Indole-3-acetic acid levels after phytochrome-mediated changes in the stem elongation rate of dark- and light-grownPisum seedlings

The effect of red (R) and far-red (FR) light on stem elongation and indole-3-acetic acid (IAA) levels was examined in dwarf and tall Pisum sativum L. seedlings. Red light reduced the extension-growth rate of etiolated seedlings by 70–90% after 3 h, and this inhibition was reversible by FR. Inhibition occurred throughout the growing zone. After 3 h of R, the level of extractable IAA in whole stem sections from the growing zone of etiolated plants either increased or showed no change. By contrast, extractable IAA from epidermal peels consistently decreased 3 h after R treatments. Decreases of 40% were observed for epidermal peels from the top 1 cm of tall plants receiving 3 h R. Brief R treatments resulted in smaller decreases in epidermal IAA levels and these decreases were not as great when FR followed R. In lightgrown plants, end-of-day FR stimulated growth during the following dark period in a photoreversible manner. The uppermost 1 cm of expanding third internodes was most responsive to the FR. Extractable IAA from epidermal peels from the upper 1 cm of third internodes increased by 30% or more 5 h after FR. When R followed the FR the increases were smaller. Levels of IAA in whole stem sections did not change and were twofold greater than in dark-grown plants. In both dark- and light-grown tall plants, IAA levels were lower in epidermal peels than in whole stem segments. These results provide evidence that IAA is compartmentalized at the tissue level within the growing stem and that phytochrome regulation of stem elongation rates may be partly based on modulating the level of IAA within the epidermis.Abbreviations IAA indole-3-acetic acid - R red light - FR farred light We thank Yu-Xian Zhu for helping to develop methods for IAA analysis, James Reid for supplying the genetic lines of Pisum and Richard Cyr for the use of microscopy equipment. This work was supported by NSF grant DCB-8801880 and by Hatch funds from the College of Agriculture and Life Sciences at Cornell University. The gas chromatograph-mass spectrometer was funded by NSF grant DMB-8505974 and funds from the College of Agriculture and Life Sciences at Cornell University. A preliminary report of some of these experiments has appeared in Plant Growth Substances, 1991 (Behringer et al. 1992 b).     

Sulfate reduction and S-oxidation in a moorland pool sediment

In an oligotrophic moorland pool in The Netherlands, S cycling near the sediment/water boundary was investigated by measuring (1) SO₄ ^2– reduction rates in the sediment, (2) depletion of SO₄ ^2– in the overlying water column and (3) release of³⁵S from the sediment into the water column. Two locations differing in sediment type (highly organic and sandy) were compared, with respect to reduction rates and depletion of SO₄ ^2– in the overlying water.Sulfate reduction rates in sediments of an oligotrophic moorland pool were estimated by diagenetic modelling and whole core³⁵SO₄ ^2– injection. Rates of SO₄ ^2– consumption in the overlying water were estimated by changes in SO₄ ^2– concentration over time in in situ enclosures. Reduction rates ranged from 0.27–11.2 mmol m^–2 d^–1. Rates of SO₄ ^2– uptake from the enclosed water column varied from –0.5, –0.3 mmol m^–2 d^–1 (November) to 0.43–1.81 mmol m^–2 d^–1 (July, August and April). Maximum rates of oxidation to SO₄ ^2– in July 1990 estimated by combination of SO₄ ^2– reduction rates and rates of in situ SO₄ ^2– uptake in the enclosed water column were 10.3 and 10.5 mmol m^–2 d^–1 at an organic rich and at a sandy site respectively.Experiments with³⁵S^2– and³⁵SO₄ ^2– tracer suggested (1) a rapid formation of organically bound S from dissimilatory reduced SO₄ ^2– and (2) the presence of mainly non SO₄ ^2–-S derived from reduced S transported from the sediment into the overlying water. A³⁵S^2– tracer experiment showed that about 7% of³⁵S^2– injected at 1 cm depth in a sediment core was recovered in the overlying water column.Sulfate reduction rates in sediments with higher volumetric mass fraction of organic matter did not significantly differ from those in sediments with a lower mass fraction of organic matter.Corresponding author     

