Control of IFN-alphaA by CD73: implications for mucosal inflammation

Inflammatory diseases influence tissue metabolism, altering regulation of extracellular adenine nucleotides, with a resultant protective influence of adenosine. Ecto-5'-nucleotidase (CD73) is a central surface enzyme generating extracellular adenosine. Thus, we hypothesized that CD73 is protective in mucosal inflammation as modeled by trinitrobenzene sulfonate (TNBS) colitis. Initial studies revealed a >3-fold induction of CD73 mRNA levels after TNBS colitis. Additionally, the severity of colitis was increased, as determined by weight loss and colonic shortening, in cd73(-/-) mice relative to cd73(+/+) controls. Likewise, enteral administration of the selective CD73 inhibitor alpha,beta-methylene ADP to cd73(+/+) mice resulted in a similar increase in severity of TNBS colitis. Gene array profiling of cytokine mRNA expression, verified by real-time PCR, revealed a >90% down-regulation of IFN-alphaA in cd73(-/-) mice and alpha,beta-methylene ADP-treated cd73(+/+) mice, compared with cd73(+/+) mice. Exogenous administration of recombinant IFN-alphaA partially protected TNBS-treated cd73(-/-) mice. Cytokine profiling revealed similar increases in both IFN-gamma and TNF-alpha mRNA in colitic animals, independent of genotype. However, IL-10 mRNA increased in wild-type mice on day 3 after TNBS administration, whereas cd73(-/-) mice mounted no IL-10 response. This IL-10 response was restored in the cd73(-/-) mice by exogenous IFN-alphaA. Further cytokine profiling revealed that this IL-10 induction is preceded by a transient IFN-alphaA induction on day 2 after TNBS exposure. Together, these studies indicate a critical regulatory role for CD73-modulated IFNalphaA in the acute inflammatory phase of TNBS colitis, thereby implicating IFN-alphaA as a protective element of adenosine signaling during mucosal inflammation.     

Combining conventional QTL analysis and whole-exome capture-based bulk-segregant analysis provides new genetic insights into tuber sprout elongation and dormancy release in a diploid potato population

Tuber dormancy and sprouting are commercially important potato traits as long-term tuber storage is necessary to ensure year-round availability. Premature dormancy release and sprout growth in tubers during storage can result in a significant deterioration in product quality. In addition, the main chemical sprout suppressant chlorpropham has been withdrawn in Europe, necessitating alternative approaches for controlling sprouting. Breeding potato cultivars with longer dormancy and slower sprout growth is a desirable goal, although this must be tempered by the needs of the seed potato industry, where dormancy break and sprout vigour are required for rapid emergence. We have performed a detailed genetic analysis of tuber sprout growth using a diploid potato population derived from two highly heterozygous parents. A dual approach employing conventional QTL analysis allied to a combined bulk-segregant analysis (BSA) using a novel potato whole-exome capture (WEC) platform was evaluated. Tubers were assessed for sprout growth in storage at six time-points over two consecutive growing seasons. Genetic analysis revealed the presence of main QTL on five chromosomes, several of which were consistent across two growing seasons. In addition, phenotypic bulks displaying extreme sprout growth phenotypes were subjected to WEC sequencing for performing BSA. The combined BSA and WEC approach corroborated QTL locations and served to narrow the associated genomic regions, while also identifying new QTL for further investigation. Overall, our findings reveal a very complex genetic architecture for tuber sprouting and sprout growth, which has implications both for potato and other root, bulb and tuber crops where long-term storage is essential.Subject terms: Genetic markers, Next-generation sequencing, Plant breeding, Agricultural genetics, Genetic mapping     

