Calcium uptake by isolated nerve endings: evidence for a rapid component mediated by the breakdown of phosphatidylinositol

Several physiological stimuli, including neuronal depolarization, increase the production of phosphatidate (PA) from phosphatidylinositol (PI) and increase calcium fluxes across cell membranes. To determine if breakdown of PI is required for neuronal calcium uptake, we tested inhibitors of PI-specific phospholipase C on depolarization-dependent uptake of calcium by isolated brain synaptosomes. At a concentration of 0.1 mM these inhibitors reduced calcium uptake produced by depolarization for 1 to 3 sec, but did not affect uptake due to more prolonged depolarization. Exogenous PA also stimulated calcium accumulation by synaptosomes and this uptake was not reduced by the enzyme inhibitors. These results suggest that the rapid calcium influx produced by neuronal depolarization may be mediated by the breakdown of PI.     

A major difference between the divergence patterns within the lines-1 families in mice and voles

L1 retroposons are represented in mice by subfamilies of interspersed sequences of varied abundance. Previous analyses have indicated that subfamilies are generated by duplicative transposition of a small number of members of the L1 family, the progeny of which then become a major component of the murine L1 population, and are not due to any active processes generating homology within preexisting groups of elements in a particular species. In mice, more than a third of the L1 elements belong to a clade that became active approximately 5 Mya and whose elements are > or = 95% identical. We have collected sequence information from 13 L1 elements isolated from two species of voles (Rodentia: Microtinae: Microtus and Arvicola) and have found that divergence within the vole L1 population is quite different from that in mice, in that there is no abundant subfamily of homologous elements. Individual L1 elements from voles are very divergent from one another and belong to a clade that began a period of elevated duplicative transposition approximately 13 Mya. Sequence analyses of portions of these divergent L1 elements (approximately 250 bp each) gave no evidence for concerted evolution having acted on the vole L1 elements since the split of the two vole lineages approximately 3.5 Mya; that is, the observed interspecific divergence (6.7%-24.7%) is not larger than the intraspecific divergence (7.9%-27.2%), and phylogenetic analyses showed no clustering into Arvicola and Microtus clades.      

(Photo)chemistry of 5-deazaflavin. A clue to the mechanism of flavin-dependent (de)hydrogenation.

The catalytic action of 5-deazaflavin in the photochemical reduction of flavin and iron proteins [Massey, V. and Hemmerich, P. (1978) Biochemistry, 17, 9--17] is shown to be due to the highly reactive 5-deazaflavosemiquinone. This radical is generated in a complex sequence of reactions, which involves (a) covalent photoaddition of the substrate residue to the deazaflavin, (b) fast secondary photoreaction of this adduct with starting deazaflavin to yield a covalent radical dimer, accompanied by the liberation of the oxidized substrate, and (c) deazaflavin-sensitized cleavage of the radical dimer to the monomers. The structure and properties of this radical (redimerisation or dismutation) and the precursor intermediates as well as the mechanism of the photoreaction are described. Deazaflavins and their natural parent compounds are compared with respect to their different redox behavior and radical stability. The syntheses of 5-deuterated deazaflavins are described and their redox reactions are compared with those of normal deazaflavins.     

Multifactorial diversity sustains microbial community stability

Maintenance of a high degree of biodiversity in homogeneous environments is poorly understood. A complex cheese starter culture with a long history of use was characterized as a model system to study simple microbial communities. Eight distinct genetic lineages were identified, encompassing two species: Lactococcus lactis and Leuconostoc mesenteroides. The genetic lineages were found to be collections of strains with variable plasmid content and phage sensitivities. Kill-the-winner hypothesis explaining the suppression of the fittest strains by density-dependent phage predation was operational at the strain level. This prevents the eradication of entire genetic lineages from the community during propagation regimes (back-slopping), stabilizing the genetic heterogeneity in the starter culture against environmental uncertainty.     

B Cell Deficient Mice Are Protected from Biliary Obstruction in the Rotavirus-Induced Mouse Model of Biliary Atresia

A leading theory regarding the pathogenesis of biliary atresia (BA) is that bile duct injury is initiated by a virus infection, followed by an autoimmune response targeting bile ducts. In experimental models of autoimmune diseases, B cells have been shown to play an important role. The aim of this study was to determine the role of B cells in the development of biliary obstruction in the Rhesus rotavirus (RRV)-induced mouse model of BA. Wild-type (WT) and B cell-deficient (Ig-α^-/-) mice received RRV shortly after birth. Ig-α^-/- RRV-infected mice had significantly increased disease-free survival rate compared to WT RRV-infected BA mice (76.8% vs. 17.5%). In stark contrast to the RRV-infected BA mice, the RRV-infected Ig-α^-/- mice did not have hyperbilirubinemia or bile duct obstruction. The RRV-infected Ig-α^-/- mice had significantly less liver inflammation and Th1 cytokine production compared to RRV-infected WT mice. In addition, Ig-α^-/- mice had significantly increased numbers of regulatory T cells (Tregs) at baseline and after RRV infection compared to WT mice. However, depletion of Tregs in Ig-α^-/- mice did not induce biliary obstruction, indicating that the expanded Tregs in the Ig-α^-/- mice were not the sole reason for protection from disease. Conclusion: B cell deficient Ig-α^-/- mice are protected from biliary obstruction in the RRV-induced mouse model of BA, indicating a primary role of B cells in mediating disease pathology. The mechanism of protection may involve lack of B cell antigen presentation, which impairs T-cell activation and Th1 inflammation. Immune modulators that inhibit B cell function may be a new strategy for treatment of BA.