Tempo and mode of early gene loss in endosymbiotic bacteria from insects

Background  

Understanding evolutionary processes that drive genome reduction requires determining the tempo (rate) and the mode (size and types of deletions) of gene losses. In this study, we analysed five endosymbiotic genome sequences of the gamma-proteobacteria (three different Buchnera aphidicola strains, Wigglesworthia glossinidia, Blochmannia floridanus) to test if gene loss could be driven by the selective importance of genes. We used a parsimony method to reconstruct a minimal ancestral genome of insect endosymbionts and quantified gene loss along the branches of the phylogenetic tree. To evaluate the selective or functional importance of genes, we used a parameter that measures the level of adaptive codon bias in E. coli (i.e. codon adaptive index, or CAI), and also estimates of evolutionary rates (Ka) between pairs of orthologs either in free-living bacteria or in pairs of symbionts.     

Endothelial targeting and enhanced antiinflammatory effects of complement inhibitors possessing sialyl Lewisx moieties

The complement inhibitor soluble complement receptor type 1 (sCR1) and a truncated form of sCR1, sCR1[desLHR-A], have been generated with expression of the selectin-reactive oligosaccharide moiety, sialyl Lewisx (sLex), as N-linked oligosaccharide adducts. These modified proteins, sCR1sLex and sCR1[desLHR-A]sLex, were assessed in the L-selectin- and P-selectin-dependent rat model of lung injury following systemic activation of complement by cobra venom factor and in the L-selectin-, P-selectin-, and E-selectin-dependent model of lung injury following intrapulmonary deposition of IgG immune complexes. In the cobra venom factor model, sCR1sLex and sCR1[desLHR-A]sLex caused substantially greater reductions in neutrophil accumulation and in albumin extravasation in lung when compared with the non-sLex-decorated forms. In this model, increased lung vascular binding of sCR1sLex and sCR1[desLHR-A]sLex occurred in a P-selectin-dependent manner, in contrast to the absence of any increased binding of sCR1 or sCR1[desLHR-A]. In the IgG immune complex model, sCR1[desLHR-A]sLex possessed greater protective effects relative to sCR1[desLHR-A], based on albumin extravasation and neutrophil accumulation. Enhanced protective effects correlated with greater lung vascular binding of sCR1[desLHR-A]sLex as compared with the non-sLex-decorated form. In TNF-alpha-activated HUVEC, substantial in vitro binding occurred with sCR1[desLHR-A]sLex (but not with sCR1[desLHR-A]). This endothelial cell binding was blocked by anti-E-selectin but not by anti-P-selectin. These data suggest that sLex-decorated complement inhibitors have enhanced antiinflammatory effects and appear to have enhanced ability to localize to the activated vascular endothelium.     

Critical role for hematopoietic cell kinase (Hck)-mediated phosphorylation of Gab1 and Gab2 docking proteins in interleukin 6-induced proliferation and survival of multiple myeloma cells

Interleukin-6 (LI-6) is a known growth and survival factor in multiple myeloma via activation of extracellular signal-regulated kinase and phosphatidylinositol 3-kinase signaling cascade. In this report we show that Grb2-associated binder (Gab) family adapter proteins Gab1 and Gab2 are expressed by multiple myeloma cells; and that interleukin-6 induces their tyrosine phosphorylation and association with downstream signaling molecules. We further demonstrate that these events are Src family tyrosine kinase-dependent and specifically identify the role of hematopoietic cell kinase (Hck) as a new Gab family adapter protein kinase. Conversely, inhibition of Src family tyrosine kinases by the pyrazolopyrimidine PP2, as in kinase-inactive Hck mutants, significantly reduces IL-6-triggered activation of extracellular signal-regulated kinase and AKT-1, leading to significant reduction of multiple myeloma cell proliferation and survival. Taken together, these results delineate a key role for Hck-mediated phosphorylation of Gab1 and Gab2 docking proteins in IL-6-induced proliferation and survival of multiple myeloma cells and identify tyrosine kinases and downstream adapter proteins as potential new therapeutic targets in multiple myeloma.     

Floral structure, breeding system and fruit-set in the threatened sub-shrub Tetratheca juncea Smith (Tremandraceae)

Tetratheca juncea Smith (Tremandraceae) has undergone a range contraction of approx. 50 km in the last 100 years and is now listed as a vulnerable sub-shrub restricted to the central and north coast regions of New South Wales, Australia. There are approx. 250 populations in a 110 km north-south distribution and populations are usually small with fewer than 50 plants/clumps. The reproductive ecology of the species was studied to determine why seed-set is reportedly rare. Flowers are bisexual, odourless and nectarless. Flowers are presented dependentally and there are eight stamens recurved around the pistil. Anthers are poricidal, contain viable pollen and basally contain a deep-red tapetal fluid that is slightly oily. Thus flowers are presented for buzz pollinators, although none were observed at flowers during our study. The species was found to be facultatively xenogamous with only one in 50 glasshouse flowers setting seed autogamously, i.e. without pollinator assistance. Field studies revealed fertile fruit in 24 populations but production varied significantly across sites from exceedingly low (0.6 fruits per plant clump) to low (17 fruits per plant clump). Fruit-set ranged from 0 to 65 %, suggesting that pollen vectors exist or that autogamy levels in the field are variable and higher than glasshouse results. Fruit production did not vary with population size, although in three of the five populations in the south-west region more than twice as much fruit was produced as in populations elsewhere. A moderately strong relationship between foliage volume and fruit : flower ratios suggests that bigger plants may be more attractive than smaller plants to pollinators. A review of Tetratheca pollination ecology revealed that several species are poorly fecund and pollinators are rare. The habitat requirements for Tetratheca, a genus of many rare and threatened species, is discussed.     

