Adaptive radiation of shrubby tarweeds (Deinandra) in the California Islands parallels diversification of the Hawaiian silversword alliance (Compositae-Madiinae)

Phylogenetic analyses of nuclear rDNA transcribed spacers and cytogenetic studies of interspecific hybrids reported here uphold Carlquist's hypothesis (1965, Island Biology) that shrubby tarweeds (Deinandra) of Guadalupe Island, Mexico, are products of in situ radiation in the California Islands, where evidence of plant diversification has been equivocal. Based on the rDNA findings, the Guadalupe Island endemics (D. frutescens, D. greeneana subsp. greeneana, and D. palmeri) constitute a clade that arose since the late Pliocene, well after the origin of Guadalupe Island and diversification of annual, mainland Californian lineages of Deinandra. High interfertility and normal meiosis in F(1) hybrids between the three endemics contrast with reduced interfertility (to complete intersterility) and meiotic irregularities in F(1) hybrids between other, mostly mainland species of Deinandra. Cloned rDNA sequences provided no convincing evidence of introgression among the Guadalupe Island deinandras; morphological, phenological, and/or habitat differences among those taxa indicate ecological barriers to gene flow and a probable role for ecological divergence in diversification. Biosystematic and molecular phylogenetic data for shrubby tarweeds of Guadalupe Island and another secondarily woody, oceanic-island tarweed lineage, the Hawaiian silversword alliance, reveal strikingly similar evolutionary histories. Both groups violate Baker's Rule by stemming from self-incompatible ancestors in western North America, and each has undergone within-island diversification without evolution of strong sterility barriers among lineages. Evolutionary parallels between these Hawaiian and California Island lineages of Madiinae, first suggested by Carlquist, may reflect characteristics of tarweeds that facilitate insular colonization and adaptive radiation.     

Multiplexed immunoassays for proteins using magnetic luminescent nanoparticles for internal calibration

Suspension arrays present a promising tool for multiplexed assays in large-scale screening applications. A simple and robust platform for quantitative multiprotein immunoanalysis has been developed with the use of magnetic Co:Nd:Fe(2)O(3)/luminescent Eu:Gd(2)O(3) core/shell nanoparticles (MLNPs) as a carrier. The magnetic properties of the MLNPs allow their manipulation by an external magnetic field in the separation and washing steps in the immunoassay. Their optical properties enable the internal calibration of the detection system. The multiplexed sandwich immunoassay involves dual binding events on the surface of the MLNPs functionalized with the capture antibodies. Secondary antibodies labeled with conventional organic dyes (Alexa Fluor) are used as reporters. The amount of the bound secondary antibody is directly proportional to the concentration of the analyte in the sample. In our approach, the fluorescence intensity of the reporter dye is related to the luminescence signal of the MLNPs. In this way, the intrinsic luminescence of the MLNPs serves as an internal standard in the quantitative immunoassay. The concept is demonstrated for a simultaneous immunoassay for three model proteins (human, rabbit, and mouse IgGs). The method uses a standard bench plate reader. It can be applied to disease diagnostics and to the detection of biological threats.     

Phylogenetic and Metabolic Diversity of Planctomycetes from Anaerobic, Sulfide- and Sulfur-Rich Zodletone Spring, Oklahoma

We investigated the phylogenetic diversity and metabolic capabilities of members of the phylum Planctomycetes in the anaerobic, sulfide-saturated sediments of a mesophilic spring (Zodletone Spring) in southwestern Oklahoma. Culture-independent analyses of 16S rRNA gene sequences generated using Planctomycetes-biased primer pairs suggested that an extremely diverse community of Planctomycetes is present at the spring. Although sequences that are phylogenetically affiliated with cultured heterotrophic Planctomycetes were identified, the majority of the sequences belonged to several globally distributed, as-yet-uncultured Planctomycetes lineages. Using complex organic media (aqueous extracts of the spring sediments and rumen fluid), we isolated two novel strains that belonged to the Pirellula-Rhodopirellula-Blastopirellula clade within the Planctomycetes. The two strains had identical 16S rRNA gene sequences, and their closest relatives were isolates from Kiel Fjord (Germany), Keauhou Beach (HI), a marine aquarium, and tissues of marine organisms (Aplysina sp. sponges and postlarvae of the giant tiger prawn Penaeus monodon). The closest recognized cultured relative of strain Zi62 was Blastopirellula marina (93.9% sequence similarity). Detailed characterization of strain Zi62 revealed its ability to reduce elemental sulfur to sulfide under anaerobic conditions, as well as its ability to produce acids from sugars; both characteristics may potentially allow strain Zi62 to survive and grow in the anaerobic, sulfide- and sulfur-rich environment at the spring source. Overall, this work indicates that anaerobic metabolic abilities are widely distributed among all major Planctomycetes lineages and suggests carbohydrate fermentation and sulfur reduction as possible mechanisms employed by heterotrophic Planctomycetes for growth and survival under anaerobic conditions.     

Knocked down and out: PACAP in development, reproduction and feeding

One approach to understanding the role of PACAP in vivo is to knockdown the translation of PACAP mRNA to protein or to knock out the PACAP gene by targeted disruption. In this paper, we review the effect of PACAP knockdown with morpholinos on early brain development in zebrafish. Also reviewed is the role of PACAP at several stages of reproduction as assessed in mice with a disrupted PACAP gene. New data are presented to analyze PACAP's action in energy homeostasis (body mass, food intake, endocrine parameters) using female PACAP-null mice. The evidence suggests PACAP is important for brain development in zebrafish and is required for normal reproduction, but not for body mass or food intake in mice maintained near thermoneutrality.     

Will knowledge of human genome variation result in changing cancer paradigms?

Our incomplete understanding of carcinogenesis may be a significant reason why some cancer mortality rates are still increasing. This lack of understanding is likely due to a research approach that relies heavily on genetic comparison between cancerous and non-cancerous tissues and cells, which has led to the identification of genes of cancer proliferation rather than differentiation. Recent observations showing that a tremendous degree of natural human genetic variation occurs are likely to lead to a shift in the basic paradigms of cancer genetics, in that there is a need to consider both the nature of the genes involved, and the idea that not every genetic variation identified in these genes may be associated with carcinogenesis. Based on studies using LCM and micro-genetic analyses, we propose that significant cancer initiating events may take place during the very early stages of development of cancer-susceptible tissues and that using such techniques might greatly help us in our understanding of carcinogenesis.     

Rab27a is a key component of the secretory machinery of azurophilic granules in granulocytes

Neutrophils kill micro-organisms using microbicidal products that they release into the phagosome or into the extracellular space. The secretory machinery utilized by neutrophils is poorly characterized. We show that the small GTPase Rab27a is an essential component of the secretory machinery of azurophilic granules in granulocytes. Rab27a-deficient mice have impaired secretion of MPO (myeloperoxidase) into the plasma in response to lipopolysaccharide. Cell fractionation analysis revealed that Rab27a and the Rab27a effector protein JFC1/Slp1 (synaptotagmin-like protein 1) are distributed principally in the low-density fraction containing a minor population of MPO-containing granules. By immunofluorescence microscopy, we detected Rab27a and JFC1/Slp1 in a minor subpopulation of MPO-containing granules. Interference with the JFC1/Slp1-Rab27a secretory machinery impaired secretion of MPO in permeabilized neutrophils. The expression of Rab27a was dramatically increased when promyelocytic HL-60 cells were differentiated into granulocytes but not when they were differentiated into monocytes. Down-regulation of Rab27a in HL-60 cells by RNA interference did not affect JFC1/Slp1 expression but significantly decreased the secretion of MPO. