Chronic photoinhibition in seedlings of tropical trees

Seedlings of five canopy species of tropical trees from Costa Rica and Puerto Rico were grown in full shade (midday range of photosynthetic photon flux density [PPFD], 100–140 μmol m^?2 s^?1), partial shade (midday PPFD, 400–600 μmol m^?2 s^?1) and full sun (midday PPFD, 1 500–1 800 μmol m^?2 s^?1) for 3 months. The species were Ochroma lagopus (Bombacaceae), a pioneer species; Inga edulis (Fabaceae), found in secondary forest; and Dipteryx panamensis (Fabaceae), Hampea appendiculata (Malvaceae), and Manilkara bidentata (Sapotaceae), three species characteristic of primary forest. After the plants were placed in the dark overnight, chlorophyll fluorescence characteristics were measured for recently expanded and mature leaves. The ratio of variable fluorescence to maximum fluorescence (F_v/F_m) was used to estimate the degree of chronic photoinhibition. Only individuals of one species, Dipteryx panamensis, showed significant depression of F_v/F_m after long-term exposure to full sun. The depression was highly correlated with quantum yield of O₂ evolution which also declined after exposure to full sun. The decline may have been related to foliar N concentration. Although all plants were supplied with ample nutrients, foliar N did not increase significantly for Dipteryx seedlings in full sun, whereas it did for Ochroma and Inga. Leaf age affected F_v/F_m only in the cases of Manilkara, where it was slightly lower in recently expanded leaves, and of Dipteryx where it interacted with the effects of light regime. We conclude that chronic photoinhibition is not common in seedlings of canopy trees of tropical rain forests except when availability of mineral nutrients may be limiting.     

HOSEpipE—a WWW-hosted data management and analysis system for STS content mapping projects

We have developed a data management system, `HOSEpipe' (High Output STS Evaluation pipeline) to aid sample tracking and data analysis in STS content mapping projects. The system is based around a World Wide Web (WWW) server that provides a number of pages including forms for sample processing and data entry accessible via a standard WWW browser application. The system is split into two main modules: firstly, a sequence evaluation and annotation module that takes de novo sequence for a potential STS, screens it against existing STSs and DNA sequence databases, followed by appropriate primer sequence design; secondly, a module that handles YAC library STS screening and includes facilities for both sample tracking and experimental data analysis. We present the design and rationale of the HOSEpipe system and its development to support a whole chromosomal physical mapping project. This software and design approach is potentially applicable to physical mapping projects of varying sizes and resolution and to similar projects, such as sample sequencing and the construction of sequence-ready maps. Received: 18 November 1996 / Accepted: 19 March 1997     

Mapping the diabetes polygene Idd3 on mouse Chromosome 3 by use of novel congenic strains

Development of novel congenic mouse strains has allowed us to better define the location of the diabetogenic locus, Idd3, on Chromosome (Chr) 3. Congenic strains were identified by use of published and newly developed microsatellite markers, their genomes fingerprinted by a rapid, fluorescence-based approach, and their susceptibility to type 1 diabetes evaluated. The maximum interval containing Idd3 is now approximately 4 cM.     

Hormonal stimulation of arachidonate release from isolated perfused organs. Relationship to prostaglandin biosynthesis

The lipids of isolated Krebs perfused rabbit kidneys and hearts were labelled with [14C]arachidonic acid. Subsequent hormonal stimulation (e.g. bradykinin, ATP) of the pre-labelled tissue resulted in dose-dependent release of [14C]prostaglandins; little or no release of the precursor [14C]arachidonic acid was observed. When fatty acid-free bovine serum albumin was added to the perfusion medium as a trap for fatty acids substantial release of [14C]arachidonic acid was detected following hormonal stimulation. The release of [14C]arachidonic acid was dose-dependent and greater than 3 fold that of [14C]prostaglandin release. Indomethacin by inhibiting the cyclo-oxygenase, completely inhibited release of [14C]prostaglandins and only slightly inhibited release of [14C]arachidonic acid. These results demonstrate that in both rabbit kidney and heart much more substrate is released by hormonal stimulation than is converted to prostaglandins. This suggests that either the deacylation reaction is not tightly coupled to the prostaglandin synthetase system or that there are two deacylation mechanisms, one which is coupled to prostaglandin synthesis while the other is non-specific. It has previously been shown that prostaglandin release due to hormones such as bradykinin is transient despite continued presence of the hormone (tachyphylaxis). By utilizing albumin to trap released fatty acid, it was found that hormone-stimulated release of arachidonic acid is also transient. This directly demonstrates that tachyphylaxis occurs at a step prior to the cyclo-oxygenase.     

Alkylation of nucleic acids by DNA-targeted 4-anilinoquinolinium aniline mustards: kinetic studies

The rate of constant for hydrolysis of a series of 4-substituted aniline mustards Ar-X-pC6H4-N(CH2CH2Cl)2, where Ar is 4-anilinoquinolinium and X = O, CH2, CONH and CO, have been measured in water and 0.02 M imidazole buffer at 37 degrees C and in 50% aqueous acetone at 66 degrees C. The equilibrium binding constants of the compounds and their hydrolysis products to nucleic acids of differing base composition have been determined at varying ionic strengths, and the results are consistent with the compounds binding as expected in the DNA minor groove. The alkylating reactivity of the mustards towards these nucleic acids has been measured in water at 37 degrees C and in 0.01 M HEPES buffer over a range of temperatures from 25 degrees C to 60 degrees C. Evaluation of the thermodynamic parameters for these kinetic and equilibrium studies suggests that the interaction with nucleic acids is via an internal SN2 mechanism involving an aziridinium ion.     

