Pollination and stigma wounding: same response, different signal?

In Petunia hybrida flowers, both pollination and stigma woundinginduced a transient Increase in ethylene production and hastenedcorolla senescence. Ethylene production by different flowerparts was measured in situ using laser photoacoustic (LPA) spectroscopy.In pollinated flowers, ethylene was exclusively produced bythe stigma/style region whereas wounding of the stigma Inducedethylene production both by the stigma/style region and by theremaining flower parts. In aminoethoxyvinylglycine (AVG)-treatedflowers, subsequent treatment of the unwounded stigma with 1-aminocyclopropane-1-carboxylicacid (ACC) induced ethylene production exclusively by the stigma/styleregion whereas treatment of a previously wounded stigma withACC induced a simultaneous increase in ethylene production bythe stigma/style region and the remaining flower parts. Theseresults suggest that following stigma wounding, either ACC orethylene is involved in inter-organ communication. Followingpollination, the signal is apparently not directly related toethylene. In vivo ACC oxidase activity of most flower parts, includingthe gynoecium, was higher in light than in dark. Light or darkdid not influence the relative contributions of stigma/styleand remaining flower parts to the total pollination, woundingor ACC-induced ethylene production, indicating that ACC is nottranslocated. Both in excised styles and intact flowers, radiolabelledACC and its analogue -aminoisobutyric acid (AIB), applied eitherto an intact or wounded stigma, were largely immobile confirmingthat ACC is not likely to play a role in inter-organ signalling. The results collectively suggest that following stigma wounding,translocation of ethylene may be the signal responsible forinitiation of corolla senescence; following pollination thesignal is not directly related to ethylene. Key words: 1-Aminocyclopropane-1-carboxylic acid (ACC), ethylene, flower senescence, Petunia hybrida, pollination, stigma wounding     

Population genetic structure of the South American species Hypochaeris lutea (Asteraceae)

The genus Hypochaeris has a recent evolutionary history caused by long‐distance dispersal in conjunction with adaptive radiation in the South American continent. Hypochaeris lutea is a perennial herb that grows mostly at altitudes of around 1000 m in cold swamps of the southern regions of Brazil. We investigated the amplified fragment length polymorphism (AFLP) in 270 individuals representing 11 Brazilian populations of H. lutea to elucidate the population genetic structure of this species. The frequencies of polymorphic loci and gene diversity ranged from 83.42% to 91.66% and from 0.26 to 0.34, respectively. Analysis of molecular variance revealed that most of the genetic variability was found within (76.67%) rather than among (23.3%) populations, agreeing with the pattern of genetic distribution within and among populations observed in other allogamous species of Hypochaeris. A Mantel test showed no correlation between genetic and geographic distances when all populations were considered. Simulations performed using a Bayesian approach consistently identified two clusters with different admixture proportions of individuals, as also revealed by a UPGMA dendrogram of populations. The pattern of genetic structure observed in H. lutea is consistent with a process of successive colonization events by long‐distance dispersal resembling the rapid and recent radiation that has been proposed to explain the origin of the South American species of Hypochaeris.     

Quantitative structure-activity relationships of estrogenic steroids substituted at C14, C15

The uterotropic activity of thirty 3-methoxyestradiol derivatives is measured and discussed on the basis of X-ray crystallographic results and quantitative structure-activity relationship analyses involving hydrophobic substituent constants pi and f as well as steric parameters Pr and L. In addition, estrogenicity is compared to data of interceptive activity and receptor binding affinity. All the biological data exhibit a high degree of intercorrelation. 17 beta-Hydroxysteroids having 14 alpha configuration reveal a generally better capability of high-affinity binding than those being 14 beta configurated. Between the uterotropic activity and the hydrophobicity of C14, C15 substituents, statistically significant correlations are found which suggest a close contact between the steroidal D-ring subsite and the receptor protein (e.g. for 14 alpha steroids: log UDD = -0.996 pi -0.392; n = 9, r = -0.943, s = 0.235, t = -7.5, alpha less than 0.001). The hydrophobic nature of both 14 alpha and 14 beta medium-sized substituents employed is shown by QSAR regressions to exert a stronger influence than steric effects. Furthermore, there are indications to additional hydrogen bonding and steric repulsion phenomena. As to the receptor-binding models discussed in the literature, it is concluded that the receptor protein has a high conformational flexibility to accommodate very different drug structures all having the common phenolic ring A. But, if an appropriate spacing of steroidal key atoms is recognized by the receptor and, consequently, the steroid-receptor complex is formed, the binding is complemented by hydrophobic interactions also in the D-ring region.     

