The kinetics and energetics of formation of a molecular hydrogen complex involving a dinuclear ruthenium(II) centre

The reversible equilibrium conversion under H₂ of [RuCl(dppb) (μ-Cl)]₂ (1) to generate (η²-H₂) (dppb) (μ-Cl)₃RuCl(dppb) in CH₂Cl₂ (dppb = Ph₂P(CH₂)₄PPh₂) has been studied at 0–25 °C by UV-Vis and ³¹P{¹H} NMR spectroscopy, and by stoppe kinetics; the equilibrium constant and corresponding thermodynamic parameters, and the forward and reverse rate constants at 25 °C have been determined. A measured ΔH° value of 0 kJ mol⁻¹ allows for an estimation of an exothermicity of 60 kJ mol⁻¹ for binding an η²-H₂ at an Ru(II) centre; a ΔS° value of 60 J mol⁻¹ K⁻¹ indicates that in solution 1 contain s coordinated CH₂Cl₂. The kinetic and thermodynamic data are compared to those obtained from a previously studied hydrogenation of styrene catalyzed by 1. Preliminary findings on related systems containing Ph₂P(CH₂)₃PPh₂ and (C₆H₁₁)₂P(C₆H₁₁)₂ are also noted.     

Phenotypic features of breast cancer cells overexpressing ornithine-decarboxylase

Polyamines (PA) have been shown to be critical mediators of estradiol-induced breast cancer cell proliferation. This finding suggests that constitutive activation of the PA pathway may promote tumor progression, possibly leading to hormone independence. To test this hypothesis, we transfected hormone-responsive MCF-7 breast cancer cells with a complementary DNA coding for ornithine-decarboxylase (ODC), the first rate-limiting enzyme in PA biosynthesis. Marked ODC over-expression observed in stably transfected clones was associated with a selective increase in cellular putrescine content, while spermidine and spermine levels were not altered. ODC-overexpressing MCF-7 cells were resistant to the antiproliferative effects of low but not high concentrations of the enzyme inhibitor, α-difluoromethylornithine. In agreement with our hypothesis, sensitivity to the growth-promoting action of estradiol was reduced by approximately one third (P < 0.001) in ODC-overexpressing MCF-7 cells compared with vector-only transfected clones. Basal growth under anchorage-dependent conditions was only marginally increased by ODC overexpression (P = 0.048), while clonogenicity in soft agar was actually reduced. These data suggest that activation of PA biosynthesis may contribute in part to the acquisition of estrogen independence by breast cancer cells. Since only putrescine content was increased as a result of ODC overexpression, these data may underestimate the overall influence of the PA pathway on breast cancer phenotype. © 1995 Wiley-Liss, Inc.     

Lipopolysaccharide inhibits activation of latent transforming growth factor-β in bovine endothelial cells

The activation of latent transforming growth factor-β (TGF-β) by vascular endothelial cells (ECs) is regulated by cellular plasminogen activator (PA)/plasmin, transglutaminase (TGase), and latent TGF-β levels. Because lipopolysaccharide (LPS) has been reported to reduce EC surface plasmin levels by increasing the production of the inhibitor of PA, PA inhibitor-1 (PAI-1), we have tested whether LPS might suppress latent TGF-β activation in ECs using two different systems, namely, bovine aortic ECs (BAECs) cocultured with smooth muscle cells (SMCs) and BAECs treated with retinol. BAECs were either cocultured with SMCs after treatment with 15 ng/ml LPS or were treated with 2 μM retinol and/or 10 ng/ml LPS, and the expression of PA, surface plasmin, TGase, and the amounts of active and latent TGF-β secreted into the culture modium were measured. The downregulation of surface PA/plasmin levels with LPS was accompanied by a profound decline of both TGase and latent TGF-β expression as well as the suppression of surface activation of latent TGF-β. The effect was dependent on the concentration of LPS and on treatment time. The formation of TGF-β did not occur in cells maintained in LPS-contaminated culture medium. © 1995 Wiley-Liss, Inc.     

