Influence of ectomycorrhizal fungi on the response of Sitka spruce and Japanese larch to forms of phosphorus

Ectomycorrhizal fungi (Paxillus involutus, Suillus grevillei and two unidentified basidiomycetes from excised Sitka spruce mycorrhizas) were isolated from stands of Sitka spruce either in monoculture or in a mixture with Japanese larch in an Irish conifer plantation. The growth of these fungi and their mycorrhizal formation in Sitka spruce and Japanese larch were examined after incubation in modified Melin-Norkrans medium containing either KH₂PO₄, Ca₃(PO₄)₂ or Fe phytate as the phosphorus (P) source. P. involutus and S. grevillei utilized all three P sources. The unidentified basidiomycetes had limited ability to utilize Fe phytate. Basidiomycete 1 showed poor growth on KH₂PO₄ whereas growth of basidiomycete 2 was low on Ca₃(PO₄)₂. Pure culture synthesis studies confirmed that P. involutus and the two basidiomycetes formed mycorrhizas with both tree species but S. grevillei was mycorrhizal only on Japanese larch. P. involutus formed more mycorrhizas in both conifers than the other fungi. Following inoculation with each of the four fungi, shoot and root dry mass of both Sitka spruce and Japanese larch seedlings was enhanced compared with uninoculated/nonmycorrhizal controls. On Fe phytate, Paxillus-inoculated Sitka spruce seedlings had the lowest primary root length and on KH₂PO₄, Suillus-inoculated Japanese larch had the greatest number of short roots. The only differences when Sitka spruce seedlings were grown in either monoculture or in a mixture with Japanese larch mycorrhizal with S. grevillei were primary root length and number of short roots after growth on media containing Fe phytate.     

Satellite DNA repeat sequence variation is low in three species of burying beetles in the genus Nicrophorus (Coleoptera: Silphidae)

Three satellite DNA families were identified in three species of burying beetles, Nicrophorus orbicollis, N. marginatus, and N. americanus. Southern hybridization and nucleotide sequence analysis of individual randomly cloned repeats shows that these satellite DNA families are highly abundant in the genome, are composed of unique repeats, and are species-specific. The repeats do not have identifiable core elements or substructures that are similar in all three families, and most interspecific sequence similarity is confined to homopolymeric runs of A and T. Satellite DNA from N. marginatus and N. americanus show single-base-pair indels among repeats, but single-nucleotide substitutions characterize most of the repeat variability. Although the repeat units are of similar lengths (342, 350, and 354 bp) and A + T composition (65%, 71%, and 71%, respectively), the average nucleotide divergence among sequenced repeats is very low (0.18%, 1.22%, and 0.71%, respectively). Transition/transversion ratios from the consensus sequence are 0.20, 0.69, and 0.70, respectively.      

Receptor Activity-modifying Proteins 2 and 3 Generate Adrenomedullin Receptor Subtypes with Distinct Molecular Properties

Adrenomedullin (AM) is a peptide hormone with numerous effects in the vascular systems. AM signals through the AM₁ and AM₂ receptors formed by the obligate heterodimerization of a G protein-coupled receptor, the calcitonin receptor-like receptor (CLR), and receptor activity-modifying proteins 2 and 3 (RAMP2 and RAMP3), respectively. These different CLR-RAMP interactions yield discrete receptor pharmacology and physiological effects. The effective design of therapeutics that target the individual AM receptors is dependent on understanding the molecular details of the effects of RAMPs on CLR. To understand the role of RAMP2 and -3 on the activation and conformation of the CLR subunit of AM receptors, we mutated 68 individual amino acids in the juxtamembrane region of CLR, a key region for activation of AM receptors, and determined the effects on cAMP signaling. Sixteen CLR mutations had differential effects between the AM₁ and AM₂ receptors. Accompanying this, independent molecular modeling of the full-length AM-bound AM₁ and AM₂ receptors predicted differences in the binding pocket and differences in the electrostatic potential of the two AM receptors. Druggability analysis indicated unique features that could be used to develop selective small molecule ligands for each receptor. The interaction of RAMP2 or RAMP3 with CLR induces conformational variation in the juxtamembrane region, yielding distinct binding pockets, probably via an allosteric mechanism. These subtype-specific differences have implications for the design of therapeutics aimed at specific AM receptors and for understanding the mechanisms by which accessory proteins affect G protein-coupled receptor function.     

Membrane resistance change of the frog taste cells in response to water and Nacl

The electrical properties of the frog taste cells during gustatory stimulations with distilled water and varying concentrations of NaCl were studied with intracellular microelectrodes. Under the Ringer adaptation of the tongue, two types of taste cells were distinguished by the gustatory stimuli. One type, termed NaCl-sensitive (NS) cells, responded to water with hyperpolarizations and responded to concentrated NaCl with depolarizations. In contrast, the other type of cells, termed water-sensitive (WS) cells, responded to water depolarizations and responded to concentrated NaCl with hyperpolarizations. The membrane resistance of both taste cell types increased during the hyperpolarizing receptor potentials and decreased during the depolarizing receptor potentials, Reversal potentials for the depolarizing and hyperpolarizing responses in each cell type were a few millivolts positive above the zero membrane potential. When the tongue was adapted with Na-free Ringer solution for 30 min, the amplitude of the depolarizing responses in the NS cells reduced to 50% of the control value under normal Ringer adaptation. On the basis of the present results, it is concluded (a) that the depolarizing responses of the NS and WS cells under the Ringer adaptation are produced by the permeability increase in some ions, mainly Na+ ions across the taste cell membranes, and (b) that the hyperpolarizing responses of both types of taste cells are produced by a decrease in the cell membrane permeability to some ions, probably Na+ ions, which is slightly enhanced during the Ringer adaptation.     

