Formation of Virus Assembly Intermediate Complexes in the Cytoplasm by Wild-Type and Assembly-Defective Mutant Human Immunodeficiency Virus Type 1 and Their Association with Membranes

We have previously identified two distinct forms of putative viral assembly intermediate complexes, a detergent-resistant complex (DRC) and a detergent-sensitive complex (DSC), in human immunodeficiency virus type 1 (HIV-1)-infected CD4(+) T cells (Y. M. Lee and X. F. Yu, Virology 243:78-93, 1998). In the present study, the intracellular localization of these two viral assembly intermediate complexes was investigated by use of a newly developed method of subcellular fractionation. In wild-type HIV-1-infected H9 cells, the DRC fractionated with the soluble cytoplasmic fraction, whereas the DSC was associated with the membrane fraction. The DRC was also detected in the cytoplasmic fraction in H9 cells expressing HIV-1 Myr- mutant Gag. However, little of the unmyristylated Gag and Gag-Pol proteins was found in the membrane fraction. Furthermore, HIV-1 Gag proteins synthesized in vitro in a rabbit reticulocyte lysate system in the absence of exogenous lipid membrane were able to assemble into a viral Gag complex similar to that of the DRC identified in infected H9 cells. The density of the viral Gag complex was not altered by treatment with the nonionic detergent Triton X-100, suggesting a lack of association of this complex with endogenous lipid. Formation of the DRC was not significantly affected by mutations in assembly domains M and L of the Gag protein but was drastically inhibited by a mutation in the assembly I domain. Purified DRC could be disrupted by high-salt treatment, suggesting electrostatic interactions are important for stabilizing the DRC. The Gag precursor proteins in the DRC were more sensitive to trypsin digestion than those in the DSC. These findings suggest that HIV-1 Gag and Gag-Pol precursors assemble into DRC in the cytoplasm, a process which requires the protein-protein interaction domain (I) in NCp7; subsequently, the DRC is transported to the plasma membrane through a process mediated by the M domain of the matrix protein. It appears that during this process, a conformational change might occur in the DRC either before or after its association with the plasma membrane, and this change is followed by the detection of virus budding structure at the plasma membrane.     

Comparative effects of various classes of mouse interferons on macrophage activation for tumor cell killing

The effects of mouse interferon-alpha (MuIFN-alpha), -beta (MuIFN-beta), and -gamma (MuIFN-gamma) on macrophage activation for tumor cell killing were determined by using proteose peptone-elicited peritoneal macrophages from C3H/HeN and C3H/HeJ mice under conditions that either included or were free of detectable endotoxin. Alone, under the conditions used, none of the interferons was able to activate macrophages directly for tumor cell killing. However, with a second signal provided to responsive macrophages by contaminating endotoxin, added bacterial lipopolysaccharide (LPS), or heat-killed Listeria monocytogenes (HKLM), all three types of interferon induced cytolytic activity, with MuIFN-gamma approximately 500 to 1000-fold more active than either MuIFN-alpha or -beta. Thus, all three interferons were able to prime macrophages for killing but required a second signal before cytolytic activity could be expressed. When MuIFN-gamma was mixed with either MuIFN-alpha or -beta and placed on macrophages, little or no killing developed. Mixtures of MuIFN-gamma with either MuIFN-alpha or -beta did increase the sensitivity of macrophages to triggering by LPS, however, compared with macrophages treated with MuIFN-gamma alone. The results are collectively important because they i) confirm that significant quantitative differences exist between the various interferons with regard to their capacity to prime macrophages for tumor cell killing; ii) indicate that to be an efficient activator each type of interferon must be combined with a second stimulus, such as LPS or HKLM; iii) show that neither MuIFN-alpha nor -beta can provide an efficient second triggering signal for macrophages that are primed by MuIFN-gamma; and iv) document that mixtures of MuIFN-gamma with either MuIFN-alpha or -beta are most efficient at inducing priming, compared with any one of the interferons used alone.     

Phylogeny and evolution of sexually selected tail ornamentation in widowbirds and bishops (Euplectes spp.)

