A phylogenetic approach to the identification of phosphoglucomutase genes

The expanding molecular database provides unparalleled opportunities for characterizing genes and for studying groups of related genes. We use sequences drawn from the database to construct an evolutionary framework for examining the important glycolytic enzyme phosphoglucomutase (PGM). Phosphoglucomutase plays a pivotal role in the synthesis and utilization of glycogen and is present in all organisms. In humans, there are three well-described isozymes, PGMI, PGM2, and PGM3. PGM1 was cloned 5 years ago; however, repeated attempts using both immunological approaches and molecular probes designed from PGM1 have failed to isolate either PGM2 or PGM3. Using a phylogenetic strategy, we first identified 47 highly divergent prokaryotic and eukaryotic PGM-like sequences from the database. Although overall amino acid identity often fell below 20%, the relative order, position, and sequence of three structural motifs, the active site and the magnesium-- and sugar-binding sites, were conserved in all 47 sequences. The phylogenetic history of these sequences was complex and marked by duplications and translocations; two instances of transkingdom horizontal gene transfer were identified. Nonetheless, the sequences fell within six well-defined evolutionary lineages, three of which contained only prokaryotes. Of the two prokaryotic/eukaryotic lineages, one contained bacterial, yeast, slimemold, invertebrate, and vertebrate homologs to human PGM1 and the second contained likely homologs to human PGM2. Indeed, an amino acid sequence, derived from a partial human cDNA, that fell within the second cross-kingdom lineage bears several characteristics expected for PGM2. A third lineage may contain homologs to human PGM3. On a general level, our phylogenetic-based approach shows promise for the further utilization of the extensive molecular database.      

Initiation of DNA Synthesis in HeLa Cell-free System

THE molecular mechanism for initiating DNA replication can be studied using a subcellular system. Rao and Johnson¹ found that HeLa cells in the pre-DNA-synthetic (G-1) period of the cell cycle initiate DNA synthesis after fusion with cells that are in the DNA synthetic (S) period. A previous subcellular system of DNA replication from HeLa cells^2–4 consisted of intact nuclei, supplemented with the four deoxy-nucleoside triphosphates, salt, ATP and a cytosol factor. The nuclei in this system appeared to be permeable to proteins and DNA synthesis was very similar to that within intact cells. We report here the initiation of DNA synthesis in nuclei isolated from HeLa cells. Our results suggest that, with the synchronization method used, a small percentage of dormant G-1 nuclei can be stimulated by S-phase cytoplasm; this would be the case if the cells were receptive to stimulation for only 30–60 min during the cell cycle. illustration

Open image in new window

     

LATE DEVONIAN (FAMENNIAN) LUNGFISHES FROM THE CATSKILL FORMATION OF PENNSYLVANIA, USA

Abstract:  Occurrences of fossil lungfishes (Dipnoi: Sarcopterygii) in the Famennian Catskill Formation of Pennsylvania are reviewed. A nearly complete dermal skull roof is assigned to a new genus and species, Apatorhynchus opistheretmus . Other recently discovered lungfish specimens include an incomplete postcranium similar to that of the Frasnian genus Fleurantia , a small parasphenoid of uncertain affinities, and isolated toothplates. Previously described dipnoan remains from the Catskill Formation include a partial skull roof of Soederberghia groenlandica , toothplates assigned to several species of Dipterus , a putative rostral or symphysial region placed in the problematic form taxon Ganorhynchus , and sedimentary structures interpreted as burrows. The toothplates attributed to Dipterus are indeterminate and are placed in open nomenclature, while the specimen identified as Ganorhynchus is not convincingly dipnoan. The status of the burrows remains uncertain pending the discovery of lungfish remains within these or similar structures in Catskill deposits. The distinct ichthyofaunas within the Catskill Formation and their lungfish components are briefly reviewed. Lungfishes are found in the Holoptychius - and Bothriolepis -dominated faunas common in the Catskill succession, as well as in the compositionally distinctive Red Hill assemblage. Many of the Devonian continental faunas that contain tetrapods also include long-snouted, denticle-bearing lungfishes ('rhynchodipterids', fleurantiids, or both). The composition of Late Devonian ichthyofaunas may have predictive qualities that will allow researchers to identify localities likely to produce the remains of early tetrapods.     

Statistical properties of the variation at linked microsatellite loci: implications for the history of human Y chromosomes [published erratum appears in Mol Biol Evol 1997 Mar;14(3):354]

It has recently been suggested that observed levels of variation at microsatellite loci can be used to infer patterns of selection in genomes and to assess demographic history. In order to evaluate the feasibility of these suggestions it is necessary to know something about how levels of variation at microsatellite loci are expected to fluctuate due simply to stochasticity in the processes of mutation and inheritance (genetic sampling). Here we use recently derived properties of the stepwise mutation model to place confidence intervals around the variance in repeat score that is expected at mutation-drift equilibrium and outline a statistical test for whether an observed value differs significantly from expectation. We also develop confidence intervals for the time course of the buildup of variation following a complete elimination of variation, such as might be caused by a selective sweep or an extreme population bottleneck. We apply these methods to the variation observed at human Y-specific microsatellites. Although a number of authors have suggested the possibility of a very recent sweep, our analyses suggest that a sweep or extreme bottleneck is unlikely to have occurred anytime during the last approximately 74,000 years. To generate this result we use a recently estimated mutation rate for microsatellite loci of 5.6 x 10(-4) along with the variation observed at autosomal microsatellite loci to estimate the human effective population size. This estimate is 18,000, implying an effective number of 4,500 Y chromosomes. One important general conclusion to emerge from this study is that in order to reject mutation-drift equilibrium at a set of linked microsatellite loci it is necessary to have an unreasonably large number of loci unless the observed variance is far below that expected at mutation-drift equilibrium.      

Ultrastructural Damage to the Malaria Parasite in the Sickled Cell

The process by which malaria parasites are killed in sickled erythrocytes was studied by electron microscopy. In vitro cultures of Plasmodium falciparum in sickle cell hemoglobin (HbS) homozygous (SS) and heterozygous (SA) red cells were deoxygenated for up to 6 h and fixed under anaerobic conditions. Parasites in SS cells appeared to be disrupted by intrusions of needle-like deoxyHbS aggregates; disintegration of cytoplasm and membranes followed. In SA red cells, the parasites were generally not disrupted. Instead, extensive vacuolization occurred, a sign of metabolic inhibition. The resistance of HbS gene carriers to malaria results partly from these causes of intracellular parasite death.     

A 28s ribosomal RNA phylogeny of certain cyclorrhaphous Diptera based upon a hypervariable region

Abstract The method of primer extension dideoxy ribosomal RNA sequencing coupled with maximum parsimony analysis was used to obtain partial 28s rRNA sequences of a variety of calyptrate and acalyptrate Diptera with which to establish a possible genealogy of these organisms. Analysis of the data shows the calyptrates to be more closely related to the drosophiloids than to the tephritoids. Thus, the acalyptrates are a paraphyletic group. The analyses also establish the oestroid character of the gasterophilids and the status of the fanniids as a family or higher taxon.