Correlation of defense gene induction defects with powdery mildew susceptibility in Arabidopsis enhanced disease susceptibility mutants

We investigated the relative importance of specific Arabidopsis thaliana genes in conferring resistance to bacterial versus fungal pathogens. We first developed a pathosystem involving the infection of Arabidopsis accession Columbia with a virulent isolate of the obligate biotrophic fungal pathogen Erysiphe orontii. E. orontii elicited the accumulation of mRNAs corresponding to the defense-related genes PR1, BGL2 (PR2), PR5 and GST1 , but did not elicit production of the phytoalexin camalexin or the accumulation of defensin ( PDF1.2 ) or thionin ( THI2.1 ) mRNAs. We tested a set of 15 previously isolated Arabidopsis phytoalexin deficient (pad), non-expresser of PR (npr) and enhanced disease susceptibility (eds) mutants that are more susceptible to Pseudomonas syringae for their susceptibility to E. orontii. Four of these mutants ( pad4–1, npr1–1, eds5–1 and a double npr1–1 eds5–1 mutant) as well as Arabidopsis lines carrying a nahG transgene exhibited enhanced susceptibility to E. orontii and reduced levels of PR gene expression . Comparison of the PR gene induction patterns in response to E. orontii in the various mutants and in the nahG transgenics suggests the existence of NPR1 -independent salicylate-dependent and NPR1 -independent salicylate-independent defense gene activation pathways. Eleven other eds and pad mutants did not show measurable enhanced susceptibility to E. orontii , suggesting that these mutants are defective in factors that are not important for the limitation of E. orontii growth.     

WAITING FOR SPECIATION: THE EFFECT OF POPULATION SUBDIVISION ON THE TIME TO SPECIATION

We study the time required for speciation in a species that is divided into small versus large populations. Following Dobzhansky and Muller, we assume that hybrid sterility or inviability is caused by “complementary genes,” that is, by the accumulation of genes that cause sterility or inviability when brought together in hybrids but that have no deleterious effect on their normal species genetic background. When divergence between populations is caused by genetic drift, we show that the time to speciation is independent of population subdivision: speciation occurs just as quickly in a species split into a few large populations as into many small populations. When divergence is driven by natural selection, however, the time to speciation is very sensitive to population subdivision and speciation occurs most rapidly when a species is split into two large populations. These results contradict several popular intuitions about the effect of population size on speciation.     

Neuronal Release of Serotonin in the Cerebellum of Behaving Rats: An In Vivo Microdialysis Study

Abstract: Release of endogenous serotonin [5-hydroxy-tryptamine (5-HT)] in the cerebellum of awake rats was characterized using in vivo microdialysis. 5-HT output was increased (∼70%) by local application of KCl (100 m M ) and was reduced (∼60%) by both tetrodotoxin (0.5 µ M ) and omission of Ca²⁺ from the perfusion fluid. 5-HT release was decreased (∼70%) by the selective 5-HT_1A agonist 8-hydroxy-2-(di- n -propylamino)tetralin (0.25 mg/kg, s.c.), and this effect was rapidly reversed by a selective 5-HT_1A antagonist, N -[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]- N -(2-pyridinyl)cyclohexane-carboxamide trihydrochloride (WAY-100635; 0.1 mg/kg, i.p.). These results indicate that a large portion of the measurable 5-HT output in the cerebellum is of neuronal origin, is dependent on impulse flow, and is sensitive to 5-HT_1A autoreceptor activation. Further studies examined the relationship between 5-HT levels and general activity of the animals across the light-dark transition and during behavioral manipulations. Both 5-HT levels and behavioral activity were significantly elevated during the dark period, with changes in 5-HT efflux closely paralleling changes in activity. Similar increases (∼40%) in 5-HT output were observed during both feeding and feeding in the presence of a stressor (tail pinch). These findings suggest that behavioral state is an important factor determining neuronal 5-HT release in cerebellum under physiological conditions.     

