Maternal transmission of type D simian retrovirus (SRV-2) in pigtailed macaques

Pregnant macaques were used as a natural model for maternal-infant transmission of SRV-2 retrovirus. Fifty-one pregnant females were placed into one of four virus/antibody groups. Nonviremic mothers produced 100% virus-negative offspring at birth. In contrast, viremic mothers produced offspring which were 17% virus-negative and 83% virus-positive at birth. SRV-2 infection occurred principally in utero by the transplacental route. Infants born to viremic mothers exhibited low birth weight, prematurity, high perinatal death, and increased incidence of SAIDS.     

Molecular location of a species-specific epitope on the hamster scrapie agent protein.

Scrapie is a transmissible neurodegenerative disease of sheep and goats. An abnormal host protein, Sp33-37, is the major protein component of the scrapie agent and the only known disease- or agent-specific macromolecule. Two monoclonal antibodies (MAbs), 4H8 (immunoglobulin G2b [IgG2b]) and 6B11 (IgG1), produced by immunizing mice with the intact hamster 263K scrapie agent protein, Sp33-37Ha, were found to have species specificity similar to that reported previously for MAb 3F4 (IgG2a), which was produced by using PrP-27-30 as the immunogen (R. J. Kascsak, R. Rubenstein, P. A. Merz, M. Tonna-DeMasi, R. Fersko, R. I. Carp, H. M. Wisniewski, and H. Diringer, J. Virol. 61:3688-3693, 1987). These antibodies all bound to Sp33-37 derived from hamster but not from mouse cells. Competitive binding assays demonstrated that all three MAbs bound to the same or overlapping sites on Sp33-37Ha. The molecular location of the epitope for these antibodies was determined to within 10 residues by using an antigen competition enzyme-linked immunosorbent assay in which synthetic peptides spanning Sp33-37Ha residues 79 to 93 or 84 to 93 specifically inhibited binding of these antibodies to plates coated with purified Sp33-37Ha. A synthetic peptide with the mouse-specific sequence (83 to 92) that differed from the hamster sequence by substitution at two positions (MetHa-87----LeuMo-86 and MetHa-90----ValMo-89) did not inhibit antibody binding to Sp33-37Ha. MAb 3F4 binding to hamster Sp33-37 was eliminated by chemical modification of Sp33-37Ha with diethylpyrocarbonate or succinic anhydride and by cleavage with CNBr or trypsin. The effect of diethylpyrocarbonate on MAb 3F4 binding was not reversed by hydroxylamine treatment. MAb 3F4 binding was not affected by prolonged exposure of Sp33-37Ha to 70% formic acid or by boiling in sodium dodecyl sulfate. We conclude that the epitope for these MAbs is a linear determinant that includes Met-87, Lys-88, and Met-90 and that Met-90 is probably the major species-specific determinant.     

Plasmid-mediated NAD independence in Haemophilus parainfluenzae.

The location of the genes coding for NAD independence in four unusual clinical isolates of Haemophilus parainfluenzae was determined by transferring these genes to plasmid-free Haemophilus influenzae Rd by transformation and analysing transformants for the presence of plasmids by agarose gel electrophoresis. All NAD-independent transformants were found to carry a single plasmid species. The plasmids, originally harboured by the four H. parainfluenzae isolates recovered from unrelated sources, were of the same size (5.25 kb). Spontaneous reversion to NAD dependence occurred with a low frequency (0.1 to 0.2% of the progeny of a single clone) in both H. parainfluenzae and H. influenzae Rd. The revertants had lost this small plasmid. Mitomycin C exhibited a plasmid 'curing' effect with a frequency of 'curing' of between 1 and 6% of the surviving clones. It was concluded that the genes conferring NAD independence were located on the small 5.25 kb plasmid.     

Selective proteolytic cleavage of recombinant human interleukin 4. Evidence for a critical role of the C-terminus

Human interleukin 4 is a 129 amino acid lymphokine secreted by activated T cells that exerts pleiotropic biological effects on B and T lymphocytes and other hematopoietic cells. Structure-function relations were studied by employing selective proteolytic cleavage of purified recombinant human interleukin 4 (rhuIL-4). Limited proteolysis with endoprotease Glu-C from Staphylococcus aureus (V8) produced two digestion products that were observed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with apparent molecular weight values of 19K (I) and 15K (II), respectively. These species were isolated by reversed-phase HPLC. Amino acid sequencing indicated that species II was an 84 amino acid core fragment extending from Gln-20 to Glu-103 and containing a hydrolyzed peptide bond at Glu-26. On the basis of known disulfide bond assignments, it was concluded that species II was stabilized by two disulfide bonds (Cys-24/Cys-65 and Cys-46/Cys-99). Analysis of its secondary structure by circular dichroism revealed a high content of alpha helix. Species I was the full-length rhuIL-4 with selective cleavage at Glu-26 and Glu-103. Both species I and II were inactive in an in vitro assay based on proliferation of peripheral blood lymphocyte blasts and lacked the ability to bind to teh rhuIL-4 receptor on Daudi cells. In order to elucidate further the role of the residues removed by S. aureus V8 protease, rabbit antisera were raised to synthetic peptides corresponding to residues 1-26 at the N-terminus and 104-129 at the C-terminus. Only antisera directed to the C-terminal peptide inhibited binding of 125I-rhuIL-4 to Daudi cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modification of lipid-related atherosclerosis risk factors by ω3 fatty acid ethyl esters in hypertriglyceridemic patients