Structural investigation of bacteriorhodopsin and some of its photoproducts by polarized Fourier transform infrared spectroscopic methods-difference spectroscopy and photoselection

The direction of selected IR-transition moments of the retinal chromophore of bacteriorhodopsin (BR) and functional active amino acid residues are determined for light- and dark-adapted BR and for the intermediates K and L of the photocycle. Torsions around single bonds of the chromophore are found to be present in all the investigated BR states. The number of twisted single bonds and the magnitude of these torsions decreases in the order K, L, light-adapted BR, dark-adapted BR. In the last, only the C₁₄—C₁₅ single bond is twisted. The orientation of molecular planes and chemical bonds of such protein side chains, which are perturbed during the transition of light-adapted BR to the respective intermediates, are deduced and the results compared with the current three dimensional model of BR. Trp 86 and Trp 185 are found to form a rigid part of the protein, whereas Asp 96 and Asp 115 perform molecular rearrangements upon formation of the L-intermediate.     

Detection of methandienone (methandrostenolone) and metabolites in horse urine by gas chromatography—mass spectrometry

The metabolic transformation of methandienone (I) in the horse was investigated. After administration of a commercial drug preparation to a female horse (0.5 mg/kg), urine samples were collected up to 96 h and processed without enzymic hydrolysis. Extraction was performed by a series of solid—liquid and liquid—liquid extractions, thus avoiding laborious purification techniques. For analysis by gas chromatography—mass spectrometry, the extracts were trimethylsilylated. Besides the parent compound I and its C-17 epimer II, three monohydroxylated metabolites were identified: 6β-hydroxymethandienone (III), its C-17 epimer (IV) and 16β-hydroxy-methandienone (V). In addition, three isomers of 6β,16-dihydroxymethandienone (VIa–c) were discovered. Apparently, reduction of the δ⁴ double bond of 16β-hyroxymethandienone (V) in the horse yields 16β,17β-dihydroxy-17α-methyl-5β-androst-1-en-3-one (VII). Reduction of the isomers VIa–c results in the corresponding 6β,16,17-trihydroxy-17-methyl-5β-androst-1-en-3-ones (VIIIa–c). The data presented here suggest that screening for the isomers of VI and VIII, applying the selected-ion monitoring technique, will be the most successful way of proving methandienone administration to a horse.     

Histidine utilization by Alcaligenes eutrophus: Regulation of histidase formation under heterotrophic conditions of growth

Histidine supported good growth of Alcaligenes eutrophus strain H 16 as a nitrogen source, but only poor growth as a carbon and energy source. The facultative chemolithoautotrophic bacterium was also able to utilize urocanic acid, the first intermediate of histidine catabolism. The products of histidine degradation were ammonium, formate and glutamate. Three enzymes of the pathway, histidase, urocanase and formiminoglutamate hydrolase, were present in histidine-grown cells. Two types of spontaneous mutants, derived from the wild type, were characterized by an increased growth rate on histidine. One of these types was found to produce histidase constitutively and at a higher activity compared with the parental strain. The second type of mutant had apparently gained an improved histidine uptake system, which is supposed to be growth rate-limiting in the wild type. From the physiological studies the conclusion was drawn that the control of histidine-degrading enzymes is based on induction by urocanate and catabolite repression by carbon sources supporting fast growth, such as succinate or pyruvate. Ammonium was found not to affect catabolite repression, however, we obtained evidence that histidine uptake is subject to a nitrogen control.Abbreviation CTAB hexadecyltrimethylammonium bromide     

Biosynthesis of cyclopentenylglycine from α-ketopimelate in Idesia polycarpa callus cultures

