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PubMed2001

Mechanisms of evolution in Rickettsia conorii and R. prowazekii.

Rickettsia conorii is an obligate intracellular bacterium that causes Mediterranean spotted fever in humans. We determined the 1,268,755-nucleotide complete genome sequence of R. conorii, containing 1374 open reading frames. This genome exhibits 804 of the 834 genes of the previously determined R. prowazekii genome plus 552 supplementary open reading frames and a 10-fold increase in the number of repetitive elements. Despite these differences, the two genomes exhibit a nearly perfect colinearity that allowed the clear identification of different stages of gene alterations with gene remnants and 37 genes split in 105 fragments, of which 59 are transcribed. A 38-kilobase sequence inversion was dated shortly after the divergence of the genus.

Adaptation, Physiological↗

PubMed2004

Divergence and redundancy of 16S rRNA sequences in genomes with multiple rrn operons.

The level of sequence heterogeneity among rrn operons within genomes determines the accuracy of diversity estimation by 16S rRNA-based methods. Furthermore, the occurrence of widespread horizontal gene transfer (HGT) between distantly related rrn operons casts doubt on reconstructions of phylogenetic relationships. For this study, patterns of distribution of rrn copy numbers, interoperonic divergence, and redundancy of 16S rRNA sequences were evaluated. Bacterial genomes display up to 15 operons and operon numbers up to 7 are commonly found, but approximately 40% of the organisms analyzed have either one or two operons. Among the Archaea, a single operon appears to dominate and the highest number of operons is five. About 40% of sequences among 380 operons in 76 bacterial genomes with multiple operons were identical to at least one other 16S rRNA sequence in the same genome, and in 38% of the genomes all 16S rRNAs were invariant. For Archaea, the number of identical operons was only 25%, but only five genomes with 21 operons are currently available. These considerations suggest an upper bound of roughly threefold overestimation of bacterial diversity resulting from cloning and sequencing of 16S rRNA genes from the environment; however, the inclusion of genomes with a single rrn operon may lower this correction factor to approximately 2.5. Divergence among operons appears to be small overall for both Bacteria and Archaea, with the vast majority of 16S rRNA sequences showing <1% nucleotide differences. Only five genomes with operons with a higher level of nucleotide divergence were detected, and Thermoanaerobacter tengcongensis exhibited the highest level of divergence (11.6%) noted to date. Overall, four of the five extreme cases of operon differences occurred among thermophilic bacteria, suggesting a much higher incidence of HGT in these bacteria than in other groups.

Gene Dosage↗

PubMed2003

Cloning of an avian adeno-associated virus (AAAV) and generation of recombinant AAAV particles.

Recent studies have proposed that adeno-associated viruses (AAVs) are not evolutionarily linked to other mammalian autonomous parvoviruses but are more closely linked to the autonomous parvoviruses of birds. To better understand the relationship between primate and avian AAVs (AAAVs), we cloned and sequenced the genome of an AAAV (ATCC VR-865) and generated recombinant AAAV particles. The genome of AAAV is 4,694 nucleotides in length and has organization similar to that of other AAVs. The entire genome of AAAV displays 56 to 65% identity at the nucleotide level with the other known AAVs. The AAAV genome has inverted terminal repeats of 142 nucleotides, with the first 122 forming the characteristic T-shaped palindromic structure. The putative Rep-binding element consists of a tandem (GAGY)(4) repeat, and the putative terminal resolution site (trs), CCGGT/CG, contains a single nucleotide substitution relative to the AAV(2) trs. The Rep open reading frame of AAAV displays 50 to 54% identity at the amino acid level with the other AAVs, with most of the diversity clustered at the carboxyl and amino termini. Comparison of the capsid proteins of AAAV and the primate dependoviruses indicate that divergent regions are localized to surface-exposed loops. Despite these sequence differences, we were able to produce recombinant AAAV particles carrying a lacZ reporter gene by cotransfection in 293T cells and were able to examine transduction efficiency in both chicken primary cells and several cell lines. Our findings indicate that AAAV is the most divergent AAV described to date but maintains all the characteristics unique to the genera of dependovirus.