Cloning of wild-type Pseudomonas solanacearum phcA, a gene that when mutated alters expression of multiple traits that contribute to virulence.

Pseudomonas solanacearum undergoes a spontaneous mutation that pleiotropically reduces extracellular polysaccharide (EPS) production, endoglucanase activity, and virulence and increases motility. We refer to the process that coordinately affects these traits as phenotype conversion (PC) and the resulting mutants as PC types. Previous research with the wild-type strain AW1 suggested that inactivation of a single locus could mimic phenotype conversion (T. P. Denny, F. W. Makini, and S. M. Brumbley, Mol. Plant-Microbe Interact. 1:215-223, 1988). Additional Tn5 mutagenesis of AW1 generated three more mutants (AW1-81, AW1-82, and AW1-84) that were indistinguishable from the PC type and one slightly leaky mutant (AW1-87); all four had single insertions in the same 4.0-kilobase (kb) EcoRI fragment that were responsible for the PC-like phenotype. Another insertion mutant, AW1-83, which lacks an insertion in this 4.0-kb fragment, resembled the PC type except that it was reversibly induced to produce wild-type levels of EPS when cultured adjacent to AW1. The wild-type region containing the gene that controls traits affected by phenotype conversion in AW1, designated phcA, was cloned on a 2.2-kb DNA fragment that restored all the phcA::Tn5 mutants and 11 independent spontaneous PC-type derivatives of AW1 to wild-type status. Homology with the phcA region was found in diverse wild-type strains of P. solanacearum, although restriction fragment length polymorphisms were seen. No major DNA alterations were observed in the phcA homologous region of PC types from strain AW1 or 82N. PC types from 7 of 11 conjugal strains of P. solanacearum were restored to EPS+ by phcA from AW1; however, only some PC types of strain K60 were restored, whereas others were not. We believe that a functional phcA gene is required to maintain the wild-type phenotype in P. solanacearum, and for most strains phenotype conversion results from a loss of phcA gene expression or the function of its gene product.     

DNA-directed alkylating ligands as potential antitumor agents: sequence specificity of alkylation by intercalating aniline mustards

The sequence preferences for alkylation of a series of novel parasubstituted aniline mustards linked to the DNA-intercalating chromophore 9-aminoacridine by an alkyl chain of variable length were studied by using procedures analogous to Maxam-Gilbert reactions. The compounds alkylate DNA at both guanine and adenine sites. For mustards linked to the acridine by a short alkyl chain through a para O- or S-link group, 5'-GT sequences are the most preferred sites at which N7-guanine alkylation occurs. For analogues with longer chain lengths, the preference of 5'-GT sequences diminishes in favor of N7-adenine alkylation at the complementary 5'-AC sequence. Magnesium ions are shown to selectively inhibit alkylation at the N7 of adenine (in the major groove) by these compounds but not the alkylation at the N3 of adenine (in the minor groove) by the antitumor antibiotic CC-1065. Effects of chromophore variation were also studied by using aniline mustards linked to quinazoline and sterically hindered tert-butyl-9-aminoacridine chromophores. The results demonstrate that in this series of DNA-directed mustards the noncovalent interactions of the carrier chromophores with DNA significantly modify the sequence selectivity of alkylation by the mustard. Relationships between the DNA alkylation patterns of these compounds and their biological activities are discussed.     

Purification and serological characterization of the major envelope glycoprotein from AKR murine leukemia virus and its reactivity with autogenous immune sera from mice.

The major envelope glycoprotein (gp71) from AKR murine leukemia virus (MuLV) was purified and its serological reactivity with heterologous and autogenous immune mouse sera was examined. Homologous and interspecies competition radioimmunoassays using antisera to Rauscher-MulV gp69/71 or Friend-MuLV gp71 or antisera to feline leukemia virus to precipitate 125I-labeled gp71 from various MuLV showed that distinct differences exist between Rauscher- or Friend-MuLV and AKR-MuLV glycoproteins. Characteristically the AKR-MuLV gp71, in contrast to FLV or RLV gp71, does not compete fully in homologous or interspecies radioimmunoassays with iodinated Friend of Rauscher glycoproteins. Purified 125I-labeled AKR-MuLV gp71, in contrast to the Rauscher- or Friend-MuLV glycoproteins, reacts with normal (autogenous immune) mouse sera in direct radioimmune precipitation assays. Competition experiments further demonstrate that this is a predominant immunological reactivity of normal mouse sera which had previously been detected by radioimmune precipitation assay against intact virions.     

Substituent effects on the binding of ethidium and its derivatives to natural DNA

The binding of eight ethidium derivatives to short (approximately 35 base-pair), random sequence DNA has been investigated using 1H-NMR. At 35 degrees C, all drugs cause upfield shifts of the DNA imino proton resonances characteristic of intercalative binding to DNA, but the line shapes vary significantly with the nature of the drug. The results confirm our previous proposal that removal of the amino group at position-3, but not at position-8, on the parent ethidium shortens the lifetime of the intercalative state (less than 1-2 ms at 35 degrees C). These results suggest that hydrogen-bonding interactions with the 3-NH2 group are involved in stabilization of the drug-DNA complex or that changes in charge distribution that accompany removal of the 3-NH2 group reduce the complex stability. The magnitude of the shift of the drug-DNA spectra indicates a slight preference for binding of the drugs adjacent to G X C base-pairs.