Molecular evidence for the rapid propagation of mouse t haplotypes from a single, recent, ancestral chromosome

Mouse t haplotypes are variant forms of chromosome 17 that exist at high frequencies in worldwide populations of two species of commensal mice. To determine both the relationship of t haplotypes to each other and the species within which they exist, 35 representative t haplotypes were analyzed by means of 10 independent molecular probes, including five DNA clones and five polypeptide spots identified by means of two- dimensional gel electrophoresis. All of the tested haplotypes were found to share restriction fragments and polypeptide spots that are absent in mice carrying wild-type forms of chromosome 17. This observation provides the first direct evidence that all of the known t haplotypes are descendents of a single ancestral chromosome. The absence of variation among t haplotypes could mean that this ancestral chromosome existed relatively recently, in which case it would be necessary to postulate introgressions of t haplotypes across species lines to explain their presence in both Mus domesticus and M. musculus. Alternatively, it is possible that the ancestral chromosome existed prior to the split between M. domesticus and M. musculus and that, by chance, our probes fail to detect polymorphisms that exist among the t haplotypes. A further result of our analysis is the characterization of a partial t haplotype in a wild population of Israeli mice.      

MOLECULAR SYSTEMATICS OF THE XANTHOPHYCEAE

The Corallinales includes ca. 40 genera of calcified red seaweeds. Species are of two distinct morphotypes; those that possess genicula (uncalcified nodes) and those that lack genicula. Most nongeniculate species take the form of crusts. The presence (or absence) of genicula, secondary pit connections, and tetrasporangial conceptacle features have traditionally been used as key characters for delimiting coralline subfamilies. In this study, nuclear encoded 18S and 26S r RNA gene sequences were determined and used to reexamine relationships among coralline taxa. Separate and combined phylogenetic analyses of these data yielded similar trees in which four major lineages are resolved. Heydrichia and Sporolithon (Sporolithaceae) are positioned at the base of the tree and appear to be distantly related to other species examined. Within the Corallinaceae, the nongeniculate Melobesioideae is resolved as a monophyletic group. All members of this subfamily produce mutiporate tetraspoangial conceptacles. The Corallinoideae, which are characterized by unizonate genicula, are resolved as sister to a clade containing species placed in the Lithophylloideae, Mastophoroideae and Metagoniolithoideae. The molecular data indicate that geniculate and nongeniculate species characterized by the presence of secondary pit connections are closely related. For example, both data sets robustly support a sister taxon relationship between Amphiroa and Titanoderma. Our results indicate that: 1) all taxa in which secondary pit connections are present should be referred to the Lithophylloideae and, 2) genicula are nonhomologous structures that are independently derived in Amphiroa, Lithothrix, Metagoniolithon and the last common ancestor of the Corallinoideae.     

Hydrogen peroxide signaling through tumor necrosis factor receptor 1 leads to selective activation of c-Jun N-terminal kinase

Binding of tumor necrosis factor-alpha (TNFalpha) to its receptor, TNF-R1, results in the activation of inhibitor of kappaB kinase (IKK) and c-Jun N-terminal kinase (JNK) pathways that are coordinately regulated and important in survival and death. We demonstrated previously that in response to hydrogen peroxide (H2O2), the ability of TNFalpha to activate IKK in mouse lung epithelial cells (C10) was inhibited and that H2O2 alone was sufficient to activate JNK and induce cell death. In the current study, we investigated the involvement of TNF-R1 in H2O2-induced JNK activation. In lung fibroblasts from TNF-R1-deficient mice the ability of H2O2 to activate JNK was inhibited compared with fibroblasts from control mice. Additionally, in C10 cells expressing a mutant form of TNF-R1, H2O2-induced JNK activation was also inhibited. Immunoprecipitation of TNF-R1 revealed that in response to H2O2, the adapter proteins, TRADD and TRAF2, and JNK were recruited to the receptor. However, expression of the adaptor protein RIP, which is essential for IKK activation by TNFalpha, was decreased in cells exposed to H2O2, and its chaperone Hsp90 was cleaved. Furthermore, data demonstrating that expression of TRAF2 was not affected by H2O2 and that overexpression of TRAF2 was sufficient to activate JNK provide an explanation for the inability of H2O2 to activate IKK and for the selective activation of JNK by H2O2. Our data demonstrate that oxidative stress interferes with IKK activation while promoting JNK signaling, creating a signaling imbalance that may favor apoptosis.