A monoclonal antibody to glycoprotein gp85 inhibits fusion but not attachment of Epstein-Barr virus.

Epstein-Barr virus (EBV) codes for at least three glycoproteins, gp350, gp220, and gp85. The two largest glycoproteins are thought to be involved in the attachment of the virus to its receptor on B cells, but despite the fact that gp85 induces neutralizing antibody, no function has been attributed to it. As an indirect approach to understanding the role of gp85 in the initiation of infection, we determined the point at which a neutralizing, monoclonal antibody that reacted with the glycoprotein interfered with virus replication. The antibody had no effect on virus binding. To examine the effect of the antibody on later stages of infection, the fusion assay of Hoekstra and colleagues (D. Hoekstra, T. de Boer, K. Klappe, and J. Wilshaut, Biochemistry 23:5675-5681, 1984) was adapted for use with EBV. The virus was labeled with a fluorescent amphiphile that was self-quenched at the high concentration obtained in the virus membrane. When the virus and cell membrane fused, there was a measurable relief of self-quenching that could be monitored kinetically. Labeling had no effect on virus binding or infectivity. The assay could be used to monitor virus fusion with lymphoblastoid lines or normal B cells, and its validity was confirmed by the use of fixed cells and the Molt 4 cell line, which binds but does not internalize the virus. The monoclonal antibody to gp85 that neutralized virus infectivity, but not a second nonneutralizing antibody to the same molecule, inhibited the relief of self-quenching in a dose-dependent manner. This finding suggests that gp85 may play an active role in the fusion of EBV with B-cell membranes.     

Recombinant human IL-2 overcomes genetic nonresponsiveness to malaria sporozoite peptides. Correlation of effect with biologic activity of IL-2

Current malaria vaccine strategies focus on subunit vaccines that contain one or a limited number of malaria Ag. However, there is widespread nonresponsiveness to many of these Ag probably resulting from Ir gene control. Using a congenic mouse model, we demonstrated that human rIL-2 (as an adjuvant) can overcome Ir gene controlled low immune responsiveness to peptide malaria Ag vaccine candidates [R32tet32, R32LR, and Th2R-NP (NANP)5NA] as determined by the antibody response, providing it is emulsified with the Ag during immunization. This effect is not caused by IL-2 merely acting as a foreign protein and stimulating noncognate help; it requires biologic activity of the IL-2, as determined by studying the effect of inactive rIL-2, which has minimal biologic activity but which has retained its antigenicity. IL-2 does not appear to be working by an effect on priming of specific Th, and IL-2 cannot overcome an Ir gene controlled low T cell proliferative response. IL-2 may have a role to play in human vaccine development where a high titer antibody response to a subunit vaccine is required.     

Autoimmune CD4+ T cells interfere with immune tolerance to a thymic-independent antigen

The ability of autoimmune T cell subsets to interfere with tolerization of B cells can be studied by using thymic-independent Ag. We have defined an abnormality within the CD4+ T cell compartment in young NZB and MRL-lpr/lpr mice by studying tolerance of spleen and B cells to the thymic independent Ag, fluorescein-Brucella abortus. Tolerization of spleen cells is defective in MRL-lpr/lpr mice, but not MRL-+/+ or C3H.lpr mice, suggesting that the defect requires both the autosomal MRL background and the lpr gene to be present. T enriched cells from NZB mice and from MRL-lpr/lpr mice (but not MRL-+/+ or C3H.lpr mice) reverse tolerance in spleen cells from [NZB X DBA/2]F1 and C3H/HeJ mice, respectively. This interference is removed by treatment with anti-CD4 antibody and C. Supernatants from cultured T cells of NZB and MRL-lpr/lpr mice also prevent tolerance in spleen cells of [NZB X DBA/2]F1 and MRL-+/+ mice, respectively, unless CD4+ cells are removed prior to T cell culture. Removal of T cells from NZB and MRL-lpr/lpr spleen cells allows normal tolerization of B cells, which is abrogated by the addition of syngeneic T cells or cultured T cell supernatants. This effect also depends on the presence of CD4+ T cells. These studies show that in MRL-lpr/lpr mice, through interaction of the lpr and MRL background genes in a T cell subset, and in NZB mice, CD4+ T cells interfere with B cell tolerance to a thymic-independent Ag.     