Amino Acid Sequence↗

PubMed2002

The putative roles of adenosine in insulin- and exercise-mediated regulation of glucose transport and glycogen metabolism in skeletal muscle.

Skeletal muscle is the primary site of whole-body glucose disposal and is vital in determining the overall insulin sensitivity and carbohydrate management. Insulin and physical exercise are important stimuli for muscle glucose transport and glycogen metabolism. While it is known that both insulin and contraction stimulate muscle glucose uptake and glycogen metabolism, the post-receptor mechanisms are not completely understood. Local metabolic factors, such as adenosine, have been suggested to play a role in insulin and contraction regulation of carbohydrate metabolism in skeletal muscle. While adenosine has clearly been shown to potentiate insulin-stimulated glucose transport in adipocytes and heart muscle, its role in carbohydrate metabolism in skeletal muscle is less clear, with numerous diverging findings published to date. This review article summarizes findings on the putative roles of adenosine in insulin and exercise-mediated regulation of carbohydrate metabolism and the signalling pathways proposed to be central to these metabolic stimuli in skeletal muscle.

Adenosine↗

PubMed2002

Early Tertiary out-of-India dispersal of Crypteroniaceae: evidence from phylogeny and molecular dating.

Phylogenetic analyses and molecular dating estimates based on chloroplast DNA sequences were used to establish the relationships of the southern and Southeast Asian Crypteroniaceae and elucidate their biogeographic history. Maximum parsimony and likelihood analyses of rbcL sequences suggested that Crypteroniaceae should be restricted to Crypteronia, Axinandra, and Dactylocladus and that Crypteroniaceae, so defined, are sister to a clade formed by three small African taxa (Oliniaceae, Penaeaceae, and Rhynchocalycaceae) and the monotypic Central and South American Alzateaceae. Three molecular dating approaches (maximum-likelihood under a molecular clock, Langley-Fitch, and penalized-likelihood) were used to infer the age of Crypteroniaceae using both paleobotanic and geologic calibrations. Comparisons among these three methods revealed significant lineage effects in rbcL sequences. Clock-independent dating estimates suggested that divergence of Crypteroniaceae from its African and South American relatives coincided with the breakup of Gondwana, and that India likely served as a "raft" transporting Crypteroniaceae to Asia, with later expansion to Southeast Asia. To our knowledge, Crypteroniaceae are the first plant group for which the out-of-India hypothesis is well corroborated by molecular-based estimates of divergence times.

Biological Evolution↗

PubMed2026

Chloroplast genome comparative analysis and phylogenetic relationships of 15 Syringa species (Oleaceae).

Syringa is a crucial shrub genus in the family Oleaceae, which has significant ornamental, economic, and medicinal value. However, research on the chloroplast genome (CPG) phylogeny and lineage diversification of this genus remains limited. In this study, all 15 Syringa CPGs exhibited a characteristic quadripartite structure, with genome lengths ranging from 154,019-158,020 bp. These CPGs were highly conserved and moderately differentiated, each containing 130-132 genes. Analysis of inverted repeat (IR) boundaries indicated structural conservation, with six genes: rps19, rpl2, ycf1, trnN, ndhF, and trnH present at the IR/single-copy (SC) junctions. The small single copy (SSC) region displayed greater sequence variability than the IR regions. ycf1, ndhH, trnL-rpl32, ndhF-ycf1, and rbcL-accD were identified as potential molecular markers and rps11, ycf2, and ycf4 may have contributed to the adaptive evolution of Syringa. Phylogenetic reconstruction based on whole CPG data supported the monophyly of the 15 species, which were divided into three distinct subclades. Molecular dating estimated that Syringa diverged from its sister genus approximately 58 million years ago, with most Syringa species diversifying further approximately 47.49 million years ago during the Eocene. Our findings will hopefully stimulate further studies on this genus that may enhance biodiversity knowledge.

