Liana diversity and distribution in four tropical dry evergreen forests on the Coromandel coast of south India

Liana diversity was inventoried in four tropical dry evergreen forest sites that are characterized by numerous trees, of short stature and small diameter, and a varying degree of anthropogenic disturbance, on the Coromandel coast of south India. A 1-ha plot was established in each of the four sites and was subdivided into 100 quadrats of 10 m× 10 m. All lianas 1 cm diameter at breast height (dbh) rooted within the plot were enumerated. The species richness and density of lianas, with respect to site disturbance and forest stature, varied across the sites. Liana density totaled 3307 individuals (range 497–1163 individuals ha^–1) and species richness totaled 39 species (range 24–29 species ha^–1) representing 34 genera and 24 families. Combretaceae, Asclepiadaceae, Capparaceae and Vitaceae were the well-represented families. The top five species Strychnos minor, Combretum albidum, Derris ovalifolia, Jasminum angustifolium and Reissantia indica contributed 55% of total density. The slopes of the species–area curves were different for each of the four sites and the curve stabilized in only one site. Of the four climbing modes recognized among the total 39 species, 18 were twiners (56% of the total density). Eight species (24% of density) were tendril climbers and 12 species (16% of density) were scramblers. Hugonia mystax was the only hook climber. All the 39 species and 88% of liana density were encountered within a category of 6 cm dbh or less, and a similar pattern prevailed in the individual sites. Of the three diaspore dispersal modes found among the 39 liana species, animal (64%) and wind (23%) dispersal were predominant over the autochorous mode (13%). Liana diversity and distribution in dry forest communities appear to be influenced by forest stature and site disturbance levels. In the light of the extent of liana diversity and sacred grove status of the study sites, the need for forest conservation, involving local people, is emphasized.     

Functional genomics tools for the analysis of zebrafish pigment

Genetic model organisms are increasingly valuable in the post-genomics era to provide a basis for comparative analysis of the human genome. For higher order processes of vertebrate pigment cell biology and development, the mouse has historically been the model of choice. A complementary organism, the zebrafish (Danio rerio), shares many of the signaling and biological processes of vertebrates, e.g. neural crest development. The zebrafish has a number of characteristics that make it an especially valuable model for the study of pigment cell biology and disease. Large-scale genetic screens have identified a collection of pigmentation mutants that have already made valuable contributions to pigment research. An increasing repertoire of genomic resources such as an expressed sequence tag-based Gene Index (The Institute for Genomic Research) and improving methods of mutagenesis, transgenesis, and gene targeting make zebrafish a particularly attractive model. Morpholino phosphorodiamidate oligonucleotide (MO) 'knockdown' of pigment gene expression provides a non-conventional antisense tool for the analysis of genes involved in pigment cell biology and disease. In addition, an ongoing, reverse-genetic, MO-based screen for the rapid identification of gene function promises to be a valuable complement to other high-throughput microarray and proteomic approaches for understanding pigment cell biology. Novel reagents for zebrafish transgenesis, such as the Sleeping Beauty transposon system, continue to improve the capacity for genetic analysis in this system and ensure that the zebrafish will be a valuable genetic model for understanding a variety of biological processes and human diseases for years to come.     

DNA damage and cytotoxicity induced by minor groove binding methyl sulfonate esters

Minor groove specific DNA equilibrium binding peptides (lex) based on N-methylpyrrole-carboxamide and/or N-methylimidazolecarboxamide subunits have been modified with an O-methyl sulfonate ester functionality to target DNA methylation in the minor groove at Ade/Thy- and/or Gua/Cyt-rich sequences. HPLC and sequencing gel analyses show that the Me-lex compounds all selectively react with DNA to afford N3-alkyladenine as a major adduct. The formation of the N3-alkyladenine lesions is sequence-dependent based on the equilibrium binding preferences of the different lex peptides. In addition to the reaction at adenine, the molecules designed to target Gua/Cyt sequences also generate lesions at guanine; however, the methylation is not sequence dependent and takes places in the major groove at the N7-position. To determine if and how the level of the different DNA adducts and the sequence selectivity for their formation affects cytotoxicity, the Me-lex analogues were tested in wild type Escherichia coli and in mutant strains defective in base excision repair (tag and/or alkA or apn). The results demonstrate the importance of 3-methyladenine, and in some cases 3-methylguanine, lesions in cellular toxicity, and the dominant protective role of the DNA glycosylases. There is no evidence that the sequence specificity is related to toxicity.     

