An initial ATP-independent step in the unwinding of a herpes simplex virus type I origin of replication by a complex of the viral origin-binding protein and single-strand DNA-binding protein

Using a spectrophotometric assay that measures the hyperchromicity that accompanies the unwinding of a DNA duplex, we have identified an ATP-independent step in the unwinding of a herpes simplex virus type 1 (HSV-1) origin of replication, Oris, by a complex of the HSV-1 origin binding protein (UL9 protein) and the HSV-1 single-strand DNA binding protein (ICP8). The sequence unwound is the 18-bp A + T-rich segment that links the two high-affinity UL9 protein binding sites, boxes I and II of Oris. P1 nuclease sensitivity of Oris and single-strand DNA-dependent ATPase measurements of the UL9 protein indicate that, at 37°C, the A + T-rich segment is sufficiently single stranded to permit the binding of ICP8. Binding of the UL9 protein to boxes I and II then results in the formation of the UL9 protein–ICP8 complex, that can, in the absence of ATP, promote unwinding of the A + T-rich segment. On addition of ATP, the helicase activity of the UL9 protein–ICP8 complex can unwind boxes I and II, permitting access of the replication machinery to the Oris sequences.

The thermodynamic origin of the stability of a thermophilic ribozyme

Understanding the mechanism of thermodynamic stability of an RNA structure has significant implications for the function and design of RNA. We investigated the equilibrium folding of a thermophilic ribozyme and its mesophilic homologue by using hydroxyl radical protection, small-angle x-ray scattering, and circular dichroism. Both RNAs require Mg2+ to fold to their native structures that are very similar. The stability is measured as a function of Mg2+ and urea concentrations at different temperatures. The enhanced stability of the thermophilic ribozyme primarily is derived from a tremendous increase in the amount of structure formed in the ultimate folding transition. This increase in structure formation and cooperativity arises because the penultimate and the ultimate folding transitions in the mesophilic ribozyme become linked into a single transition in the folding of the thermophilic ribozyme. Therefore, the starting point, or reference state, for the transition to the native, functional thermophilic ribozyme is significantly less structured. The shift in the reference state, and the resulting increase in folding cooperativity, is likely due to the stabilization of selected native interactions that only form in the ultimate transition. This mechanism of using a less structured intermediate and increased cooperativity to achieve higher functional stability for tertiary RNAs is fundamentally different from that commonly proposed to explain the increased stability of thermophilic proteins.

Origin of���Genes

We discuss two tests of the hypothesis that the first genes were assembled from exons. The hypothesis of exon shuffling in the progenote predicts that intron phases will be correlated so that exons will be an integer number of codons and predicts that the exons will be correlated with compact regions of polypeptide chain. These predictions have been tested on ancient conserved proteins (proteins without introns in prokaryotes but with introns in eukaryotes) and hold with high statistical significance. We conclude that introns are correlated with compact features of proteins 15-, 22-, or 30-amino acid residues long, as was predicted by “The Exon Theory of Genes.”

Phylogenetics and the origin of���species

A recent criticism that the biological species concept (BSC) unduly neglects phylogeny is examined under a novel modification of coalescent theory that considers multiple, sex-defined genealogical pathways through sexual organismal pedigrees. A competing phylogenetic species concept (PSC) also is evaluated from this vantage. Two analytical approaches are employed to capture the composite phylogenetic information contained within the braided assemblages of hereditary pathways of a pedigree: (i) consensus phylogenetic trees across allelic transmission routes and (ii) composite phenograms from quantitative values of organismal coancestry. Outcomes from both approaches demonstrate that the supposed sharp distinction between biological and phylogenetic species concepts is illusory. Historical descent and reproductive ties are related aspects of phylogeny and jointly illuminate biotic discontinuity.

Genetics and the origin of bird species

External (environmental) factors affecting the speciation of birds are better known than the internal (genetic) factors. The opposite is true for several groups of invertebrates, Drosophila being the outstanding example. Ideas about the genetics of speciation in general trace back to Dobzhansky who worked with Drosophila. These ideas are an insufficient guide for reconstructing speciation in birds for two main reasons. First, speciation in birds proceeds with the evolution of behavioral barriers to interbreeding; postmating isolation usually evolves much later, perhaps after gene exchange has all but ceased. As a consequence of the slow evolution of postmating isolating factors the scope for reinforcement of premating isolation is small, whereas the opportunity for introgressive hybridization to influence the evolution of diverging species is large. Second, premating isolation may arise from nongenetic, cultural causes; isolation may be affected partly by song, a trait that is culturally inherited through an imprinting-like process in many, but not all, groups of birds. Thus the genetic basis to the origin of bird species is to be sought in the inheritance of adult traits that are subject to natural and sexual selection. Some of the factors involved in premating isolation (plumage, morphology, and behavior) are under single-gene control, most are under polygenic control. The genetic basis of the origin of postmating isolating factors affecting the early development of embryos (viability) and reproductive physiology (sterility) is almost completely unknown. Bird speciation is facilitated by small population size, involves few genetic changes, and occurs relatively rapidly.

Regulatory evolution and the origin of the bilaterians

The adult body plan of bilaterians is achieved by imposing regional specifications on pluripotential cells. The establishment of spatial domains is governed in part by regulating expression of transcription factors. The key to understanding bilaterian evolution is contingent on our understanding of how the regulation of these transcription factors influenced bilaterian stem-group evolution.