Journal Article↗

PubMed2004

T29C polymorphism in the transforming growth factor beta1 gene and postmenopausal breast cancer risk: the Multiethnic Cohort Study.

Laboratory studies suggest a dual role for the transforming growth factor-beta (TGF-beta) signaling pathway in breast cancer. The normal antiproliferative activity of TGF-beta in early breast tumor development is replaced by a promoting effect in later stages. A T29C transition polymorphism in the TGFB1 gene has been associated with higher circulating TGF-beta1 levels, and inconsistently with breast cancer risk in three recent studies. We tested the association of this variant with invasive breast cancer in a case-control study of 1123 cases and 2314 controls nested in the Multiethnic Cohort (MEC) Study. This study is a large prospective study being conducted in Hawaii and Los Angeles that includes Japanese, white, African American, Latino, and Native Hawaiian women who were predominantly postmenopausal at baseline. After adjustment for breast cancer risk factors, the odds ratio (OR) and 95% confidence interval (95% CI) for the TGFB1 29 CC genotype was 0.95 (95% confidence interval: 0.76-1.18), compared to the TT genotype. Analyses stratified by race/ethnicity, stage, or age category did not reveal any association of this variant with breast cancer. Given the strong biological rationale and the scarce and divergent epidemiologic data to date, additional investigations of the relationship between breast cancer and genetic variants in the TGF-beta signaling pathway appear warranted.

Aged↗

PubMedDate not supplied

Molecular analysis of trkC in the cat visual cortex.

trkC belongs to the trk family of neurotrophin receptors. Several isoforms of trkC have been cloned to date; a full-length catalytic form containing a tyrosine kinase (TK) domain, three full-length isoforms with amino-acid insertions (14, 25, and 39 amino acids) in the TK domain, and five noncatalytic truncated forms that completely lack the TK domain. These isoforms have been studied in several mammalian species, including the pig, rat, mouse, monkey, and human. In this article we report the cloning and sequencing of five trkC isoforms isolated from 30-d postnatal cat visual cortex. The first isoform corresponded to the previously reported full-length trkC transcript containing the 14 amino-acid insert. To search for the presence of other inserts, reverse transcription polymerase chain reaction (RT-PCR) was performed on 30-d postnatal cat visual cortex mRNA using primers that flank the insertion site in the TK domain. Both the isoform containing the 14 amino-acid insert and the isoform lacking any insertion were present in abundant amounts, whereas the other two insert containing isoforms (TK25 and TK39) were much less abundant. The fifth isoform discovered corresponds to the previously reported truncated transcript. Overall, there is a high degree of identity (89-98%) and homology (97-99%) between the cat trkC nucleotide and amino-acid sequences among all mammals. The extracellular juxtamembrane domain was found to be highly divergent among all mammals that have been studied to date. This divergent region also included a proline deletion in the cat trkC sequence. This is the first report of the cloning, sequencing, and RT-PCR analysis of trkC in cat visual cortex, a system extensively studied using anatomical and physiological approaches.

Amino Acid Sequence↗

PubMed1997

Intermittent divergence of the protein tyrosine kinase family during animal evolution.

The protein tyrosine kinases (PTKs) are a large protein family consisting of many subfamilies with a variety of domain structures. The basic functions are thought to differ for different subfamilies. To know the dates at which the subfamilies diverged by gene duplications, a phylogenetic tree of the PTKs was inferred by comparing sequences from a wide range of species covering diploblasts and triploblasts. The PTK tree revealed that almost all of the gene duplications that gave rise to different subfamilies occurred rapidly before the diploblast-triploblast split, accompanying with rapid amino acid substitutions. This type of gene duplication was, however, rarely observed after that split. Long after the subfamily divergence, another type of gene duplication that gave rise to diverse tissue-specific genes occurred in each subfamily on the chordate lineage since the separation from arthropods. This type of gene duplication occurred frequently before the fish-tetrapod split, accompanying with rapid amino acid substitutions. In contrast, both the frequency of gene duplications and the rate of the amino acid substitutions were considerably reduced after that split. These results strongly suggest that the PTKs diverged intermittently, but not gradually, during animal evolution.

