Production and composition of phenylpyrrole metabolites prodcued by Pseudomonas cepacia

Summary In shaken cultures, a strain of Pseudomonas cepacia isolated from apple leaves produced pyrrolnitrin and four other phenylpyrrole antibiotics. The concentrations of these metabolites were determined at intervals for 7 days in three different media at two initial pH levels. Optical density measurements revealed maximum cell concentrations after 24 h in nutrient broth, after 48 h in King's B medium, and after 96 h in minimum salts solution. The effects caused by initiating fermentations at pH 5.8 rather than 7.0 were in most cases not dramatic, although in some instances, especially in minimum salts broth, higher concentrations of metabolites were produced with the lower initial pH. Concentrations of the phenylpyrrole antibiotics were greatly affected by choice of culture medium and incubation time. Concentrations of the two nitrophenyl metabolites, pyrrolnitrin and 2-chloropyrrolnitrin, rose throughout the 7-day incubation and were more than 20 times greater in minimum salts medium than in either King's B medium or nutrient broth. The maximum concentrations of each of the three aminophenyl metabolites (dichloroamino, trichloroamino and monochloroamino) occurred in different media, the monochloro compound in nutrient broth, the dichloro compound in Kings B medium and the trichloro compound in minimum salts medium. The time dependence of the concentrations of the five metabolites supports the proposed biosynthesis of these pyrroles from tryptophan by successive chlorinations followed by oxidation of the amino group at the end of the pathway.     

The Roles of CRE, TRE, and TRE-Adjacent S1 Nuclease Sensitive Element in the Regulation of Tyrosine Hydroxylase Gene Promoter Activity by Angiotensin II

Abstract: The cis elements mediating activation of the tyrosine hydroxylase gene by angiotensin II were examined by transfecting tyrosine hydroxylase promoter-luciferase constructs into cultured bovine adrenal medullary cells. Angiotensin II-responsive elements are located within −54/+25-bp and −269/−55-bp promoter regions and were identified, respectively, as cyclic AMP (CRE)- and 12- O -tetradecanoylphorbol 13-acetate responsive element (TRE)-like sequences. Unlike CRE, TRE also supports basal promoter activity. Mutations of TRE or CRE that reduced angiotensin II stimulation abolished in vitro binding of nuclear proteins to those elements, suggesting that proteins forming CRE- and TRE-inducible complexes may mediate angiotensin II stimulation. The TRE is adjacent to a dyad symmetry element. Those two sites form a common regulatory unit in which the dyad symmetry element acts as a repressor of the TRE site. Isolated dyad symmetry element did not bind nuclear proteins in vitro. In supercoiled DNA it exhibited S1 nuclease sensitivity and was recognized by a DNA cruciform-specific antibody consistent with the extrusion of a cruciform structure that overlaps with the TRE. A mutation that abolished formation of the cruciform correlated with a loss of repressor activity. We propose a novel model of tyrosine hydroxylase gene regulation in which functions of the TRE are modulated via structural transition in the adjacent DNA.     

Alfalfa Nuclei Contain Cold-Responsive Phosphoproteins and Accumulate Heat-Stable Proteins during Cold Treatment of Seedlings

We have examined whether low temperature, as a pervasive thermodynamicstimulus, is sensed independently in different parts of thecell by studying low temperature responses of phosphoproteinsin isolated nuclei. The isolated alfalfa (Medicago saliva) nucleirespond to cold by rapid and reversible changes in phosphorylationlevel of their proteins. The population of such cold-regulatedphosphoproteins and the cold-stimulation of their phosphorylationare greater in a freezing-tolerant cultivar Apica than in asensitive cultivar Trek. With a 4-day cold treatment of theseedlings, additional proteins showing cold-stimulated phosphorylationappear in the nucleus of Apica while there is little changein the case of Trek. Furthermore, nuclei from cold-treated seedlingsof Apica, but not of Trek, show a large accumulation of heat-stableproteins. These results support the view that the low temperaturesensing and acclimation occur in all vital parts of the celland that accumulation of heat-stable nuclear proteins may berelated to freezing tolerance. (Received October 14, 1996; Accepted November 11, 1996)     

Overall Informativity,OI, in DNA Polymorphisms Revealed by inter-AluPCR: Detection of Genomic Rearrangements

We studied two systems of multilocus markers revealed by PCR using primers directing amplification betweenAlurepeats in a tail-to-tail orientation. Genomic polymorphisms were detected as the presence or absence of the electrophoretic bands representing DNA fragments of a given length. A total of 104 such fragments segregating as Mendelian markers in a panel of eight CEPH families were analyzed by two-point linkage analysis. Fifty-one of these fragments were localized with respect to CEPH markers; they represented 33 loci, 7 of which were multiallelic. Locus-specific oligonucleotides were developed and used as hybridization probes to identify the mapped loci within a complex pattern of inter-AluPCR products. A great proportion of inter-AluPCR polymorphisms represented length variants within amplified DNA segments, while others were presumably due to mutations within the priming sites. To describe the expected number of informative loci per typing experiment we introduced a parameter called overall informativity (OI), which provides a single measure of the multiplex ratio and the informativity of markers contributing to a multilocus system (OIof a single locus is equivalent to its heterozygosity and cannot exceed 0.5 for a biallelic codominant marker). HighOIvalues (5.8 and 11.5) of the two presented systems of inter-AluPCR markers of random chromosomal distribution render them suitable for mapping genomic rearrangements such as genomic deletions in tumoral tissues. This was illustrated by the detection of loss of heterozygosity in the 9q22–qter region in sporadic colon cancer.     

