Key residues characteristic of GATA N-fingers are recognized by FOG

Protein-protein interactions play significant roles in the control of gene expression. These interactions often occur between small, discrete domains within different transcription factors. In particular, zinc fingers, usually regarded as DNA-binding domains, are now also known to be involved in mediating contacts between proteins. We have investigated the interaction between the erythroid transcription factor GATA-1 and its partner, the 9 zinc finger protein, FOG (Friend of GATA). We demonstrate that this interaction represents a genuine finger-finger contact, which is dependent on zinc coordinating residues within each protein. We map the contact domains to the core of the N-terminal zinc finger of GATA-1 and the 6th zinc finger of FOG. Using a scanning substitution strategy we identify key residues within the GATA-1 N-finger which are required for FOG binding. These residues are conserved in the N-fingers of all GATA proteins known to bind FOG, but are not found in the respective C-fingers, This observation may, therefore, account for the particular specificity of FOG for N-fingers, Interestingly, the key N-finger residues are seen to form a contiguous surface, when mapped onto the structure of the N-finger of GATA-1.

Social identity and perceptions of media persuasion: Are we always less influenced than others?

People typically perceive negative media content (e.g., violence) to have more impact on others than on themselves (a third-person effect). To examine the perceived effects of positive content (e.g., public-service advertisements) and the moderating role of social identities, we examined students' perceptions of the impact of AIDS advertisements on self, students (in- group), nonstudents (out-group), and people in general. Perceived self-other differences varied with the salience of student identity. Low identifiers displayed the typical third-person effect, whereas high identifiers were more willing to acknowledge impact on themselves and the student in-group. Further, when influence was normatively accept able within the in-group, high identifiers perceived self and students (us) as more influenced than nonstudents (them). The theoretical and practical implications of this reversal in third-person perceptions are discussed.

HPV6b virus like particles are potent immunogens without adjuvant in man

Subjects with genital warts were immunized three times or more with HPV6b VLPs without adjuvant. All immunized subjects had DTH to HPV6b L1 protein. Of 32 subjects, nine had HPV6b specific antibody prior to immunization and 22 acquired antibody with immunization. VLP specific antibody increased following a single immunization in 6 of 8 subjects with low level antibody at recruitment. Complete regression of genital warts was observed in 25 of 33 evaluable subjects over the 20-week observation period. We conclude that immunization with HPV6b L1 VLPs without adjuvant induces immunity to the L1 protein epitopes recognised during natural infection, and may accelerate regression of warts. (C) 2000 Elsevier Science Ltd. All rights reserved.

The salicylate-derived mycobactin siderophores of Mycobacterium tuberculosis are essential for growth in macrophages

Mycobacterium tuberculosis is an important pathogen of mammals that relies on 2-hydroxyphenyloxazoline-containing siderophore molecules called mycobactins for the acquisition of iron in the restrictive environment of the mammalian macrophage, These compounds have been proposed to be biosynthesized through the action of a cluster of genes that include both nonribosomal peptide synthase and polyketide synthase components. One of these genes encodes a protein, MbtB, that putatively couples activated salicylic acid with serine or threonine and then cyclizes this precursor to the phenyloxazoline ring system. We have used gene replacement through homologous recombination to delete the mbtB gene and replace this with a hygromycin-resistance cassette in the virulent strain of M. tuberculosis H37Rv, The resulting mutant is restricted for growth in iron-limited media but grows normally in iron-replete media. Analysis of siderophore production by this organism revealed that the biosynthesis of all salicylate-derived siderophores was interrupted. The mutant was found to be impaired for growth in macrophage-like THP-1 cells, suggesting that siderophore production is required for virulence of M. tuberculosis, These results provide conclusive evidence linking this genetic locus to siderophore production.