Amino Acids↗

PubMedDate not supplied

Entamoeba histolytica encodes a highly divergent beta-tubulin.

The microtubules of the amitochondrial parasite Entamoeba histolytica are atypical in certain respects. Consistent with this, we report that E. histolytica encodes the most divergent beta-tubulin identified to date, with only 54% to 58% identity to beta-tubulins from various species. A similarly divergent beta-tubulin is encoded by the related Entamoeba invadens; single gene copies appear to be present in both organisms. The Entamoeba sequences were compared with a database of 101 beta-tubulins, including the highly divergent sequence from another amitochondrial protozoan, Trichomonas vaginalis. A total of 81 residues were universally conserved, and 76 residues varied only once. Correlations with previous studies indicate that microtubule function is altered when most, but not all, conserved residues are mutated.

Amino Acid Sequence↗

PubMed2006

Integration of flowering dates in phenology and pollen counts in aerobiology: analysis of their spatial and temporal coherence in Germany (1992-1999).

We studied the possibility of integrating flowering dates in phenology and pollen counts in aerobiology in Germany. Data were analyzed for three pollen types (Betula, Poaceae, Artemisia) at 51 stations with pollen traps, and corresponding phenological flowering dates for 400 adjacent stations (< 25 km) for the years 1992-1993 and 1997-1999. The spatial and temporal coherence of these data sets was investigated by comparing start and peak of the pollen season with local minima and means of plant flowering. Our study revealed that start of birch pollen season occurred on average 5.7 days earlier than local birch flowering. For mugwort and grass, the pollen season started on average after local flowering was observed; mugwort pollen was found 4.8 days later and grass pollen season started almost on the same day (0.6 days later) as local flowering. Whereas the peak of the birch pollen season coincided with the mean flowering dates (0.4 days later), the pollen peaks of the other two species took place much later. On average, the peak of mugwort pollen occurred 15.4 days later than mean local flowering, the peak of grass pollen catches followed 22.6 days after local flowering. The study revealed a great temporal divergence between pollen and flowering dates with an irregular spatial pattern across Germany. Not all pollen catches could be explained by local vegetation flowering. Possible reasons include long-distance transport, pollen contributions of other than phenologically observed species and methodological constraints. The results suggest that further research is needed before using flowering dates in phenology to extrapolate pollen counts.

Artemisia↗

PubMed2006

Contrasting phylogeographical patterns between mainland and island taxa of the Pinus luchuensis complex.

Species whose geographical distribution encompasses both mainland and island populations provide an ideal system for examining isolation and genetic divergence. In this study, paternally transmitted chloroplast DNA (cpDNA) and maternally transmitted mitochondrial DNA (mtDNA) were used to estimate population structure and phylogeography of Pinus luchuensis, a species found in eastern China (ssp. hwangshanensis), Taiwan (ssp. taiwanensis), and the Ryukyu Archipelago (ssp. luchuensis). Gene genealogies of both mtDNA and cpDNA reveal two major lineages. Molecular dating indicates that these lineages diverged before the colonization of P. luchuensis subspecies in Taiwan and the Ryukyu Archipelago. Both mtDNA and cpDNA show a lack of correspondence between molecular phylogeny and subspecies designation. Phylogeographical analysis suggests that paraphyly of the subspecies is the result of recent divergence rather than secondary contacts. In spite of the short divergence history of P. luchuensis on islands, the island populations show the same degree of genetic divergence as mainland populations. Low levels of genetic diversity in the mainland ssp. hwangshanensis suggest demographic bottlenecks. In contrast, the high heterogeneity of genetic composition for island populations is likely to be associated with a history of multiple colonization from the mainland. The spatial apportionment of organelle DNA polymorphisms is consistent with a pattern of stepwise colonization on island populations.

China↗

PubMed2004

Estimating metazoan divergence times with a molecular clock.