Regulation of GTP Cyclohydrolase I Gene Expression and Tetrahydrobiopterin Content in Cultured Sympathetic Neurons by Leukemia Inhibitory Factor and Ciliary Neurotrophic Factor

Abstract: Cultures of neonatal rat superior cervical ganglia (SCG) were used to test the hypothesis that the cytokines leukemia inhibitory factor (LIF) and ciliary neurotrophic factor (CNTF) control GTP cyclohydrolase I (GTPCH) gene expression and 5,6,7,8-tetrahydrobiopterin (BH4) content as traits of the noradrenergic phenotype. Treatment for 7 days with 1 ng/ml of LIF was found to produce the characteristic switch in the SCG neurotransmitter phenotype reported by others, as evidenced by a 60% decline in tyrosine hydroxylase (TH) activity and a 75% increase in choline acetyltransferase activity. This LIF treatment paradigm decreased BH4 levels in a concentration-dependent manner, with a maximal decline of 60% observed at 1 ng/ml. Analysis of the time course of this response indicated that LIF decreased BH4 levels by 60% following 3–7 days of treatment. Treatment of cultures with CNTF (2 ng/ml) resulted in a decline in BH4 levels that was of equal magnitude and followed the same time course as that produced by LIF. The LIF-dependent decline in BH4 levels resulted from a reduction in GTPCH enzyme activity, which decreased by 75% following 7 days of treatment. Nuclease protection assays of RNA extracted from cells treated for 7 days with 2 ng/ml of LIF or CNTF detected a 78–96% reduction in GTPCH mRNA content relative to β-actin mRNA content. Concomitant decreases in TH and GTPCH gene expression in response to LIF or CNTF demonstrate a coordinated regulation of gene expression for this BH4-dependent enzyme and the rate-limiting enzyme in the synthesis of its essential cofactor, BH4. Moreover, these results indicate that GTPCH gene expression in SCG neurons should be regarded as a trait of the noradrenergic phenotype.     

Dura mater secretes soluble heparin-binding factors required for cranial suture morphogenesis

Summary Cranial sutures play a critical role in calvarial morphogenesis, serving as bone growth centers during skull enlargement. Defective suture morphogenesis, resulting in premature osseous obliteration of sutures and their failure to function appropriately, causes severe craniofacial anomalies. Previously published data demonstrated osseous obliteration of coronal suturesin vitro in the absence of dura mater and the rescue of sutures from osseous obliteration in rudiments cocultured with dura mater on the opposite sides of 0.45-μm polycarbonate filters. With thisin vitro culture system, experiments were designed to examine the nature of the soluble signal secreted by dura mater, required for maintaining intact sutures. The signal remained active in conditioned medium produced from dura mater, which was capable of rescuing coronal sutures from osseous obliteration in calvaria cultured without dura mater. When conditioned medium was segregated into heparin-binding and non-heparin-binding fractions, the signal capable of maintaining intact coronal sutures cosegregated with the heparin-binding component and remained functional in the absence of the non-heparin-binding component of conditioned medium. Evidence indicates that soluble, heparin-binding factors secreted by the dura mater act as osteoinhibitory signals at the suture site.     

Derivatives of the red-violet cluster of copper and penicillamine prepared by mixed ligand formation reactions or direct additions

Mixed ligand clusters of the type Cu(I)₈Cu(II)₆L'_(n)L_(12?n)Cl are easily distinguished by zone electrophoresis if ligands L and L' differ in formal charge. Approximately random species distributions were observed when L was penicillamine and L' a related ligand lacking the negative charge of a carboxylate group, provided the L'/L molar ratio was less than $\text{1}\text{9}$ in the reaction mixture. Depending on the choice of L or L', it should be possible to prepare clusters with a variety of pendant groups. The cluster containing only penicillamine apparently forms addition products with carbodiimides and aziridines based on reactions involving the cluster's peripheral carboxylate groups.     

Athletes'' pseudoanaemia

To characterize the so-called pseudoanaemia of endurance-trained athletes, the plasma volume (PV), red cell volume (RCV) and total blood volume (TBV) of 12 male and 12 female athletes and 5 male and 5 female nonexercising controls were measured using 125I-labelled human serum albumin and 51Cr-labelled erythrocytes. The mean PV of the male athletes (52.8 ml.kg-1) was 37.5% higher than that of the controls (38.4 ml.kg-1), while the 18.1% increase measured in the female runners (51.5 ml.kg-1) over the controls (43.6 ml.kg-1) was a novel observation. Although the RCV was significantly greater (34.7%) in male athletes (32.6 ml.kg-1 vs 24.2 ml.kg-1 in the controls), a similar elevation (3.6%) was not found in the female athletes (25.9 ml.kg-1) compared to the sedentary women (22.8 ml.kg-1). This could have been due to iron-limited erythropoiesis because the RCV of the female athletes defined as clinically anaemic was markedly lower that of the nonanaemic women (P less than 0.05). The elevated plasma protein mass and concentration measured in the athletes partly accounted for their expanded PV. It was concluded that the decreased blood haemoglobin levels reported in the endurance athletes was largely a dilutional effect.     

