the reactive center, which has been noted previously for serine proteinase inhibitors, was driven by some sort of a positive Darwinian selection that was exerted in a taxon-specific manner. We suggest that evolution of alpha 1PI genes of murine rodents has been characterized by both modification of gene copy number and rapid reactive center divergence. These processes may have resulted in a broadened repertoire of proteinase inhibitors that was evolutionarily advantageous during Mus speciation.

Amino Acid Sequence↗

PubMed1990

Evolution of the Simiiformes and the phylogeny of human chromosomes.

This paper is based on the results of Primate chromosome studies obtained using high resolution techniques in our and other laboratories. We discuss the origin and the evolution of the chromosomes in the human karyotype and the time in evolution of the Simiiformes when they acquired their present morphology. Our results indicate that the chromosomes that underwent a higher number of reorganizations during the evolution of the Simiiformes coincide with the chromosomes most often implicated in human chromosome pathology. We describe the main reorganizations that took place during Primate evolution. Centromere activation and inactivation and heterochromatin changes are discussed as mechanisms of chromosome evolution.

Animals↗

PubMed1979

Chromosomal evolution in primates: tentative phylogeny from Microcebus murinus (Prosimian) to man.

The karyotypes of more than 60 species of Primates are studied and compared, with the use of almost all existing banding techniques. There is a very close analogy of chromosome banding between the Simians studied and man. The quantitative or qualitative variations detected all involve the heterochromatin. It is very likely that all the euchromatin (nonvariable R and Q bands) is identical in all the species. Approximately 70% of the bands are common to the Simians and to the Lemurs (Prosimians). In the remaining 30%, technical difficulties prevented a valuable comparison, but this does not exclude the possibility that a complete analogy may exist. Thus, it is very likely that chromosomal evolutions of the Simians, and probably of all the Primates, has occurred without duplication or deficiency of the euchromatin. Approximately 150 rearrangements could be identified and related to the human chromosomes. The types of rearrangement vary from one group (suborder, family, genus) to another. For instance, Robertsonian translocations are preponderant among the Lemuridae (44/57), but are nonexistent among the Pongidae. Chromosome fissions are very frequent amng the Cercopithecidae (10/23), but were not found elsewhere, and pericentric inversions are preponderant in the evolution of Pongidae and man (17/28). This suggest that the chromosomal evolution may be directed by the genic constitution (favouring the occurrence of a particular type of rearrangement, by enzymatic reaction), by the chromosomal morphology (the probability that Robertsonian translocation will be formed depends at least partially on the number of acrocentrics), and by the reproductive behaviour of the animals. Reconstitution of the sequence of the chromosomal rearrangements allowed us to propose a fairly precise genealogy of many Primates, giving the positions of the Catarrhines, the Platyrrhines, and the Prosimians. It was also possible to reconstruct the karyotypes of ancestors that died out several dozen million years ago. The possible role of chromosomal rearrangements in evolution is discussed. It appears necessary to consider different categories of rearrangements separately, depending on their behaviour. The 'nonfavoured' rearrangements, such as pericentric inversions, need to occur in an isolated small population for implanting, by an equivalent of genic derivation. The 'favoured' rearrangements, e.g., Robertsonian translocations, may occur and diffuse in panmictic populations, and accumulate. Their role of gametic barrier could be much more progressive. For discrimination between these two categories, it was necessary to differentiate the selective advantage or disadvantage of the rearrangement itself. It was not possible to show that chromosomal rearrangements play a direct role in modification of the phenotype by position effect. Comparison of the rearrangement that have occurred during evolution and those detected in the human population shows a strong correlation for some of them...

Animals↗

PubMed1991

Parallel evolution in mammalian and avian brains: comparative cytoarchitectonic and cytochemical analysis.

Comparative morphology, which is based on the selection theory of evolution, analyses the impact of function upon structure and, therefore, emphasizes the adaptive events and biological advantage during the evolution of organs. A comparison based on analogies is described here as an adequate method. The hypothesis is proposed that the evolution of the brain follows the same trends in birds as in mammals. This hypothesis is proved by (1) allometric studies of brain weight and brain structure volume in relation to body weight in mammals and birds; (2) architectonic studies using image analysis on cell and fibre stains as well as on histochemical preparations and receptor autoradiography; and (3) hodological studies with injections of [3H]leucin, HRP and WGA-HRP. The results reveal a vast amount of structural and functional similarities in avian and mammalian brain organization, especially an expansion of structures that permit multimodal integration capacity in the telencephalon. Thus, a parallel evolution occurred in these two groups of vertebrates. It is argued that this may be a general phenomenon in evolution. A cladistic approach, which is based on the concept of homologies (plesio-, apomorphies), pushes aside the existence of analogies. For this reason, cladism does not seem to be a method to answer questions of evolutionary morphology adequately.