The efficacy of ω3 fatty acid ethyl esters was evaluated in 10 mildly hypertriglyceridemic patients in this randomized, placebo-controlled, double-blind, crossover trial. Patients were given capsules (1 per 10 kg body weight) containing 640 mg/g of ω3 fatty acids or an olive oil placebo for two 4-week treatment periods separated by a 1-week washout phase. Plasma lipids, lipoproteins, and apolipoproteins: phospholipid FA composition; the susceptibility to oxidation of the apolipoprotein B-100 containing lipoproteins; and bleeding times were determined at the end of each period. Plasma triglyceride levels were reduced by 37% (P < 0.001), whereas low density lipoprotein cholesterol and the cholesterol content of subfraction 2 of high density lipoproteins increased by 23 and 56%, respectively (both P < 0.02). Changes in plasma lipid parameters and in phospholipid FA patterns occurred rapidly, usually stabilizing within 1 week, and returned to baseline levels within 10 days after stopping supplementation with ω3 fatty acids. Bleeding times were not changed. However, the susceptibility of lipoproteins to oxidation was increased during the ω3 fatty acid period. We conclude that ω3 fatty acid ethyl esters are effective hypotriglyceridemic agents, and that they impact lipoprotein metabolism very quickly. How they may alter the atherogenic process is not clear from this study because some risk factors worsened and other improved.     

Effects of Foliar Endophytic Fungi on the Preference and Performance of the Leaf Beetle Chelymorpha alternans in Panama

Foliar endophytic fungi live inside healthy plant leaves, and in some cases they confer herbivore resistance to the host. All previous studies of endophyte–herbivore interactions have occurred in temperate areas, and many use correlations rather than experiments. In Panama, Glomerella cingulata is a common endophyte species found in healthy leaves, and Chelymorpha alternans is a common herbivore on Merremia umbellata , a tropical vine. We manipulated the abundance of G. cingulata in the leaves of M. umbellata . We then assessed the effects of high and low endophyte densities on the food choice, development, and reproductive success of the leaf beetle, C. alternans . In 'choice' experiments, adult females with a history of feeding on wild plants showed no preference when offered food plants with high and low endophyte densities. Further, in 'no-choice' experiments, C. alternans larvae that were fed high- or low-density endophyte leaves did not differ in development or survivorship. However, when larvae fed on leaves with low endophyte densities became adults, they produced 80 percent more offspring. This suggests high endophyte levels in hosts can have a negative effect on herbivore fecundity. Further experiments are necessary to understand whether the reproductive effects are due to feeding on low-density endophytes in the larval or adult stages, and whether changes in reproductive success are motivated by the adult's perception of food quality or by physiological constraint due to food quality.     

Wolbachia modification of sperm does not always require residence within developing sperm

Wolbachia are maternally inherited intracellular bacteria known to manipulate the reproduction of their arthropod hosts. Wolbachia commonly affect the sperm of infected arthropods. Wolbachia-modified sperm cannot successfully fertilize unless the female is infected with the same Wolbachia type. A study of spermatogenesis in the parasitic wasp Nasonia vitripennis reveals that Wolbachia are not required in individual spermatocytes or spermatids to modify sperm. In N. vitripennis, Wolbachia modify nearly all sperm, but are found only in approximately 28% of developing sperm, and are also found in surrounding cyst and sheath cells. In the beetle Chelymorpha alternans, Wolbachia can modify up to 90% of sperm, but were never observed within the developing sperm or within the surrounding cyst cells; they were abundant within the outer testis sheath. We conclude that the residence within a developing sperm is not a prerequisite for Wolbachia-induced sperm modification, suggesting that Wolbachia modification of sperm may occur across multiple tissue membranes or act upstream of spermiogenesis.     

Structure-guided discovery of cyclin-dependent kinase inhibitors

CDK2 inhibitors containing the related bicyclic heterocycles pyrazolopyrimidines and imidazopyrazines were discovered through high-throughput screening. Crystal structures of inhibitors with these bicyclic cores and two more related ones show that all but one have a common binding mode featuring two hydrogen bonds (H-bonds) to the backbone of the kinase hinge region. Even though ab initio computations indicated that the imidazopyrazine core would bind more tightly to the hinge, pyrazolopyrimidines gain an advantage in potency through participation of N4 in an H-bond network involving two catalytic residues and bridging water molecules. Further insight into inhibitor/CDK2 interactions was gained from analysis of additional crystal structures. Significant gains in potency were obtained by optimizing the fit of hydrophobic substituents to the gatekeeper region of the ATP binding site. The most potent inhibitors have good selectivity.