The cucumber mosaic virus 1a protein regulates interactions between the 2b protein and ARGONAUTE 1 while maintaining the silencing suppressor activity of the 2b protein

The cucumber mosaic virus (CMV) 2b viral suppressor of RNA silencing (VSR) is a potent counter-defense and pathogenicity factor that inhibits antiviral silencing by titration of short double-stranded RNAs. It also disrupts microRNA-mediated regulation of host gene expression by binding ARGONAUTE 1 (AGO1). But in Arabidopsis thaliana complete inhibition of AGO1 is counterproductive to CMV since this triggers another layer of antiviral silencing mediated by AGO2, de-represses strong resistance against aphids (the insect vectors of CMV), and exacerbates symptoms. Using confocal laser scanning microscopy, bimolecular fluorescence complementation, and co-immunoprecipitation assays we found that the CMV 1a protein, a component of the viral replicase complex, regulates the 2b-AGO1 interaction. By binding 2b protein molecules and sequestering them in P-bodies, the 1a protein limits the proportion of 2b protein molecules available to bind AGO1, which ameliorates 2b-induced disease symptoms, and moderates induction of resistance to CMV and to its aphid vector. However, the 1a protein-2b protein interaction does not inhibit the ability of the 2b protein to inhibit silencing of reporter gene expression in agroinfiltration assays. The interaction between the CMV 1a and 2b proteins represents a novel regulatory system in which specific functions of a VSR are selectively modulated by another viral protein. The finding also provides a mechanism that explains how CMV, and possibly other viruses, modulates symptom induction and manipulates host-vector interactions.     

Mechanisms of polyclonal B-cell activation in autoimmune B6-lpr/lpr mice

The influence of the lpr gene on spontaneous and lipopolysaccharide (LPS)-induced immunoglobulin production was studied in B6 mice homozygous for the mutant lpr gene (B6-lpr/lpr). Male and female mice of this congenic strain were followed for 1 year and sera serially tested by the enzyme-linked immunosorbent assay (ELISA) for the production of antibodies to single-stranded DNA (anti-sDNA), immunoglobulin (anti-IgG), and keyhole limpet hemocyanin (anti-KLH), models of autoantibody and non-autoantibody responses, respectively. Female B6-lpr/lpr mice demonstrated marked spontaneous responses to all three antigens; the responses of male B6-lpr/lpr mice were significantly lower but still exceeded those of the congenic B6-+/+ controls. These results demonstrate a generalized influence of sex on lpr associated responses. To determine whether this sex difference could be demonstrated with other forms of B-cell activation, young B6-+/+ and B6-lpr/lpr male and female mice were immunized with lipopolysaccharide and the induced responses determined. This immunization caused significant increases in the IgM response only. The levels of the induced responses produced after LPS treatment were comparable for +/+ and lpr/lpr mice. These results indicate that the enhanced responsiveness of female mice to lpr action is not reflected in the polyclonal response to LPS, which, furthermore, was unaffected by the presence of lpr. The differential influence of sex on lpr and LPS-induced responses and their apparent independence suggests that lpr and LPS promote B-cell activation by dissimilar mechanisms.     

Leukotriene C4 and D4 formation by particulate enzymes

The homogenate of rat basophilic leukemia cells, when incubated with arachidonic acid, glutathione, and calcium, formed 3 isomers of 5,12-dihydroxyeicosatetraenoic acid and 2 isomers of 5,6-dihydroxyeicosatetraenoic acid, as well as leukotriene (LT) C4 and D4. The products were identified by high pressure liquid chromatography, ultraviolet spectral analysis, co-migration with standards, bioassay, and gas chromatography-mass spectrometry. The enzymes responsible for the formation of LTC4 and LTD4 from LTA4 were found in the 10,000 x g pellet and, therefore, appear to be particulate. The possibility that these enzymes are bound to the cell membrane suggest that the formation of these leukotrienes might be important in the basophil and mast cells release reaction.     

Developmental Changes in Polypeptide Composition of, and Precursor Incorporation into, Cellular and Subcellular Fractions of Rat Cerebral Cortex

Abstract: Neuronal-enriched and glial-enriched fractions from rat cerebral cortex at 2. 5, 9, 14 and 23 days postnatally, and subcellular fractions from 2, 14 and 46 day old rat were prepared. The polypeptide composition of all fractions was analysed by sodium dodecyl sulphate (SDS) polyacrylamide gel electro-phoresis and quantified by densitometry. Fifty-nine polypeptides (mol. wts., 13,200–251,000) were resolved in the cell fractions of which the majority remained unchanged throughout postnatal development. Three polypeptides (mol. wts., 102,000, 56,000, 53,700) were found to increase in amount devel-opmentally in both cellular fractions, the latter two showing a peak in relative amount on day 14 and a subsequent decline. Three polypeptides (mol. wts., 47,000, 28,200, 17,400) were found to be common to the glial-enriched fraction as well as the myelin fraction, and all showed a developmental increase. The neuronal-enriched fraction was found to be enriched in five polypeptides of which one (mol. wt., 51,900) showed a developmental increase after ten days postnatally, the others (mol. wts., 178,700, 142,000, 109,000, 24,000) showing a decrease. In vitro incorporation of [³⁵S]-methionine into the glial-enriched fraction was carried out, and a developmental decline was observed in the labelling of a polypeptide of 42,000 mol. wt.     

Cervical lymphadenitis in guinea pigs: infection via intact ocular and nasal mucosa by Streptococcus zooepidemicus.

The traditional view regarding the pathogenesis of cervical lymphadenitis in guinea pigs is that Lancefield Group C Streptococcus gains access to cervical lymph nodes via an abraded oral mucosa. In this study, it is established that inoculation of intact nasal and conjunctival mucous membranes with Streptococcus zooepidemicus (Lancefield Group C) also can produce the disease. Weanling (SPF) guinea pigs (Cavia porcellus) were divided into two experimental groups of 10 and two control groups of four each. Guinea pigs from each group were individually housed in separate cubicles. Group I was inoculated with 0.05 ml of culture containing 2.8 x 10(7) CFU/ml of S. zooepidemicus into the conjunctiva of the left eye. Group II received a similar inoculum into the left nares. Control groups received 0.05 ml of TSB broth in the same sites. Five of ten guinea pigs in Group II died four to nine days postinoculation. Surviving guinea pigs were euthanatized at intervals between days 4-13 postinoculation. All guinea pigs were necropsied, cultured and examined for evidence of infection. S. zooepidemicus was recovered from 30/50 and 39/46 sites cultured from Groups I and II, respectively. Lymphadenitis was found in cervical lymph nodes from 8/10 guinea pigs in Group I and 10/10 in Group II. The conjunctival and nasal mucosa, therefore, represent potential sites of entry resulting in cervical lymphadenitis in guinea pigs.