Structural requirements of SLP-76 in signaling via the high-affinity immunoglobulin E receptor (Fc epsilon RI) in mast cells

The adapter SLP-76 plays an essential role in Fc epsilon RI signaling, since SLP-76(-/-) bone marrow-derived mast cells (BMMC) fail to degranulate and release interleukin-6 (IL-6) following Fc epsilon RI ligation. To define the role of SLP-76 domains and motifs in Fc epsilon RI signaling, SLP-76(-/-) BMMC were retrovirally transduced with SLP-76 and SLP-76 mutants. The SLP-76 N-terminal and Gads binding domains, but not the SH2 domain, were critical for Fc epsilon RI-mediated degranulation and IL-6 secretion, whereas all three domains are essential for T-cell proliferation following T-cell receptor (TCR) ligation. Unexpectedly, the three tyrosine residues in SLP-76 critical for TCR signaling, Y112, Y128, and Y145, were not essential for IL-6 secretion, but were required for degranulation and mitogen-activated protein kinase activation. Furthermore, a Y112/128F SLP-76 mutant, but not a Y145F mutant, strongly reconstituted mast cell degranulation, suggesting a critical role for Y145 in Fc epsilon RI-mediated exocytosis. These results point to important differences in the function of SLP-76 between T cells and mast cells.     

ABCA1 is essential for efficient basolateral cholesterol efflux during the absorption of dietary cholesterol in chickens

The ATP-binding cassette transporter A1 (ABCA1) participates in the efflux of cholesterol from cells. It remains unclear whether ABCA1 functions to efflux cholesterol across the basolateral or apical membrane of the intestine. We used a chicken model of ABCA1 dysfunction, the Wisconsin hypoalpha mutant (WHAM) chicken, to address this issue. After an oral gavage of radioactive cholesterol, the percentage appearing in the bloodstream was reduced by 79% in the WHAM chicken along with a 97% reduction in the amount of tracer in high density lipoprotein. In contrast, the percentage of radioactive cholesterol absorbed from the lumen into the intestine was not affected by the ABCA1 mutation. Liver X receptor (LXR) agonists have been inferred to decrease cholesterol absorption through activation of ABCA1 expression. However, the LXR agonist T0901317 decreased cholesterol absorption equally in both wild type and WHAM chickens, indicating that the effect of LXR activation on cholesterol absorption is independent of ABCA1. The ABCA1 mutation resulted in accumulation of radioactive cholesterol ester in the intestine and the liver of the WHAM chicken (5.0- and 4.4-fold, respectively), whereas biliary lipid concentrations were unaltered by the WHAM mutation. In summary, ABCA1 regulates the efflux of cholesterol from the basolateral but not apical membrane in the intestine and the liver.     

Effects of Pluronic F-68 on shoot regeneration from cultured jute cotyledons and on growth of transformed roots

The effects have been studied of the non-ionic surfactant, Pluronic F-68, on the growth in culture of jute (Corchorus capsularis L.) cotyledons with attached petioles, cotyledon explants and transformed roots. Supplementation of culture medium with 0.001–0.5% (w/v) of either commercial grade Pluronic F-68 or a purified fraction prepared by passage through silica gel, stimulated shoot production from the petioles of C. capsularis var. D154 and C134 cotyledons. This effect was most marked in C134, because of the failure of control cotyledons to produce shoots in the absence of Pluronic. Plants regenerated from Pluronic-treated cotyledons were morphologically normal. Growth of transformed roots of C. capsularis var. D154 was stimulated in medium supplemented with commercial grade or purified Pluronic F-68, with maximum increases in both fresh and dry weights with 0.1% (w/v) of the surfactant. Roots cultured in the presence of Pluronic F-68 could be maintained without sub-culture for up to 70 days, whereas roots cultured in the absence of Pluronic required subculture every 7 days, to prevent necrosis. Transformed roots also produced callus in the presence of 0.001–1.0% (w/v) of either commercial grade or purified Pluronic. The biotechnological implications of these results are discussed in relation to the potential value of non-ionic surfactants as growth-stimulating additives to plant culture media.Abbreviations NAA -naphthaleneacetic acid - BA 6-benzyladenine - IAA indole-3-acetic acid - MS Murashige & Skoog (1962)     

Utilization of exogenous linolenic and oleic acids for plasma membrane phosphoglyceride systhesis in L-Fibroblasts

The inability of the strain L-fibroblast to synthesize quatitatively significant amounts of polyenoic fatty acid and the apparent lack of turnover of their phosphoglyceride acyl groups under the usual conditions of cell culture makes them especially well suited for studies concerning the effect of fatty acid unsaturation on biological membranes. Such cells grown in the absence of exogenous lipid sources have in their phosphoglycerides only traces of polyenoic fatty acid.By infusing fatty acid supplements into suspension cultures of logarithmically growing cultures of L-fibroblasts it is possible to increase singnificantly their phosphoglyceride polyenoic fatty content to as much as 50% of the total lipid phosphoglyceride fatty acids. The infusion of fatty acid supplements at a constant rate over a 48 h time period diminishes the toxic effects which may accompany single doses of unesterified fatty acid and reduces considerably the accumulation of cytoplasmic lipid droplets. Cultures supplemented in this way have virtually the same generation times as non-supplemented control cultures. The data show that alterations in surface membrane and homogenate polyenoic fatty acid composition are minimal when oleic acid is supplied to the culture. During exposure to large amounts of polyenoic fatty acid, however, the unsaturation of plasma membrane total phosphoglyceride fraction is less than that of the cell homogenate. This effect is more pronounced in the phosphatidylethanolamine than in the phosphatidylcholine fraction.