Increased hepatobiliary and fecal cholesterol excretion upon activation of the liver X receptor is independent of ABCA1

The ATP-binding cassette transporter ABCA1 is essential for high density lipoprotein (HDL) formation and considered rate-controlling for reverse cholesterol transport. Expression of the Abca1 gene is under control of the liver X receptor (LXR). We have evaluated effects of LXR activation by the synthetic agonist T0901317 on hepatic and intestinal cholesterol metabolism in C57BL/6J and DBA/1 wild-type mice and in ABCA1-deficient DBA/1 mice. In wild-type mice, T0901317 increased expression of Abca1 in liver and intestine, which was associated with an approximately 60% rise in HDL. Biliary cholesterol excretion rose 2.7-fold upon treatment, and fecal neutral sterol output was increased by 150-300%. Plasma cholesterol levels also increased in treated Abca1(-/-) mice (+120%), but exclusively in very low density lipoprotein-sized fractions. Despite the absence of HDL, hepatobiliary cholesterol output was stimulated upon LXR activation in Abca1(-/-) mice, leading to a 250% increase in the biliary cholesterol/phospholipid ratio. Most importantly, fecal neutral sterol loss was induced to a similar extent (+300%) by the LXR agonist in DBA/1 wild-type and Abca1(-/-) mice. Expression of Abcg5 and Abcg8, recently implicated in biliary excretion of cholesterol and its intestinal absorption, was induced in T0901317-treated mice. Thus, activation of LXR in mice leads to enhanced hepatobiliary cholesterol secretion and fecal neutral sterol loss independent of (ABCA1-mediated) elevation of HDL and the presence of ABCA1 in liver and intestine.     

A model for signaling specificity of Wnt/Frizzled combinations through co-receptor recruitment

Wnts control mammalian developmental morphogenesis and are critical for adult stem cell maintenance. Wnts initiate several intracellular signaling cascades, such as Wnt/β-catenin-, Wnt/Ca²⁺- and Wnt/ROR2-signaling. Signaling preference of Wnts for these various pathways is thought to depend on the repertoire of receptors present on recipient cells. Here, we propose a further refinement of this receptor model and hypothesize that Wnt signaling specificity depends on co-receptor recruitment upon binding of Wnt to Frizzled receptor molecules. In this model, recruitment of LRP5/6 leads to activation of Wnt/β-catenin signaling, whereas signaling through other pathways is mediated by recruiting ROR2.     

Discovery of novel small molecule activators of β-catenin signaling

Wnt/β-catenin signaling plays a major role in embryonic development and adult stem cell maintenance. Reduced activation of the Wnt/β-catenin pathway underlies neurodegenerative disorders and aberrations in bone formation. Screening of a small molecule compound library with a β-galactosidase fragment complementation assay measuring β-catenin nuclear entry revealed bona fide activators of β-catenin signaling. The compounds stabilized cytoplasmic β-catenin and activated β-catenin-dependent reporter gene activity. Although the mechanism through which the compounds activate β-catenin signaling has yet to be determined, several key regulators of Wnt/β-catenin signaling, including glycogen synthase kinase 3 and Frizzled receptors, were excluded as the molecular target. The compounds displayed remarkable selectivity, as they only induced β-catenin signaling in a human osteosarcoma U2OS cell line and not in a variety of other cell lines examined. Our data indicate that differences in cellular Wnt/β-catenin signaling machinery can be exploited to identify cell type-specific activators of Wnt/β-catenin signaling.     

Analysis of alcohol dependence phenotype in the COGA families using covariates to detect linkage

Linkage analysis methods that incorporate etiological heterogeneity of complex diseases are likely to demonstrate greater power than traditional linkage analysis methods. Several such methods use covariates to discriminate between linked and unlinked pedigrees with respect to a certain disease locus. Here we apply several such methods including two mixture models, ordered subset analysis, and a conditional logistic model to genome scan data on the DSM-IV alcohol dependence phenotype on the Collaborative Studies on Genetics of Alcoholism families, and compare the results to traditional nonparametric linkage analysis. In general, there was little agreement among the various covariate-based linkage statistics. Linkage signals with empirical p-values less than 0.001 were detected on chromosomes 3, 4, 7, 10, and 12, with the highest peak occurring at the GABRB1 gene using the ecb21 covariate.     