Animals↗

PubMed1995

A short introduction to the origin and molecular evolution of viruses.

The present review deals with conceptual and experimental approaches to two aspects of the origin and molecular evolution of viruses. In the section "Role of Retrons, Retroelements, and Reverse Transcriptase in the Evolution of Retroviruses and in Eukaryotic Genome Plasticity", Temin's concept that retrons are an ancient genetic element that during evolution of the species gave rise to retroviruses is presented. An opposing view of Xiong and Eickbush that the most probable ancestor of current retroelements is a retrotransposable element with gag- and pol-like genes is presented. Minus-strand RNA viruses are also discussed. The second aspect of this review is the molecular evolution of viruses at the level of the virus genome. Spiegelman's experiment on the evolution of self-replicating nucleic acid molecules outside living cells and Eigen's experimental and conceptual approaches to this subject are presented, along with studies on the evolutionary rates of base substitutions in viral RNA and defective molecules generated during replication.

Animals↗

PubMed1976

Evolution of DNA structure: direction, mechanism, rate.

On the basis of the results of an analysis of frequencies of pyrimidine oligonucleotides, the degree of pyrimidine clustering of DNA in species from different taxa has been determined. A tendency for an increase in the index of clustering of DNA was revealed in the sequence: invertebrates, fishes, amphibians, reptiles, birds, mammals. A mechanism is postulated, according to which the increase in the degree of clustering of DNA d-ring the evolution may be associated with the accumulation of mutations, Purine equalibrium Pyrimidine transversions, resulting in a selective enrichment of one of the chains of DNA with pyrimidines and the other- with purines, i.e. in an increase in the degree of purine-pyrimidine imbalance (asymmetry) of DNA complementary chains. This mechanism of DNA evolution is supported by the presence of positive correlation between the degree of clustering and the degree of the chain asymmetry of natural DNAs, as well as the character of the amino acid substitutions in cytochromes c in different species. The progressive evolution of different groups of organisms on the whole may have been accompanied by an acceleration of the rates of evolution of the DNA structure. On the basis of the amino acid sequence of cytochromes c in different species the degree of clustering and the degree of the chain asymmetry of the corresponding structural genes of DNA was found to have a general tendency towards an increase in the following order: invertebrates, fishes, amphibians, reptiles, birds, mammals. Thus, evolution of cytochrome c cistron is a vector process based on a selection of mutations which, on the one hand, are neurtral to protein, and, on the other hand, result in the sense chain of DNA being enriched with pyrimidines and the nonsense one (and the corresponding mRNA)- with purines. Hence, it is the polynucleotide template rather than protein, that must have been the "object of selection". The frequency of substitutions in cytochromes c cistron for vertebrates is 1.56x13(-9) per nucleotide per year. It is believed that the evolutionary modification of the DNA structure may be associated with an increase in the interference resistance of the translation, i.e. with selection for codons of highest readout stability.

Amino Acid Sequence↗

PubMed1996

The molecular evolution of vertebrate growth hormones: a pattern of near-stasis interrupted by sustained bursts of rapid change.

It has been demonstrated previously that in mammals the evolution of pituitary growth hormone shows an unusual pattern, with an underlying slow rate and at least two sustained bursts of rapid evolution (in the artiodactyls and primates), during which the rate increased at least 25-fold. It is demonstrated here that a similar pattern applies for growth hormone evolution throughout the vertebrates, with a basal rate similar to that seen in mammals, but bursts of rapid evolution in the amphibia and the elasmobranchs, and several bursts in the teleosts. The placental growth-hormone-like proteins of primates show a similar pattern. It is argued that the bursts of evolution seen for growth hormone are a consequence of selection and that this may reflect changes in the functions of the hormone additional to its basic growth-promoting actions.

Animals↗