The chimeric eukaryote: Origin of the nucleus from the karyomastigont in amitochondriate protists

We present a testable model for the origin of the nucleus, the membrane-bounded organelle that defines eukaryotes. A chimeric cell evolved via symbiogenesis by syntrophic merger between an archaebacterium and a eubacterium. The archaebacterium, a thermoacidophil resembling extant Thermoplasma, generated hydrogen sulfide to protect the eubacterium, a heterotrophic swimmer comparable to Spirochaeta or Hollandina that oxidized sulfide to sulfur. Selection pressure for speed swimming and oxygen avoidance led to an ancient analogue of the extant cosmopolitan bacterial consortium “Thiodendron latens.” By eubacterial-archaebacterial genetic integration, the chimera, an amitochondriate heterotroph, evolved. This “earliest branching protist” that formed by permanent DNA recombination generated the nucleus as a component of the karyomastigont, an intracellular complex that assured genetic continuity of the former symbionts. The karyomastigont organellar system, common in extant amitochondriate protists as well as in presumed mitochondriate ancestors, minimally consists of a single nucleus, a single kinetosome and their protein connector. As predecessor of standard mitosis, the karyomastigont preceded free (unattached) nuclei. The nucleus evolved in karyomastigont ancestors by detachment at least five times (archamoebae, calonymphids, chlorophyte green algae, ciliates, foraminifera). This specific model of syntrophic chimeric fusion can be proved by sequence comparison of functional domains of motility proteins isolated from candidate taxa.

On the origin of and phylogenetic relationships among living amphibians

The phylogenetic relationships among the three orders of modern amphibians (Caudata, Gymnophiona, and Anura) have been estimated based on both morphological and molecular evidence. Most morphological and paleontological studies of living and fossil amphibians support the hypothesis that salamanders and frogs are sister lineages (the Batrachia hypothesis) and that caecilians are more distantly related. Previous interpretations of molecular data based on nuclear and mitochondrial rRNA sequences suggested that salamanders and caecilians are sister groups to the exclusion of frogs. In an attempt to resolve this apparent conflict, the complete mitochondrial genomes of a salamander (Mertensiella luschani) and a caecilian (Typhlonectes natans) were determined (16,656 and 17,005 bp, respectively) and compared with previously published sequences from a frog (Xenopus laevis) and several other groups of vertebrates. Phylogenetic analyses of the mitochondrial data supported with high bootstrap values the monophyly of living amphibians with respect to other living groups of tetrapods, and a sister group relationship of salamanders and frogs. The lack of phylogenetically informative sites in the previous rRNA data sets (because of its shorter size and higher among-site rate variation) likely explains the discrepancy between our results and those based on previous molecular data. Strong support of the Batrachia hypothesis from both molecule- and morphology-based studies provides a robust phylogenetic framework that will be helpful to comparative studies among the three living orders of amphibians and will permit better understanding of the considerably divergent vertebral, brain, and digit developmental patterns found in frogs and salamanders.

The origin of 15R-prostaglandins in the Caribbean coral Plexaura homomalla: Molecular cloning and expression of a novel cyclooxygenase

The highest concentrations of prostaglandins in nature are found in the Caribbean gorgonian Plexaura homomalla. Depending on its geographical location, this coral contains prostaglandins with typical mammalian stereochemistry (15S-hydroxy) or the unusual 15R-prostaglandins. Their metabolic origin has remained the subject of mechanistic speculations for three decades. Here, we report the structure of a type of cyclooxygenase (COX) that catalyzes transformation of arachidonic acid into 15R-prostaglandins. Using a homology-based reverse transcriptase–PCR strategy, we cloned a cDNA corresponding to a COX protein from the R variety of P. homomalla. The deduced peptide sequence shows 80% identity with the 15S-specific coral COX from the Arctic soft coral Gersemia fruticosa and ≈50% identity to mammalian COX-1 and COX-2. The predicted tertiary structure shows high homology with mammalian COX isozymes having all of the characteristic structural units and the amino acid residues important in catalysis. Some structural differences are apparent around the peroxidase active site, in the membrane-binding domain, and in the pattern of glycosylation. When expressed in Sf9 cells, the P. homomalla enzyme forms a 15R-prostaglandin endoperoxide together with 11R-hydroxyeicosatetraenoic acid and 15R-hydroxyeicosatetraenoic acid as by-products. The endoperoxide gives rise to 15R-prostaglandins and 12R-hydroxyheptadecatrienoic acid, identified by comparison to authentic standards. Evaluation of the structural differences of this 15R-COX isozyme should provide new insights into the substrate binding and stereospecificity of the dioxygenation reaction of arachidonic acid in the cyclooxygenase active site.

Computer-modeling origin of a simple genetic apparatus

This computer simulation is based on a model of the origin of life proposed by H. Kuhn and J. Waser, where the evolution of short molecular strands is assumed to take place in a distinct spatiotemporal structured environment. In their model, the prebiotic situation is strongly simplified to grasp essential features of the evolution of the genetic apparatus without attempts to trace the historic path. With the tool of computer implementation confining to principle aspects and focused on critical features of the model, a deeper understanding of the model's premises is achieved. Each generation consists of three steps: (i) construction of devices (entities exposed to selection) presently available; (ii) selection; and (iii) multiplication of the isolated strands (R oligomers) by complementary copying with occasional variation by copying mismatch. In the beginning, the devices are single strands with random sequences; later, increasingly complex aggregates of strands form devices such as a hairpin-assembler device which develop in favorable cases. A monomers interlink by binding to the hairpin-assembler device, and a translation machinery, called the hairpin-assembler-enzyme device, emerges, which translates the sequence of R1 and R2 monomers in the assembler strand to the sequence of A1 and A2 monomers in the A oligomer, working as an enzyme.