Differentiated dendritic cells expressing nuclear RelB are predominantly located in rheumatoid synovial tissue perivascular mononuclear cell aggregates

Objective. Differentiated dendritic cells (DC) and other antigen-presenting cells are characterized by the nuclear location of RelB, a member of the nuclear factor kappa B/Rel family. To characterize and enumerate differentiated DC in rheumatoid arthritis (RA) peripheral blood (PB), synovial fluid (SF), and synovial tissue (ST), the expression and location of RelB were examined. Methods. RelB protein expression and cellular location were determined in RA PB, SF, and ST by flow cytometry and immunohistochemical analysis of purified cells or formalin-fixed tissue. DNA-binding activity of RelB was determined by electrophoretic: mobility shift-Western immunoblotting assays. Results. Circulating RA PBDC resembled normal immature PBDC in that they did not express intracellular RelB protein. In RA ST serial sections, cells containing nuclear RelB (nRelB) were enriched in perivascular regions. A mean +/- SD of 84 +/- 10% of these cells were DC. The remaining nRelB+,HLA-DR+ cells comprised B cells and macrophages. Only 3% of sorted SFDC contained nRelB, However, RelB present in the nucleus of these SFDC was capable of binding DNA, and therefore capable of transcriptional activity. Conclusion. Circulating DC precursors differentiate and express RelB after entry into rheumatoid ST. Differentiated DC can thus be identified by immunohistochemistry in formalin-fixed ST. Signals for DC maturation may differ between RA ST and SF, resulting in nuclear location of RelB predominantly in ST. This is likely to have functional consequences for the DC in these sites.

Neurons in the hilus region of the rat hippocampus are depleted in number by exposure to alcohol during early postnatal life

We have previously shown that exposing rats to a relatively high dose of ethanol during early postnatal life resulted in a deficit in spatial learning ability. This ability is controlled, at least in part, by the hippocampal formation. The purpose of the present study was to determine whether exposure of rats to ethanol during early postnatal life affected the number of specific neurons in the hippocampus. Wistar rats were exposed to a relatively high daily dose of ethanol between postnatal days 10 and 15 by placing them for 3 h each day in a chamber containing ethanol vapor. The blood ethanol concentration was about 430 mg/dl at the end of the exposure period. Groups of ethanol-treated (ET) rats, separation controls (SC), and mother-reared controls (MRC) were anesthetized and killed at 16 days of age by perfusion with phosphate-buffered glutaraldehyde (2.5%). The Cavalieri principle was used to determine the volume of various subdivisions of the hippocampal formation (CA1, CA2+CA3, hilus, and granule cell layer), and the physical disector method was used to estimate the numerical densities of neurons within each subdivision. The total number of neurons was calculated by multiplying estimates of the numerical density with the volume. There were, on average, about 441,000 granule cells in the granule cell layer and 153,000 to 177,000 pyramidal cells in both the CA1 and CA2+CA3 regions in all three treatment groups. In the hilus region, ET rats had about 27,000 neuronal cells. This was significantly fewer than the average of 38,000 such neurons estimated to be present in both MRC and SC animals. Thus, neurons in the hilus region may be particularly vulnerable to the effects of a high dose of ethanol exposure during early postnatal life. (C) 2000 Wiley-Liss, Inc.

Relationships among climate, latitude and migration: Australian butterflies are not temperate-zone birds