Fatty acid binding proteins in the three types of rat skeletal muscle

By means of Sephadex G-50 column chromatography, a Mr 12,000 fatty acid binding protein (FABP) was found to be present in all three types of skeletal muscle. FABP concentrations in muscle cytosols (105,000g supernatant) were fiber type specific with binding levels (expressed as pmole [14C]oleate bound/mg protein) of 70 +/- 7 in fast-twitch white (FTW) (heart FABP = 469 +/- 33). Cytosols from all three fiber types cross-reacted with antibody to pure heart FABP on Ouchterlony immunodiffusion analysis. FABP content, determined by radial immunodiffusion, followed the same pattern in the muscle types as that in the binding assay. The values (in micrograms/mg protein) were 3.3 +/- 0.1 in FTW, 17.0 +/- 0.4 in FTR, and 31.7 +/- 1.4 in STR fibers (heart = 55). Disc gel electrophoresis revealed a protein band in each fiber type that had migration identical to that of pure heart FABP and was proportional to the amounts determined by Sephadex G-50 chromatography and immunoassay. In addition, Western blots of tissue cytosols, using antibody to heart FABP, detected single protein bands identical in size to pure heart FABP in all three types of skeletal muscle. These results show the presence of a FABP in all skeletal muscle types with an immunologic and electrophoretic characterization identical to that of heart FABP.     

Recovery of murine lens epithelial cells from single and fractionated doses of X rays and neutrons

Subpopulations of mouse lens epithelial cells, differing in proliferative status, were irradiated with either X rays or fission spectrum neutrons given singly or in four weekly fractions. After various times, epithelia were mitogenically stimulated by wounding and DNA synthesis responses were determined by incorporation of [3H]thymidine. At 1 h following both X and neutron irradiations, significant suppression of the wound response after single doses and a sparing effect of fractionation were evident in both the mitotically quiescent and the slowly proliferating subpopulations. At 1 week following single or fractionated doses of both radiations, recovery was evident in both subpopulations. By 4 weeks, the quiescent subpopulation showed significant recovery after both single and fractionated doses of X rays or neutrons. In contrast, a marked decreased ability to respond after neutron irradiation and, in addition, a significant enhancement effect of neutron fractionation were observed for the slowly proliferating subpopulation. Per gray, neutrons were about 7.5 times more effective than X rays as a single dose and 25 times more effective in four equal fractions. The shift from an initial sparing to a final enhancing effect of neutron fractionation for the slowly proliferating subpopulation has importance for understanding divergent early and late radiation responses following dose fractionation.     

Stimulation of ciliary beat frequency by autonomic agonists: in vivo

beta 2-Adrenergic bronchodilator and muscarinic cholinergic bronchoconstrictor agonists both stimulate ciliary activity in vitro. To test the hypothesis that increases in autonomic activity would result in increases in ciliary beat frequency (CBF) in vivo, a correlation analysis heterodyne laser light-scattering system was developed and validated to measure the stimulating effects of sympathomimetic and parasympathomimetic agonists on tracheal CBF in intact, anesthetized beagles. The mean baseline CBF from 42 studies of 274 measurements in 9 (5 male and 4 female) adult beagles was 6.6 +/- 1.1 Hz. The stimulating effects of a beta 2-adrenergic agonist, fenoterol, and a muscarinic cholinergic agonist, methacholine, on CBF were studied on four and eight beagles, respectively. The studies were randomized and blinded. Aerosolized 10(-5) M fenoterol stimulated the CBF from the base line of 6.8 +/- 2.5 to 32.0 +/- 17.9 Hz in four dogs. Aerosolized methacholine stimulated the CBF from the base line of 5.8 +/- 0.7 to 9.4 +/- 3.0 Hz for 10(-8) M, and to 12.6 +/- 3.1 Hz for 10(-6) M in eight dogs. These are the first data obtained in intact animals that demonstrate CBF in the lower respiratory tract is regulated by autonomic agonists.     

The effects of low levels of chloramphenicol and methotrexate on somatic embryogenesis inCitrus

Summary The effects of the antibiotics methotrexate and chloramphenicol on somatic embryogenesis inCitrus were evaluated. Relatively low levels (0.1 to 1.0 μg/ml) of these antibiotics did not inhibit embryo production from undeveloped ovules of ‘Key’ lime [C. aurantifolia) (Christm.) Swing.]. Surprisingly, both antibiotics induced the formation of embryogenic callus in these cultures. This is usually a rare event in cultures of undevelopedCitrus ovules, and ‘Key’ lime is especially recalcitrant. The effects of these antibiotics on embryogenic callus appeared to be limited to the induction stage, because there was no consistent effect, either stimulatory or inhibitory, on established, lines of embryogenic callus. Florida Agricultural Experiment Station Journal Series No. 8958. This research was supported in part by a grant to Moore and Cline from the Competitive Grants Office of the SEA, USDA (85-CRCR-1-1623).     

Optimal alignments in linear space

Space, not time, is often the limiting factor when computingoptimal sequence alignments, and a number of recent papers inthe biology literature have proposed space-saving strategies.However, a 1975 computer science paper by Hirschberg presenteda method that is superior to the new proposals, both in theoryand in practice. The goal of this paper is to give Hirschberg'sidea the visibility it deserves by developing a linear-spaceversion of Gotoh's algorithm, which accommodates affine gappenalties. A portable C-software package implementing this algorithmis available on the BIONET free of charge. Received on October 14, 1987; accepted on December 19, 1987