Proteins reacting with anti-spectrin antibodies are present in Chlamydomonas cells.

It was found either in Western-blot analysis or in indirect immunofluorescence microscopy that cells of the alga Chlamydomonas reinhardtii contain polypeptides cross-reacting with antibodies directed against red blood cell spectrin. The protein could also be detected by immunoprecipitation with anti-spectrin antibodies. C. reinhardtii cells contain distinct polypeptide chains reacting with antibodies directed against either α- or β-spectrin subunits. This protein was extracted from the cells with low ionic strength solution but was not with nonionic detergent.     

Non-random needle orientation in 1-year-old Scots pine (Pinus sylvestris L.) seedlings when adjacent to non-shading vegetation

Summary Two experiments were performed to test needle architecture of Scots pine (Pinus sylvestris L.) seedlings when affected by non-shading neighbouring vegetation. Both experiments showed that seedlings reacted to the presence of such adjacent vegetation by changes in needle distribution around the main stem: more needles were found on the opposite side of the main stem to where the vegetation was located. However, this reaction was confined to needles from the main stem. Responses of seedlings to dead neighbours differed from the response to live neighbours and also from the response to a black curtain substituted for plant neighbours. It is suggested that reactions are mediated by the red/far red ratio of incident light.     

Epidermal growth factor,phorbol esters,and aurintricarboxylic acid are survival factors for MDA-231 cells exposed to adriamycin

Summary The ability of epidermal growth factor (EGF), insulinlike growth factor-1 (IGF-1), insulin, 12-O-tetradecanoylphorbol-13-acetate (TPA), and aurintricarboxylic acid (ATA) to protect the human breast cancer cell line MDA-231 from death induced by the anticancer drug adriamycin was investigated. Cell death was induced in the MDA-231 cells either by a short-time exposure to a high dose of adriamycin (2 μg · ml⁻¹ · 1 h⁻¹) and further culturing in the absence of the drug, or by continuous exposure to a low dose of adriamycin (0.3μg/ml). Cell death was evaluated after 48 h of incubation by several techniques (trypan blue dye exclusion, lactic dehydrogenase activity, cellular ATP content, transmission electron microscopy, and DNA fragmentation). EGF, TPA, and ATA, each at an optimal concentration of 20 ng/ml, 5 ng/ml, and 100μg/ml respectively, substantially enhanced survival of cells exposed either to a high or low dose of adriamycin. Neither IGF-1 nor insulin, each at concentrations of 20 ng/ml, had an effect on cell survival. The three survival factors enhanced protein synthesis in the untreated cells and attenuated the continuous decrease in protein synthesis in the adriamycin-treated cells. Moreover, the three survival factors protected the MDA-231 cells from death in the absence of protein synthesis (cycloheximide 30μg/ml). These results suggest that EGF, TPA, and ATA promote survival of adriamycin pretreated cells by at least two mechanisms: enhancement of protein synthesis and by a protein synthesis independent process, probably a posttranslational modification effect.     

Ecological and methodological drivers of non-stationarity in tree growth response to climate

Radial tree growth is sensitive to environmental conditions, making observed growth increments an important indicator of climate change effects on forest growth. However, unprecedented climate variability could lead to non-stationarity, that is, a decoupling of tree growth responses from climate over time, potentially inducing biases in climate reconstructions and forest growth projections. Little is known about whether and to what extent environmental conditions, species, and model type and resolution affect the occurrence and magnitude of non-stationarity. To systematically assess potential drivers of non-stationarity, we compiled tree-ring width chronologies of two conifer species, Picea abies and Pinus sylvestris, distributed across cold, dry, and mixed climates. We analyzed 147 sites across the Europe including the distribution margins of these species as well as moderate sites. We calibrated four numerical models (linear vs. non-linear, daily vs. monthly resolution) to simulate growth chronologies based on temperature and soil moisture data. Climate–growth models were tested in independent verification periods to quantify their non-stationarity, which was assessed based on bootstrapped transfer function stability tests. The degree of non-stationarity varied between species, site climatic conditions, and models. Chronologies of P. sylvestris showed stronger non-stationarity compared with Picea abies stands with a high degree of stationarity. Sites with mixed climatic signals were most affected by non-stationarity compared with sites sampled at cold and dry species distribution margins. Moreover, linear models with daily resolution exhibited greater non-stationarity compared with monthly-resolved non-linear models. We conclude that non-stationarity in climate–growth responses is a multifactorial phenomenon driven by the interaction of site climatic conditions, tree species, and methodological features of the modeling approach. Given the existence of multiple drivers and the frequent occurrence of non-stationarity, we recommend that temporal non-stationarity rather than stationarity should be considered as the baseline model of climate–growth response for temperate forests.