Replication asynchrony between homologs 15q11.2: Cytogenetic evidence for genomic imprinting

Summary Replication kinetics of the Prader-Willi syndrome critical region (15q11.2) was investigated in seven normal healthy adult females using RBG replication bands. Replication asynchrony between homologs 15q11.2 was identified consistently in about 40% of cells in all individuals. It was limited to the stages in which Xp22, Xp11, Xq13 and Xq24/26 were visible in the late-replicating X chromosome. This asynchrony suggested that replication timing overlapped between 15q11.2 and the early replicating R-bands of the late X chromosome in some cells, and that the difference in replication timing between homologs was probably related to genomic imprinting; the latter has been suggested as a pathogenetic basis of Prader-Willi syndrome. As a result of an analysis of the proportions of asynchronous and synchronous cells in each replication stage, two types of cells were deduced providing 11 methylation mosaicism of genomic imprinting was assumed. The first type was composed of cells with normal replication in one homolog and delayed replication in the other. The second type was composed of cells with normal replication in both homologs. Our results provide cytogenetic evidence of methylation mosaicism for mammalian genomic imprinting.     

Involvement of IL-6 in mesangial proliferative glomerulonephritis

In this study, we demonstrated that IL-6 was a possible autocrine growth factor for rat mesangial cells (MC). rIL-6 induced in vitro growth of rat MC at a concentration of 2 to 200 ng/ml and IL-6 activity was found in the supernatant of cultured rat MC. Northern blot analysis as well as in situ hybridization revealed that IL-6 mRNA was expressed in the cultured MC. Of urine samples from patients with mesangial proliferative glomerulonephritis (PGN) 50% were found to contain significant IL-6 activity (ranging from 30 to 126 pg/ml). Urine samples from other type of primary glomerular diseases such as minimal change nephrotic syndrome or healthy volunteers contained no detectable IL-6 activity. Only 2 of 27 urine samples from membranous nephropathy contained detectable amount of IL-6. Furthermore, there was some relationship between the levels of urine IL-6 and the progressive stage of PGN. Finally, by immunohistochemical staining using an anti-IL-6 mAb, it was shown that MC in the affected glomeruli of PGN patients produced IL-6, whereas MC obtained from the patients with membranous nephropathy, minimal change nephrotic syndrome or normal kidney were not found to produce IL-6. These data suggest that deregulated production of IL-6 is involved in PGN and the measurement of urine IL-6 is helpful for the differential diagnosis of PGN as well as for monitoring the progression of PGN.     

Phenazine derivatives as the mutagenic reaction product from o- or m-phenylenediamine derivatives with hydrogen peroxide

8 Kinds of o- and m-phenylenediamine (PD) derivatives, which are used as oxidative-type hair dyes, were treated with hydrogen peroxide (H2O2). Both before and after H2O2 treatment, their mutagenicity was tested by using Salmonella typhimurium TA98 in the presence or absence of a mammalian metabolic activation system (S9 mix). After H2O2 treatment, the mutagenic potencies of p-nitro-o-phenylenediamine, 3,4-diaminotoluene, p-nitro-m-phenylenediamine and 2,4-diaminophenol did not vary or slightly increased in comparison with those of the starting materials. The mutagenicity of o-PD, p-chloro-o-phenylenediamine (p-Cl-o-PD), m-PD and 2,4-diaminoanisole (p-OMe-m-PD) was enhanced remarkably by treatment with H2O2 and all the oxidation products required metabolic activation by S9 mix for their mutagenesis. In a gas chromatography/mass spectrometric study, 2,3-diaminophenazine and 2,7-diaminophenazine were identified with authentic samples in o-PD and m-PD oxidation mixture, respectively. The oxidation mixture obtained from p-Cl-o-PD and p-OMe-m-PD was separated into several fractions by repeated column chromatography. Brownish yellow crystals were isolated from oxidized p-Cl-o-PD and the structure of the compound was determined to be 2,3-diamino-7-chlorophenazine from physicochemical and chemical evidence. Two reddish yellow crystals, obtained from oxidized p-OMe-m-PD, were 2,7-diamino-3,8-dimethoxyphenazine and 2,7-diamino-3-methoxyphenazine. The number of revertants induced by 1 nmole of phenazines detected from oxidized PD derivatives was as follows; 2,3-diaminophenazine: 349 rev.; 2,3-diamino-7-chlorophenazine; 406 rev.: 2,7-diaminophenazine: 12 110 rev.; 2,7-diamino-3,8-dimethoxyphenazine: 4229 rev.; 2,7-diamino-3-methoxyphenazine: 24 640 rev. in S. typhimurium TA98 strain with 25 microliters S9 per plate.