Accurately dating when the first bilaterally symmetrical animals arose is crucial to our understanding of early animal evolution. The earliest unequivocally bilaterian fossils are approximately 555 million years old. In contrast, molecular-clock analyses calibrated by using the fossil record of vertebrates estimate that vertebrates split from dipterans (Drosophila) approximately 900 million years ago (Ma). Nonetheless, comparative genomic analyses suggest that a significant rate difference exists between vertebrates and dipterans, because the percentage difference between the genomes of mosquito and fly is greater than between fish and mouse, even though the vertebrate divergence is almost twice that of the dipteran. Here we show that the dipteran rate of molecular evolution is similar to other invertebrate taxa (echinoderms and bivalve molluscs) but not to vertebrates, which significantly decreased their rate of molecular evolution with respect to invertebrates. Using a data set consisting of the concatenation of seven different amino acid sequences from 23 ingroup taxa (giving a total of 11 different invertebrate calibration points scattered throughout the bilaterian tree and across the Phanerozoic), we estimate that the last common ancestor of bilaterians arose somewhere between 573 and 656 Ma, depending on the value assigned to the parameter scaling molecular substitution rate heterogeneity. These results are in accord with the known fossil record and support the view that the Cambrian explosion reflects, in part, the diversification of bilaterian phyla.

Animals↗

PubMed1998

The complete mitochondrial DNA sequence of the domestic sheep (Ovis aries) and comparison with the other major ovine haplotype.

The complete mitochondrial DNA (mtDNA) molecule of the domestic sheep, Ovis aries, was sequenced, together with part of the mtDNA of a specimen representing the other major O. aries haplotype group. The length of the complete ovine mtDNA presented is 16,616 nucleotides (nt). This length is not absolute, however, due to heteroplasmy caused by the occurrence of different numbers of a 75-nt-long tandem repeat in the control region. The sequence data were included in analyses of intraspecific ovine molecular differences, molecular comparisons with bovine mtDNAs, and phylogenetic analyses based on complete mtDNAs. The comparisons with bovine mtDNAs were based on the central domains of the ovine control regions, representing both major ovine haplotype groups, and the corresponding domains of Bos taurus and B. indicus. The comparisons showed that the difference between the bovids was 1.4 times greater than the intraspecific ovine difference. These findings suggest that the strains of wild sheep from which domestic sheep originated were more closely related than were the B. primigenius subspecies which gave rise to B. indicus and B. taurus cattle. Datings based on complete mtDNAs suggest that the bovine and ovine lineages diverged about 30 million years before present. This dating is considerably earlier than that proposed previously.

Animals↗

PubMed2002

Evolution of hematophagy in ticks: common origins for blood coagulation and platelet aggregation inhibitors from soft ticks of the genus Ornithodoros.

Identification and characterization of antihemostatic components from hematophagous organisms are useful for the elucidation of the evolutionary mechanisms involved in adaptation to a highly complex host hemostatic system. Although many bioactive components involved in the regulation of the host's hemostatic system have been described, the evolutionary mechanisms of how arthropods adapted to a blood-feeding environment have not been elucidated. This study describes common origins of both blood coagulation inhibitors and platelet aggregation inhibitors (PAIs) from soft ticks of the genus Ornithodoros. Neighbor-joining analysis indicates that fXa, thrombin, and PAIs share a common ancestor. Maximum parsimony analysis and a phylogeny based on root mean square deviation values of alpha-carbon backbone structures suggest a novel evolutionary pathway by which different antihemostatic functions have evolved through a series of paralogous gene duplication events. In this scenario, the thrombin inhibitors preceded the fXa and PAIs. This evolutionary model explains why the tick serine protease inhibitors have inhibition mechanisms that differ from that of the canonical bovine pancreatic trypsin inhibitor (BPTI)-like inhibitors. Higher nonsynonymous-to-synonymous substitution rates indicate positive Darwinian selection for the fXa and PAIs. Comparison with hemostatic inhibitors of hard ticks suggests that the two main tick families have independently evolved novel antihemostatic mechanisms. Independent evolution of these mechanisms in ticks points to a rapid divergence between tick families that could be dated between 120 and 92 MYA. This coincides with current molecular phylogeny views on the early divergence of modern birds and placental mammals in the Late Cretaceous, which suggests that this event might have been a driving force in the evolution of hematophagy in ticks.