Colony sound facilitates sexual and agonistic activities in royal penguins

A unique playback system was used to exaggerate continuously the number of calls heard in royal penguin, Eudyptes schlegeli, colonies to determine whether social facilitation increases a female's exposure to both sexual and agonistic activities, a process that could influence the timing of reproduction in colonies. We also determined whether the source of colony sound or the phase of the breeding cycle influenced the facilitation of sexual or agonistic activities. The system recorded and then played a new 12-min colony sound sample every 24 min throughout the first four phases of the breeding cycle. Because each broadcast was unique, this playback system avoids the possibility of habituation and can modify the acoustic environment in social groups for extended periods of time. The activities of penguins in control colonies (with no playback) were compared with those in colonies that heard samples derived from either their own or a different colony. Colony sound facilitated both sexual and agonistic activities from male and female royal penguins during all four phases of the cycle that we examined. Playbacks from the perceiver's own colony facilitated a greater response than sound originating from a different colony. Studies comparing egg-laying dates in colonies exposed continuously to the sounds of agonistic activities with those in control colonies, or colonies exposed continuously to the sounds of sexual activities are now needed to determine whether the facilitation of agonistic activities opposes the effects of social stimulation on the hormonal changes leading to ovulation in colony-dwelling birds. Copyright 2000 The Association for the Study of Animal Behaviour.     

Protein expression profiling of endometrial malignancies reveals a new tumor marker: chaperonin 10

Endometrial carcinoma is a common malignancy in women, being exceeded in incidence only by that of breast, lung, and colorectal cancers. At present, no serum tumor markers are available for the monitoring of endometrial carcinoma patients, and patients with recurrent disease are detected only following the development of symptoms or abnormalities in imaging assessments. Similarly, no screening tools are available for endometrial carcinoma. Protein profiling by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) has proven to be a sensitive and fast method of analysis for small proteins or peptides to yield specific biomarkers. In this study, a variety of normal and malignant endometrial tissue samples were fractionated and analyzed by SELDI-TOF MS (SELDI is a version of MALDI utilizing protein "chips"). A number of proteins displayed differential expression in malignant endometrial tissues. One of the prominent proteins fractionated by weak cation exchange chromatography and displaying enhanced expression in these malignant tissues was identified as chaperonin 10. The increased expression of chaperonin 10 in malignant endometrial tissues was further confirmed by parallel Western blot and immunohistochemistry analyses.     

An aspirin-triggered lipoxin A4 stable analog displays a unique topical anti-inflammatory profile

Lipoxins and 15-epi-lipoxins are counter-regulatory lipid mediators that modulate leukocyte trafficking and promote the resolution of inflammation. To assess the potential of lipoxins as novel anti-inflammatory agents, a stable 15-epi-lipoxin A(4) analog, 15-epi-16-p-fluorophenoxy-lipoxin A(4) methyl ester (ATLa), was synthesized by total organic synthesis and examined for efficacy relative to a potent leukotriene B(4) (LTB(4)) receptor antagonist (LTB(4)R-Ant) and the clinically used topical glucocorticoid methylprednisolone aceponate. In vitro, ATLa was 100-fold more potent than LTB(4)R-Ant for inhibiting neutrophil chemotaxis and trans-epithelial cell migration induced by fMLP, but was approximately 10-fold less potent than the LTB(4)R-Ant in blocking responses to LTB(4). A broad panel of cutaneous inflammation models that display pathological aspects of psoriasis, atopic dermatitis, and allergic contact dermatitis was used to directly compare the topical efficacy of ATLa with that of LTB(4)R-Ant and methylprednisolone aceponate. ATLa was efficacious in all models tested: LTB(4)/Iloprost-, calcium ionophore-, croton oil-, and mezerein-induced inflammation and trimellitic anhydride-induced allergic delayed-type hypersensitivity. ATLa was efficacious in mouse and guinea pig skin inflammation models, exhibiting dose-dependent effects on edema, neutrophil or eosinophil infiltration, and epidermal hyperproliferation. We conclude that the LXA(4) and aspirin-triggered LXA(4) pathways play key anti-inflammatory roles in vivo. Moreover, these results suggest that ATLa and related LXA(4) analogs may have broad therapeutic potential in inflammatory disorders and could provide an alternative to corticosteroids in certain clinical settings.     

