Electron transport between cytochrome c and alpha tocopherol.

Using liposomes we have demonstrated an electron transfer between tocopherol (vitamin E) and cytochrome c. Reduced cytochrome c protects vitamin E from oxidation induced either directly by ultraviolet light or indirectly by soybean lipoxygenase-catalyzed oxidation of arachidonic acid. Oxidized cytochrome c is reduced by tocopherol and tocopherol homologues (chromanols) resulting in accumulation of tocopheroxyl radicals which we detected by ESR. The peak height of the ESR spectrum of tocopheroxyl radicals (which is proportional to the amount of radical present) is proportional to the ratio of reduced to oxidized cytochrome c. In mitochondrial membranes succinate-cytochrome c reduction is inhibited by antimycin A. Addition of exogenous chromanols facilitates a by-pass of the antimycin A blocked electron pathway, and succinate-dependent cytochrome c reductase activity is restored. Cytochrome c may act as a water-soluble complement to the lipid-soluble ubiquinol in regenerating mitochondrial tocopherol from tocopheroxyl radical.     

Demonstration of binding of dengue virus envelope protein to target cells.

The nature of the initial interaction of dengue virus with target cells and the extent to which this interaction defines tropism are unknown. Infection of some cells may involve antidengue antibody-mediated immune adherence to cells bearing immunoglobulin Fc receptors; however, this mechanism does not explain primary infection or the infection of cells without Fc receptors. We hypothesized that dengue virus envelope protein mediates initial binding to target cells. To test this hypothesis, a recombinant chimeric form of dengue type 2 virus envelope protein was used as a probe to investigate binding to the surfaces of potential target cells. Envelope protein was expressed amino terminal to the heavy-chain constant region of human immunoglobulin G containing the Fc receptor binding motif; the binding mediated by envelope determinants was distinguishable from the binding mediated by immunoglobulin Fc determinants. We found that the recombinant chimera bound to Vero, CHO, endothelial, and glial cells through envelope protein determinants and to monocytes and U937 cells by Fc-Fc receptor interactions. The highest level of binding was to Vero cells; binding was dose and time dependent and saturable. Examination of partial-length recombinant envelope proteins indicated that the binding motif was expressed between amino acids 281 and 423. Recombinant envelope protein inhibited infection of Vero cells by dengue virus, indicating the functional significance of the interaction of envelope protein and target cells in infectivity. These results suggest that envelope protein binding to a non-Fc receptor could explain the cell and tissue tropism of primary dengue virus infection.     

The coupling of crossing over and homologous synapsis in maize inversions

Because fresh initiations of synapsis must occur for homologous synapsis of internal heterozygously inverted chromosome segments, attention has been directed at homologous synapsis and crossing over in overlapping paracentric inversions in the long arm of chromosome 1 of maize. In an earlier study with a relatively short inversion (where double crossovers within the inversion were rare), a recombination nodule (RN) was generally found at pachytene in reverse paired (homologously synapsed) inverted regions. Crossover frequency within the inversion, which could be independently estimated from analysis of bridge and fragment frequency at anaphase I and II, closely corresponded to crossover frequency estimated from observed RN frequency in pachytene inversion loops. These findings were consistent with the interpretation that establishment of homologous synapsis in this case is generally coupled to crossing over. This coupling suggests that there is very early commitment to the form of resolution of recombination intermediates that results in reciprocal recombination events instead of conversion only or other noncrossover events. This study examines another, larger paracentric inversion in the long arm of chromosome 1 that completely overlaps the first inversion. It is sufficiently longer than the first inversion that double crossover events are found within it with substantial frequency and interference considerations are feasible. This study confers additional insight into the interrelationships of synapsis and crossing over and the probable sequence in which the various involved processes usually occur. It raises the strong possibility that crossovers can be initiated during the alignment phase that precedes synapsis.     