We examined the distribution of butterflies over the mostly arid and semi-arid continent of Australia and analyzed the proportion of migrant species and species diversity with respect to an array of climatic and geographic variables. On a continent-wide scale, latitude explained virtually no variance in either proportion of migrants (r(2) = 0.01) or species diversity (r(2) = 0.03) in Australian butterflies. These results are in marked contrast to those for temperate-zone birds from three continents where latitude explained between 82 and 98% of the variance in frequency of migrants and also accounted for much of the variance in bird species diversity. In eastern Australia where rainfall regimes are similar to those in temperate Europe and North and South America, latitude explains 78% of the variance in frequency of butterfly migrants. In both eastern and central Australia, latitude also accounts for relatively high proportions of the variance in species diversity. Rainfall patterns and especially soil moisture are negatively associated with migration frequency in Australian butterfly faunas, both alone and in combination with other climate variables. Where moisture levels are relatively high, as in eastern Australia, measures of temperature are associated with migration frequency, a result consistent with findings for temperate-zone birds, suggesting latitude is a surrogate for temperature. The ultimate causes of migration in temperate-zone birds and Australian butterflies are the uneven temporal, and in Australia also spatial, distribution of resources. Uneven distribution is brought about primarily by temperature in temperate regions and by erratic rainfall over much of arid Australia. As a key determinant of productivity, especially in the tropics and subtropics, aridity is likely to be an important determinant of the global distributions of migrants.

Osmolarity effects on observed insect cell size after baculovirus infection are avoided using growth medium for sample dilution

Rates of cell size increase are an important measure of success during the baculovirus infection process. Batch and fed batch cultures sustain large fluctuations in osmolarity that can affect the measured cell volume if this parameter is not considered during the sizing protocol. Where osmolarity differences between the sizing diluent and the culture broth exist, biased measurements of size are obtained as a result of the cell osmometer response. Spodoptera frugiperda (Sf9) cells are highly sensitive to volume change when subjected to a change in osmolarity. Use of the modified protocol with culture supernatants for sample dilution prior to sizing removed the observed error during measurement.

Multiple conformational epitopes are recognized by natural and induced immunity to the E7 protein of human papilloma virus type 16 in man

The reactivity of sera from patients with cervical cancer with the E7 protein of human papilloma virus type 16 (HPV16) was estimated using a novel non-radioactive immunoprecipitation assay and four established protein-and peptide-based immunoassays. Six of 14 sera from patients with cervical cancer and 1 of 10 sera from healthy laboratory staff showed repeated reactivity with E7 in at least one assay. Four of the 7 reactive sera were consistently reactive in more than one assay, but only one was reactive in all four assays. Following immunization with E7, 2 of 5 patients with cervical cancer had increased E7-specific reactivity, measurable in one or more assays. No single assay was particularly sensitive for E7 reactivity, or predictive of cervical cancer. Mapping of E7 reactivity to specific E7 peptides was unsuccessful, suggesting that natural or induced E7 reactivity in human serum is commonly directed to conformational epitopes of E7, These results suggest that each assay employed with is study measures a different aspect of E7 reactivity, and that various reactivities to E7 may manifest following HPV infection or immunization. This finding is of significance for monitoring of E7 immunotherapy and for serological screening for cervical cancer. Copyright (C) 2000 S.Karger, AG. Basel.

Resident tissue macrophages within the normal rat iris lack immunosuppressive activity and are effective antigen-presenting cells

Despite extensive study of the numerous immunoregulatory mechanisms that contribute to the immune-privileged nature of the anterior chamber (AC) of the eye, little is known of the functional nature of antigen-presenting cells (APC) present in the tissues adjoining the AC. In the present study, we have compared the antigen-presenting capacity of dendritic cells (DC) and macrophages isolated from the normal rat iris. Whereas iris DC exhibited a potent ability to stimulate resting allogeneic T cells in MLR cultures (an in-vitro correlate of the ability to induce primary T cell responses), resident iris macrophages displayed negligible MLR-stimulatory capacity. Significantly, iris macrophages could efficiently elicit proliferation of primed antigen-specific T cells (an in-vitro correlate of the ability to act as local APC in secondary responses). This antigen-presenting activity was approximately half that of fully mature iris DC and considerably greater than that of freshly isolated iris DC. A key contributor to the effectiveness of resident iris macrophage antigen presentation was considered to be the absence of lymphocytostatic control of T cell proliferation exerted by these cells. The results indicate dichotomous but complementary roles for DC (immune surveillance) and macrophages (local antigen presentation in secondary responses) in this tissue.