Despite similar ecology, mating systems and female preferences for supernormal tails, the 17 species of African widowbirds and bishops (Euplectes spp.) show astonishing variation in male tail ornamentation. Whereas bishops retain their brown nonbreeding tails in nuptial plumage, widowbirds grow black nuptial tails, varying in length from a few centimetres in E. axillaris to the extreme half metre train of E. progne. Here, we phylogenetically reconstruct the evolution of the discrete trait, nuptial tail and the continuous trait, tail length, using a molecular phylogeny of 33 Euplectes subspecies. Unlike many recent findings of labile evolution of plumage ornaments, our results suggest that the nuptial tail of Euplectes is a derived and phylogenetically conserved ornamental trait that, once gained, shows directional evolution in its expression. Directionality is demonstrated in the trivial sense of a short‐tailed ancestor, and by contingency and randomization tests suggesting that branches with increasing tail length are overrepresented. This supports an early origin and strong retention of directional female mate choice in widowbirds and bishops, as previously indicated by empirical and experimental results, and provides a less labile, yet rapid scenario of sexually selected diversification.     

Pollen-based biome reconstruction for southern Europe and Africa 18,000 yr bp

Pollen data from 18,000 ¹⁴C yr bp were compiled in order to reconstruct biome distributions at the last glacial maximum in southern Europe and Africa. Biome reconstructions were made using the objective biomization method applied to pollen counts using a complete list of dryland taxa wherever possible. Consistent and major differences from present‐day biomes are shown. Forest and xerophytic woods/scrub were replaced by steppe, both in the Mediterranean region and in southern Africa, except in south‐western Cape Province where fynbos (xerophytic scrub) persisted. Sites in the tropical highlands, characterized today by evergreen forest, were dominated by steppe and/or xerophytic vegetation (cf. today’s Ericaceous belt and Afroalpine grassland) at the last glacial maximum. Available data from the tropical lowlands are sparse but suggest that the modern tropical rain forest was largely replaced by tropical seasonal forest while the modern seasonal or dry forests were encroached on by savanna or steppe. Montane forest elements descended to lower elevations than today.     

Characterization and partial purification of <Emphasis Type="Italic">Candida albicans</Emphasis> Secretory IL-12 Inhibitory Factor

Background  

We have previously shown that supernatant from Candida albicans (CA) culture contains a Secretory Interleukin (IL)-12 Inhibitory Factor (CA-SIIF), which inhibits IL-12 production by human monocytes. However, the effect of CA-SIIF on secretion of other cytokines by monocytes is unknown, and detailed characterization of this factor has not been performed.     

Comparison of the expression of cell surface poly-N-acetyllactosamine-type oligosaccharides in PC12 cells with those in its variant PC12D.

To explore the biological role of carbohydrate chains in the process of nerve cell differentiation, we carried out a characterization of the carbohydrate structure of glycoproteins by comparing conventional PC12 cells with variant cells (PC12D). In vitro metabolic labeling of cells with either [(3)H] glucosamine or [(3)H] threonine, together with tomato lectin staining, revealed that nerve growth factor (NGF) stimulation caused a decrease in the poly-N-acetyllactosamine synthesis of high-molecular-weight glycopeptides from PC12 cells. By comparison, the amount of glycopeptides with poly-N-acetyllactosamine from PC12D cells was already significantly low and it was not changed by NGF stimulation. By assaying the glycosyltransferases that participate in poly-N-acetyllactosamine synthesis, the decrease in the amount of the poly-N-acetyllactosamine in PC12D cells as well as NGF-stimulated PC12 cells could be accounted for by a reduction in the activity of poly-N-acetyllactosamine extension enzyme (GnT-i), because the amount of poly-N-acetyllactosamine in both cells precisely correlated with changes in GnT-i activity, whereas the activities of N-acetylglucosaminyltransferase V (GnT-V) and beta 1-4 galactosyltransferase remained unchanged. These results demonstrate that the decrease in poly-N-acetyllactosamine synthesis in PC12 cells occurred prior to neurite formation, whereas PC12D cells were insensitive to this effect. Next, we showed that GnT-i but not GnT-V catalyzed a rate-limiting reaction in the expression of poly-N-acetyllactosamine chains, especially in pheochromocytoma.