Structure of a c-kit product complex reveals the basis for kinase transactivation

The c-Kit proto-oncogene is a receptor protein-tyrosine kinase associated with several highly malignant human cancers. Upon binding its ligand, stem cell factor (SCF), c-Kit forms an active dimer that autophosphorylates itself and activates a signaling cascade that induces cell growth. Disease-causing human mutations that activate SCF-independent constitutive expression of c-Kit are found in acute myelogenous leukemia, human mast cell disease, and gastrointestinal stromal tumors. We report on the phosphorylation state and crystal structure of a c-Kit product complex. The c-Kit structure is in a fully active form, with ordered kinase activation and phosphate-binding loops. These results provide key insights into the molecular basis for c-Kit kinase transactivation to assist in the design of new competitive inhibitors targeting activated mutant forms of c-Kit that are resistant to current chemotherapy regimes.     

Synthetic shuffling expands functional protein diversity by allowing amino acids to recombine independently

We describe synthetic shuffling, an evolutionary protein engineering technology in which every amino acid from a set of parents is allowed to recombine independently of every other amino acid. With the use of degenerate oligonucleotides, synthetic shuffling provides a direct route from database sequence information to functional libraries. Physical starting genes are unnecessary, and additional design criteria such as optimal codon usage or known beneficial mutations can also be incorporated. We performed synthetic shuffling of 15 subtilisin genes and obtained active and highly chimeric enzymes with desirable combinations of properties that we did not obtain by other directed-evolution methods.     

Breakdown of Ficus and Eucalyptus leaves in an organically polluted river in India: fungal diversity and ecological functions

1. At two organically polluted sites in the River Nethravathi, banyan and eucalypt leaves were colonized by one or two species of aquatic hyphomycetes. A total of three or four species were identified at the two sites in samples of water and naturally occurring leaves.
2. Spore production from stream‐exposed leaves by aquatic hyphomycetes was lower by a factor of up to 1 million compared with an earlier study in geographically close but unpolluted streams.
3. Exponential decay rates and loss rates of phosphorus and calcium, were not statistically different from an earlier study in unpolluted streams. Nitrogen increased during decomposition more slowly in the current study.
4. The microbial community on both leaves released enzymes active against starch, pectin, cellulose and xylan.
5. Banyan leaves conditioned for 12 weeks were more palatable to the gastropod Notopala sp. than unconditioned leaves.
6. Together with earlier data from unpolluted streams, the study provides evidence that organic pollution severely restricts diversity of aquatic hyphomycetes and their reproductive output, but does not have an equally strong effect on ecological functions generally associated with this group.     

Cytotoxicity of glucose analogues in V79 multicell spheroids

2-Deoxy-D-glucose (2DG) and 5-thio-D-glucose (5TG) are glucose antimetabolites that are known to be selectively toxic to hypoxic cells grown as single cells or as monolayer cultures. These analogues were toxic to Chinese hamster V79 cells grown as multicell spheroids even under aerobic conditions. When spheroids, 500- to 600-microns diameter, were exposed to 7.5 mM of these chemicals for 3 days, the number of clonogenic cells per spheroid dropped to 50% for 5-thio-D-glucose and 20% for 2-deoxy-D-glucose, relative to control values. Survivals were reduced to less than 1% when the experiment was repeated in glucose-free medium. Scanning electron photomicrographs of spheroids treated with 7.5 mM of either analogue showed extensive damage to the outer cells. The cell killing observed was much more than could be predicted on the basis of the hypoxic fraction known to be present in these spheroids. The crowded tumor-like environment may make the cells vulnerable to the cytotoxic action of glucose analogues and other glycolytic inhibitors.     

Effect of radiosensitizing agents on electron transport systems

Experiments have been carried out to study the interaxtion between chemical radiosensitizing agents and model electron transport systems. Using an NAD(P)H:O2 oxidoreductase enzyme as such a model, it was demonstrated that radiosensitizers can act as intermediates in the transfer of electrons from NADH to O2, even in the presence of classical inhibitors of electron transport, with anefficiency related to both their redox potentials and their radiosensitizing abilities. This work which was further confirmed in mammalian mitochondria and microsomes as well as in a cultured cell system indicated that these sensitizers can accept electrons from a variety of organelle systems. This action was shown to be related to the concentration of reduced pyridine nucleotides present both in vivo and in vitro. Of the electron-affinic agents tested, those whose redox potential was more negative than -0.39 V may possibly serve as better radiotherqpeutic mediators.