Evidence for a correlation between ambient cholesterol levels and soluble plasma sialyltransferase enzyme activity

While soluble forms of the sialyltransferase (sialyl-T) enzyme have been detected in significant quantities in serum, the exact source(s) of the enzyme, or the factors controlling its secretion are poorly understood. In this study, we have examined the relationship between ambient plasma cholesterol concentrations and sialyl-T activities and also levels of constituent plasma sialoglycoproteins (SGP). There was an inverse relationship between levels of the 2,6 sialyl-T enzyme and both total plasma cholesterol and HDL, although no such relationship was observed for the 2,3 enzyme. While there was no correlation between total cholesterol and the levels of plasma SGPs, there was an inverse relationship between the HDL component and 2,3 SGPs.     

Synaptic defects of asynaptic homozygotes in maize at the electron microscope level.

More detailed observations of the synaptonemal complex (SC) in asynaptic maize plants have been faciliated by superior silver-staining procedures. These suggest that central region components of the SC are strongly implicated as defective in asynaptic. Apparently homologous axial elements tend to follow roughly parallel courses within the nucleus at pachytene, in some short segments apparently synapsed and in others at wider separation than normal synapsis yet close enough to allow observation of thin central element segments and also occasional thin transverse element-type structures. This kind of transverse filament may be weakened and severely stretched yet associated with both axial elements. Small nodules, similar to recombination nodules, appear at corresponding positions in widely separated axial elements. Key words : synaptonemal complex, central element, transverse filament, recombination nodule.     

Detecting non-neutral heterogeneity across a region of DNA sequence in the ratio of polymorphism to divergence

Natural selection, in the form of balancing selection or selective sweeps, can result in a decoupling of the amounts of molecular polymorphism and divergence. Thus natural selection can cause some areas of DNA sequence to have greater silent polymorphism, relative to divergence between species, than other areas. It would be useful to have a statistical test for heterogeneity in the polymorphism to divergence ratio across a region of DNA sequence, one that could identify heterogeneity greater than that expected from the neutral processes of mutation, drift, and recombination. The only currently available test requires that a region be arbitrarily divided into sections that are compared with each other, and the subjectivity of this division could be problematic. Here a test is proposed in which runs of polymorphic and fixed sites are counted, where a "run" is a set of one or more sites of one type preceded and followed by the other type. The number of runs is smaller than otherwise expected if polymorphisms are clumped together. By simulating neutral evolution and comparing the observed number of runs to the simulations, a statistical test is possible which does not require any a priori decisions about subdivision.      

Succinate-ubiquinone reductase linked recycling of alpha-tocopherol in reconstituted systems and mitochondria: requirement for reduced ubiquinone.

Studies have demonstrated that accumulation of mitochondrial tocopheroxyl radical, the primary oxidation product of alpha-tocopherol, accompanies rapid consumption of tocopherol. Enzyme-linked electron flow lowers both the steady-state concentration of the radical and the consumption of tocopherol. Reduction of tocopheroxyl radical by a mitochondrial electron carrier(s) seems a likely mechanism of tocopherol recycling. Succinate-ubiquinone reductase (complex II) was incorporated into liposomes in the presence of tocopherol and ubiquinone-10. After inducing formation of tocopheroxyl radical, it was possible to show that reduced ubiquinone prevents radical accumulation and tocopherol consumption. There was no evidence of direct reduction of tocopheroxyl radical by succinate-reduced complex II. These reactions were also measured using ubiquinone-1 and alpha-C-6-chromanol (2,5,7,8-tetramethyl-2-(4'-methylpentyl)-6-chromanol) which are less hydrophobic analogues of ubiquinone-10 and alpha-tocopherol. Mitochondrial membranes were made deficient in ubiquinone but sufficient in alpha-tocopherol and were reconstituted with added quinone. With these membranes it was shown that mitochondrial enzyme-linked reduction of ubiquinone protects alpha-tocopherol from consumption, and there is a requirement for ubiquinone. This complements the observations made in liposomes and we propose that reduced mitochondrial ubiquinones have a role in alpha-tocopherol protection, presumably through efficient reduction of the tocopheroxyl radical.