Cyclic-AMP inhibits cell growth and negatively interacts with platelet membrane glycoprotein expression on the Dami human megakaryoblastic cell line

Intracellular signaling processes by which hematopoietic growth factors regulate megakaryocytopoiesis remain uncompletely understood. Cyclic AMP (cAMP) has been shown to be implicated in the regulation of growth and differentiation in various normal and malignant cell types. Since a few studies have suggested the possible involvement of the cAMP pathway as one of the intracellular mechanisms whereby megakaryocytopoiesis may be regulated, we investigated the functional effects of cAMP on the human megakaryoblastic Dami cell line. We observed that exposure of Dami cells to cAMP analogs or to agents elevating intracellular cAMP levels yielded dose-dependent cell growth inhibition. Cell cycle progression analysis of cells predominantly synchronized at the G1/S boundary by prior treatment with hydroxyurea revealed that cAMP transiently accumulated cells in the G2/M phase, then slowing down cell cycle. On the other hand, immunofluorescence and Northern blot analysis of megakaryocytic differentiation marker expression showed that probes we have used significantly inhibited GPlb expression. Moreover, although these agents used alone did not affect GPllb/llla expression, they markedly reversed phorbol ester-induced GPllb/llla expression increase. These inhibitory cAMP actions on glycoprotein expression were not the result of cell cycle perturbation since we observed that GPlb and GPllb/llla expression were not cell cycle dependent. All these data may then be consistent with a potential negative regulatory role of the cAMP intracellular signaling pathway during megakaryocytopoiesis. © 1995 Wiley-Liss, Inc.     

Molecular cloning and complete nucleotide sequence of the repeated unit and flanking gene of the scallop Pecten maximus mitochondrial DNA: Putative replication origin features

In the bivalve mollusc Pecten maximus, the size of the mitochondrial DNA molecules ranges from 20 to 25.8 kbp. This variability is mainly correlated with the occurrence of a variable domain composed with two to five 1.6-kbp repeated units tandemly arrayed in the genome. DNA fragments spanning the 1,586-base-pair-long repeated element and the nearest flanking gene have been cloned and sequenced. This sequence was analyzed regarding its base composition and potential secondary structures. The repeated unit domain was positioned and oriented with regard to the known flanking gene. It ends 2 base pairs upstream relative to the beginning of the tRNA^gly gene. The peculiar properties of the repeated unit were compared with those of the 1,442-bp repeated element found in the mitochondrial genome of the deep sea scallop Placopecten magellanicus. This comparison provided evidence for the absence of nucleotide conservation, except for a small sequence engaged in a secondary structure, but argued for a strong pressure maintaining domains with specific nucleotide content. A possible role for the conserved sequence is discussed.Correspondence to: A. Rigaa     

Biodiversity of sweetpotato (iPomoea batatas,Convolvulaceae) in Southern Mexico

Between 1987 and 1992 the phytogeographic region of southern Mexico was explored during three collecting trips made in search of cultivated and wild germplasm of the sweetpotato (Ipomoea batatas). The first trip was made in 1987, when we collected wild species in Ipomoea section Batatas found in the southeastern and southwestern regions of Mexico. A second trip was made in 1990, when we collected accessions of the cultivated species as well as wild species in the southeast, southwest and northeast. The third and final trip was oriented at identification, characterization and collecting seeds in the ecological niches ofI.tabascana andI.umbraticola. As a result of the three trips we collected 165 accessions of cultivated and wild germplasm with populations dispersed in 147 localities in 15 states of the Mexican region: Veracruz, Tabasco, Campeche, Chiapas, Oaxaca, Yucatán, Guerrero, Michoacán, San Luis Potosí, Hidalgo, Querétaro, Tamaulipas, Guanajuato, Puebla and México. Of the total accessions some 64 (38.3%) were of the cultivated species including nine accessions of feral material, and 103 accessions (61.7%) were of wild species made up of 59 accessions of seven species in the section Batatas, 37 of other species in the family Convolvulaceae, and seven yet to be determined. We have identified the largest genetic biodiversity in six localities of five states: Tabasco, Oaxaca, Michoacán, San Luis Potosí, and Puebla. Biodiversity maintenance in this region is associated with the day-of-the-dead festivities.     