Amino Acid Sequence↗

PubMed1996

Comparative HPLC, SDS-PAGE, and immunoblot analyses of dental enamel proteins.

The primary structures of amelogenins expressed from different genes vary because of DNA sequence divergence and variations in alternative RNA splicing. The pattern of splicing is unique for each amelogenin gene yet investigated, even when two copies of the gene are expressed in the same cell. Despite the high conservation of amelogenin sequences, diversity in the pattern of RNA splicing leads to significant differences in the number and character of amelogenin isoforms in the developing enamel matrix. Since conservation of molecular structure is an indicator of functional significance, we compared enamel protein preparations from rat, porcine, rabbit, and opossum developing tooth organs. Enamel extracts were fractionated by reversed-phase high-performance liquid chromatography (HPLC) and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Western blot analyses were performed with polyclonal antibodies raised against recombinant murine amelogenin and the polypeptide encoded by murine exon 4. The opossum enamel extract produced the simplest chromatogram, suggesting that fewer proteins are secreted into the developing enamel matrix. The predominant opossum amelogenin has an apparent molecular mass of 28 kDa and reacts strongly with the recombinant amelogenin antibody but is not recognized by the murine exon 4 antibody. Opossum amelogenin mRNA was amplified with murine amelogenin primers specific for the amino- and carboxyl-terminal coding regions. The mobility of the amplification products on 4% agarose gels indicates that the leucine-rich amelogenin polypeptide (LRAP) is expressed in the opossum and that the major amelogenin is larger than its homologue in the mouse. We conclude that the alternative splicing of amelogenins pre-dates the metatherian and eutherian divergence over 100 million years ago.

Alternative Splicing↗

PubMed2005

Initial diversification of living amphibians predated the breakup of Pangaea.

The origin and divergence of the three living orders of amphibians (Anura, Caudata, Gymnophiona) and their main lineages are one of the most hotly debated topics in vertebrate evolution. Here, we present a robust molecular phylogeny based on the nuclear RAG1 gene as well as results from a variety of alternative independent molecular clock calibrations. Our analyses suggest that the origin and early divergence of the three living amphibian orders dates back to the Palaeozoic or early Mesozoic, before the breakup of Pangaea, and soon after the divergence from lobe-finned fishes. The resulting new biogeographic scenario, age estimate, and the inferred rapid divergence of the three lissamphibian orders may account for the lack of fossils that represent plausible ancestors or immediate sister taxa of all three orders and the heretofore paradoxical distribution of some amphibian fossil taxa. Furthermore, the ancient and rapid radiation of the three lissamphibian orders likely explains why branch lengths connecting their early nodes are particularly short, thus rendering phylogenetic inference of implicated relationships especially difficult.

Amphibians↗

PubMed1997

The complete mitochondrial DNA sequence of the white rhinoceros, Ceratotherium simum, and comparison with the mtDNA sequence of the Indian rhinoceros, Rhinoceros unicornis.

The complete nucleotide sequence of the mitochondrial genome of the white rhinoceros, Ceratotherium simum, was determined. The length of the reported sequence is 16,832 nucleotides. This length can vary, however, due to pronounced heteroplasmy caused by differing numbers of a repetitive motif (5'-CG-CATATACA-3') in the control region. The 16,832 nucleotide sequence presented here is the longest version of the molecule and contains 35 copies of this motif. Comparison between the complete mitochondrial sequences of the white and the Indian (Rhinoceros unicornis) rhinoceroses allowed an estimate of the date of the basal evolutionary divergence among extant rhinoceroses. The calculation suggested that this divergence took place approximately 27 million years before present.

Animals↗