The nonproteinogenic amino acid, cyclopentenylglycine, is found in certain Flacourtiaceae. This compound may be synthesized by two C₁-chain elongations of -ketoglutarate via -ketopimelate (C₅+2C₁) or by condensation of C₄ and C₃ units (C₄+C₃), a pathway not involving -ketopimelate. The following experimental design elucidated the biosynthetic pathway: Idesia polycarpa callus cultures were freshly established from leaf petioles; synthetic -[1,2-¹⁴C]ketopimelate was added to the medium and cultures were incubated for 3 weeks. After isolation and separation of free amino acids from the tissues, the radioactivity incorporated into cyclopentenylglycine was determined. The results establish -ketopimelate as a precursor for cyclopentenylglycine, thus providing evidence for the C₅+2C₁ biosynthetic path.     

Quantitative evaluation of gel electrophoretic patterns by videodensitometry

Videodensitometry based on television technique has been shown to be suitable for recording gel electrophoretic patterns. Its speed of operation is about 60 times as fast as that of conventional densitometers, whereas the patterns recorded are practically the same.     

Post-transformational rearrangement of an in vitro reconstructed group-A streptococcal erythromycin resistance plasmid

Cleavage of the group-A streptococcal macrolide, lincosamide, and streptogramin B (MLS) resistance plasmid pSM19035 yields 2 fragments [13 and 4 megadaltons (MD)] with EcoRI, and 15 fragments with HindIII, 12 of which are 6 pairs of identical fragments derived from the inverted repeats that comprise about 80% of the pSM19035 genome. The large EcoRI fragment was isolated, ligated, and used to transform the Challis strain of Streptococcus sanguis to erythromycin resistance. Plasmids (pDB101, pDB102, and pDB103) isolated from three different transformants had lower molecular masses than the original large EcoRI fragment. HindIII digestion of these molecules and subsequent analysis of fragment radioactivity distributions indicated the loss of plasmid segments of various sizes. The deletions, all of which occurred in the palindrome, did not affect the level and the inducible nature of pSM19035-determined antibiotic resistance. Only pDB101 retained the unique EcoRI cleavage site. The results of this analysis allowed the construction of an EcoRI and HindIII cleavage-site map of pSM19035 and promise to simplify future studies of genetic functions specified by streptococcal MLS resistance plasmids.     

Studies on the organization of the α-satellite DNA from African green monkey cells using restriction nucleases and molecular cloning

α-Satellite DNA from African green monkey cells was analysed with restriction nucleases in some detail confirming and complementing our earlier results. With EcoRI and HaeIII (or BsuRI isoschizomer), about 25 and 10%, respectively, of the satellite DNA were cleaved into a series of fragments of the 172 bp repeat length and multiples thereof. To allow studies with fragments of homogeneous sequence unit length, HindIII fragments were covalently joined with the plasmid pBR313. After transformation 19 clones were obtained, containing up to three monomer fragments. Nine of the clones were characterized by digestion with EcoRI. Three of these had cleavage sites for this nuclease in the satellite DNA portion. In the six clones tested with HaeIII no cleavage site was detected in the cloned DNA. The results are discussed in relation to the nucleotide sequence data recently published by Rosenberg et al. (1978) and in the context of random and nonrandom processes in satellite DNA evolution.     

Indomethacin inhibition of cell proliferation induced by the phorbolester TPA is reversed by prostaglandin E2 in mouse epidermis in vivo

The proliferative response of mouse epidermis to the phorbolester TPA (10 nmoles) in vivo is completely inhibited by a single topical application of indomethacin one hour before TPA. DNA labeling in normal mouse epidermis is not significantly depressed by the drug. The inhibition can be reversed by applying prostaglandin E₂ (>3 nmoles) simultaneously with TPA, whereas prostaglandin F_2α (100 nmoles) is ineffective. The indomethacin-sensitive event is restricted to the first hour after phorbol ester treatment. TPA-induced skin inflammation is not influenced by the drug. It is proposed that prostaglandin E₂, or a closely related compound, mediates the mitogenic effect of TPA in mouse skin.