Fas-induced apoptosis of T cells occurs independently of ceramide generation

The Fas receptor is one of a number of important physiological inducers of programmed cell death (apoptosis). Current models for regulation of this process involve rapid conversion of sphingomyelin to ceramide by cellular sphingomyelinases. Induced changes in cellular levels of such sphingosine-based ceramides are normally extrapolated from measurements of sphingomyelinase activity or following their conversion to ceramide phosphate by treatment of cellular lipid extracts with bacterial diacylglycerol kinase (DAGK). To allow direct study of cellular sphingosine- and sphinganine-based ceramide levels, we developed a mass spectrometric technique capable of determining inducible changes in both overall ceramide levels and species distribution in cellular lipid preparations. Contrary to current models, we detected no changes in cellular ceramide levels up to 2 hr poststimulation of Jurkat T cells with an anti-Fas IgM, although this treatment did induce apoptosis. We also determined in the same system that, when utilizing the DAGK assay, increased phosphorylation of substrates that comigrated with ceramide standards was apparent but that this effect was due to an enhancement of DAGK activity rather than increases in levels of cellular ceramides as substrates per se. Thus, the first direct measurement of ceramides present in cells undergoing apoptosis indicates that, insofar as it can be measured, the induction of apoptosis does not involve the generation of sphingosine-based ceramides, contrary to many published accounts.

Homospermidine synthase, the first pathway-specific enzyme of pyrrolizidine alkaloid biosynthesis, evolved from deoxyhypusine synthase

Pyrrolizidine alkaloids are preformed plant defense compounds with sporadic phylogenetic distribution. They are thought to have evolved in response to the selective pressure of herbivory. The first pathway-specific intermediate of these alkaloids is the rare polyamine homospermidine, which is synthesized by homospermidine synthase (HSS). The HSS gene from Senecio vernalis was cloned and shown to be derived from the deoxyhypusine synthase (DHS) gene, which is highly conserved among all eukaryotes and archaebacteria. DHS catalyzes the first step in the activation of translation initiation factor 5A (eIF5A), which is essential for eukaryotic cell proliferation and which acts as a cofactor of the HIV-1 Rev regulatory protein. Sequence comparison provides direct evidence for the evolutionary recruitment of an essential gene of primary metabolism (DHS) for the origin of the committing step (HSS) in the biosynthesis of pyrrolizidine alkaloids.

Sexually antagonistic coevolution of a postmating-prezygotic reproductive character in desert Drosophila

Rapid divergence in postmating-prezygotic characters suggests that selection may be responsible for generating reproductive barriers between closely related species. Theoretical models indicate that this rapid divergence could be generated by a series of male adaptations and female counteradaptations by means of sexual selection or conflict, but empirical tests of particular mechanisms are generally lacking. Moreover, although a male–female genotypic interaction in mediating sperm competition attests to an active role of females, molecular or morphological evidence of the female's participation in the coevolutionary process is critically needed. Here we show that postmating-prezygotic variation among populations of cactophilic desert Drosophila reflects divergent coevolutionary trajectories between the sexes. We explicitly test the female's role in intersexual interactions by quantifying differences in a specific postmating-prezygotic reproductive character, the insemination reaction mass, in two species, Drosophila mojavensis and Drosophila arizonae. A series of interpopulation crosses confirmed that population divergence was propelled by male–female interactions, a prerequisite if the selective forces derive from sexual conflicts. An association between the reaction mass and remating and oviposition behavior argues that divergence has been propelled by sexually antagonistic coevolution, and potentially has important implications for speciation.