GENETIC CONSTRAINTS ON MACROEVOLUTION: THE EVOLUTION OF HOST AFFILIATION IN THE LEAF BEETLE GENUS OPHRAELLA

We hypothesize that the evolution of an ecologically important character, the host associations of specialized phytophagous insects, has been influenced by limitations on genetic variation. Using as a historical framework a phylogenetic reconstruction of the history of host associations in the beetle genus Ophraella (Chrysomelidae), we have employed quantitative-genetic methods to screen four species for genetic variation in larval survival, oviposition (in one species only), and feeding responses to their congeners' host plants, in the Asteraceae. We here report results of studies of one species and evaluate the results from all four. Analysis of half-sib/full-sib families and of progenies of wild females of O. notulata, a specialist on Iva (Ambrosiinae), provided evidence of genetic variation in larval consumption of five of six test plants and in adult consumption of four of six. Larval mortality was complete on five plants; only on Ambrosia, a close relative of the natural host, was there appreciable, and genetically variable, survival. Oviposition on Ambrosia showed marginally significant evidence of genetic variation; a more distantly related plant elicited no oviposition at all. In compiling results from four Ophraella species, reported in this and two other papers, we found no evidence of genetic variation in 18 of 39 tests of feeding responses and 14 of 16 tests of larval survival on congeners' hosts. This result is consistent with the hypothesis that absence or paucity of genetic variation may constrain or at least bias the evolution of host associations. The lower incidence of genetic variation in survival than in feeding behavior may imply, according to recent models, that avoidance is a more common evolutionary response to novel plants than adaptation. The usually great disparity between mean performance on congeners' hosts and the species' natural hosts, and an almost complete lack of evidence for negative genetic correlations, argue against the likelihood that speciation has occurred by sympatric host shift. The presence versus apparent absence of genetic variation in consumption was correlated with the propinquity of relationship between the beetle species tested and the species that normally feeds on the test plant, suggesting that the history of host shifts in Ophraella has been guided in part by restrictions on genetic variation. It was also correlated with the propinquity of relationship between a test plant and the beetle's natural host. The contributions of plant relationships and insect relationships, themselves correlated in part, to the pattern of genetic variation, are not readily distinguishable, but together accord with phylogenetic evidence that these and other phytophagous insects adapt most readily to related plants. In this instance, therefore, the macroevolution of an ecologically important character appears to have been influenced by genetic constraints. We hypothesize that absence of the structural prerequisites for genetic variation in complex characters may affect genetic variation and the trajectory of evolution.     

P1 clones from Drosophila melanogaster as markers to study the chromosomal evolution of Muller's A element in two species of the obscura group of Drosophila

Thirty P1 clones from the X chromosome (Muller's A element) of Drosophila melanogaster were cross-hybridized in situ to Drosophila subobscura and Drosophila pseudoobscura polytene chromosomes. An additional recombinant phage Dsuby was also used as a marker. Twenty-three (77%) of the P1 clones gave positive hybridization on D. pseudoobscura chromosomes bat only 16 (53%) did so with those of D. subobscura. Eight P1 clones gave more than one hybridization signal on D. pseudoobscura and/or D. subobscura chromosomes. All P1 clones and Dsuby hybridized on Muller's A element (X chromosome) of D. subobscura. In contrast, only 18 P1 clones and Dsuby hybridized on Muller's D element (XR chromosomal arm) of D. pseudoobscura; 4 additional P1 clones hybridized on Muller's D element (XR chromosomal arm) of this species and the remaining P1 clone gave on hybridization signal on each arm of the X chromosome. This latter clone may contain one breakpoint of a pericentric inversion that may account for the interchange of genetic material between Muller's A and D elements in D. pseudoobscura. In contrast to the rare interchange of genetic material between chromosomal elements, profound differences in the order and spacing of markers were detected between D. melanogaster, D. pseudoobscura and D. subobscura. In fact, the number of chromosomal segments delimited by identical markers and conserved between pairwise comparisons is small. Therefore, extensive reorganization within Muller's A element has been produced during the divergence of the three species. Rough estimates of the number of cytologically detectable inversions contributing to differentiation of Muller's A element were obtained. The most reliable of these estimates is that obtained from the D. pseudoobscura and D. melanogaster comparison since a greater number of markers have been mapped in both species. Tentatively, one inversion breakpoint about every 200 kb has been produced and fixed during the divergence of D. pseudoobscura and D. melanogaster.     

Ribosomal proteins ofThermomyces lanuginosus — characterisation by two-dimensional gel electrophoresis and differential disassembly

One- and two-dimensional gel electrophoresis were employed to characterise the proteins derived from the ribosomes of the thermophilic fungusThermomyces lanuginosus. Approximately 32 (29 basic and 3 acidic) and 45 (43 basic and 2 acidic) protein spots were resolved fromTh. lanuginosus small and large ribosomal subunits, respectively. The molecular weight of the small subunit proteins ranged from 9,800–36,000 Da with a number average molecular weight of 20,300 Da. The molecular weight range for the large subunit proteins was 12,000–48,500 Da with a number average molecular weight of 25,900 Da. Most proteins appeared to be present in unimolar amounts. These data are comparable with but not identical to those from other eukaryotic ribosomes. The sensitivities of the ribosomal proteins to increasing concentrations of NH₄Cl were also evaluated by two-dimensional gel electrophoresis. Most ribosomal proteins were gradually released over a wide range of salt concentrations but some were preferentially enriched in one or two salt conditions.