Abcg5/Abcg8-independent pathways contribute to hepatobiliary cholesterol secretion in mice

The ATP-binding cassette (ABC) half-transporters ABCG5 and ABCG8 heterodimerize into a functional complex that mediates the secretion of plant sterols and cholesterol by hepatocytes into bile and their apical efflux from enterocytes. We addressed the putative rate-controlling role of Abcg5/Abcg8 in hepatobiliary cholesterol excretion in mice during (maximal) stimulation of this process. Despite similar bile salt (BS) excretion rates, basal total sterol and phospholipid (PL) output rates were reduced by 82% and 35%, respectively, in chow-fed Abcg5(-/-) mice compared with wild-type mice. When mice were infused with the hydrophilic BS tauroursodeoxycholate, similar relative increases in bile flow, BS output, PL output, and total sterol output were observed in wild-type, Abcg5(+/-), and Abcg5(-/-) mice. Maximal cholesterol and PL output rates in Abcg5(-/-) mice were only 15% and 69%, respectively, of wild-type values. An infusion of increasing amounts of the hydrophobic BS taurodeoxycholate increased cholesterol excretion by 3.0- and 2.4-fold in wild-type and Abcg5(-/-) mice but rapidly induced cholestasis in Abcg5(-/-) mice. Treatment with the liver X receptor (LXR) agonist T0901317 increased the maximal sterol excretion capacity in wild-type mice (fourfold), concomitant with the induction of Abcg5/Abcg8 expression, but not in Abcg5(-/-) mice. In a separate study, mice were fed chow containing 1% (wt/wt) cholesterol. As expected, hepatic expression of Abcg5 and Abcg8 was strongly induced (fivefold and fourfold) in wild-type but not LXR-alpha-deficient (Lxra(-/-)) mice. Surprisingly, hepatobiliary cholesterol excretion was increased to the same extent, i.e., 2.2-fold in wild-type mice and 2.0-fold in Lxra(-/-) mice, upon cholesterol feeding. Our data confirm that Abcg5, as part of the Abcg5/Abcg8 heterodimer, strongly controls hepatobiliary cholesterol secretion in mice. However, our data demonstrate that Abcg5/Abcg8 heterodimer-independent, inducible routes exist that can significantly contribute to total hepatobiliary cholesterol output.     

Phase transitions and permeability changes in dry membranes during rehydration

Dry phospholipid bilayers are known to undergo transient changes in permeability during rehydration. In this review, we present evidence from which we suggest that this permeability change is due to a gel to liquid-crystaline phase transition accompanying rehydration. If the transition is avoided, as in lipids that remain in gel phase whether dry or rehydrated, the problem of leakage during rehydration is obviated, at least in part. Further, the evidence that the transition temperature for dry bilayers can be depressed by certain sugars is discussed. Finally, we show that these principles can be extended to intact cells. Using pollen grains as a model, we have measured the transition temperature for membrane phospholipids and show that the transition is correlated with physiological measurements including permeability changes and subsequent germination. From theT _m values taken from pollen grains at different water contents, we have constructed a phase diagram for the intact pollen that has high predictive value for physiological properties.     

Properties of proteins and the glassy matrix in maturation-defective mutant seeds of Arabidopsis thaliana

In situ Fourier transform infrared microspectroscopy was used to study the heat stability of proteins and hydrogen bonding interactions in dry maturation-defective mutant seeds of Arabidopsis thaliana . α-Helical, turn and β-sheet conformations were the major protein secondary structures in all of these seeds. On heating, intermolecular extended β-sheet structures, typical of protein denaturation, were formed in abscisic acid-insensitive ( abi3 ) and leafy cotyledon ( lec ) mutant seeds. Proteins in dry wild-type seeds did not denature up to 150°C, but those in dry desiccation-sensitive, lec1–1 , lec1–3 and abi3–5 seeds did at 68, 89 and 87°C, respectively. In the desiccation-tolerant abi3–7 and abi3–1 seeds, denaturation commenced above 120 and 135°C, respectively. Seeds of the aba1–1 abi3–1 double mutant showed signs of denaturation already upon drying. The molecular packing in the seeds was studied by observing the shift in the position of the OH-stretching vibration band with temperature. The maximal rate of change of this band with temperature was much higher in the desiccation-sensitive abi3–5 , aba1–1 abi3–1 , lec1–1 , and lec1–3 mutant seeds than in the desiccation-tolerant wild-type, abi3–1 , abi3–7 , and lec2–1 seeds. We interpret this to mean that the molecular packing density is higher in dry desiccation-tolerant than in dry desiccation-sensitive seeds, which is associated with a higher or lower protein denaturation temperature, respectively. The results are discussed in relation to the physiological and biochemical characteristics of these mutant seeds.