Genomic affinities in Arachis section Arachis (Fabaceae): molecular and cytogenetic evidence

Section Arachis is the largest of nine sections in the genus Arachis and includes domesticated peanut, A. hypogaea L. Most species are diploids (x=10) with two tetraploids and a few aneuploids. Three genome types have been recognized in this section (A, B and D), but the genomes are not well characterized and relationships of several newly described species are uncertain. To clarify genomic relationships in section Arachis, cytogenetic information and molecular data from amplified fragment length polymorphism (AFLP) and the trnT-F plastid region were used to provide an additional insight into genome composition and species relationships. Cytogenetic information supports earlier observations on genome types of A. cruziana, A. herzogii, A. kempff-mercadoi and A. kuhlmannii but was inconclusive about the genome composition of A. benensis, A. hoehnei, A. ipaensis, A. palustris, A. praecox and A. williamsii. An AFLP dendrogram resolved species into four major clusters and showed A. hypogaea grouping closely with A. ipaensis and A. williamsii. Sequence data of the trnT-F region provided genome-specific information and showed for the first time that the B and D genomes are more closely related to each other than to the A genome. Integration of information from cytogenetics and biparentally and maternally inherited genomic regions show promise in understanding genome types and relationships in Arachis.     

CD1d-dependent macrophage-mediated clearance of Pseudomonas aeruginosa from lung

CD1d-restricted T cells are implicated as key players in host defense against various microbial infections. However, the mechanisms involved and the role they play, if any, at the mucosal surfaces where pathogenic infections are initiated is unknown. In a murine pneumonia model established by intranasal application of Pseudomonas aeruginosa, CD1d(-/-) mice showed markedly reduced pulmonary eradication of P. aeruginosa compared with wild-type mice; this was associated with significantly lower amounts of macrophage inflammatory protein-2 and reduced numbers of neutrophils within the bronchoalveolar lavage fluid. Corollarily, treatment of mice with alpha-galactosylceramide--a lipid that activates CD1d-restricted T cells--increased the amount of interferon-gamma; this was associated with rapid pulmonary clearance through enhanced phagocytosis of P. aeruginosa by alveolar macrophages. These results reveal a crucial role played by CD1d-restricted T cells in regulating the antimicrobial immune functions of macrophages at the lung mucosal surface.     

Two groups control light-induced schiff base deprotonation and the proton affinity of asp(85) in the Arg(82)His mutant of bacteriorhodopsin

Arg(82) is one of the four buried charged residues in the retinal binding pocket of bacteriorhodopsin (bR). Previous studies show that Arg(82) controls the pK(a)s of Asp(85) and the proton release group and is essential for fast light-induced proton release. To further investigate the role of Arg(82) in light-induced proton pumping, we replaced Arg(82) with histidine and studied the resulting pigment and its photochemical properties. The main pK(a) of the purple-to-blue transition (pK(a) of Asp(85)) is unusually low in R82H: 1.0 versus 2.6 in wild type (WT). At pH 3, the pigment is purple and shows light and dark adaptation, but almost no light-induced Schiff base deprotonation (formation of the M intermediate) is observed. As the pH is increased from 3 to 7 the M yield increases with pK(a) 4.5 to a value approximately 40% of that in the WT. A transition with a similar pK(a) is observed in the pH dependence of the rate constant of dark adaptation, k(da). These data can be explained, assuming that some group deprotonates with pK(a) 4.5, causing an increase in the pK(a) of Asp(85) and thus affecting k(da) and the yield of M. As the pH is increased from 7 to 10.5 there is a further 2.5-fold increase in the yield of M and a decrease in its rise time from 200 &mgr;s to 75 &mgr;s with pK(a) 9. 4. The chromophore absorption band undergoes a 4-nm red shift with a similar pK(a). We assume that at high pH, the proton release group deprotonates in the unphotolyzed pigment, causing a transformation of the pigment into a red-shifted "alkaline" form which has a faster rate of light-induced Schiff base deprotonation. The pH dependence of proton release shows that coupling between Asp(85) and the proton release group is weakened in R82H. The pK(a) of the proton release group in M is 7.2 (versus 5.8 in the WT). At pH < 7, most of the proton release occurs during O --> bR transition with tau approximately 45 ms. This transition is slowed in R82H, indicating that Arg(82) is important for the proton transfer from Asp(85) to the proton release group. A model describing the interaction of Asp(85) with two ionizable residues is proposed to describe the pH dependence of light-induced Schiff base deprotonation and proton release.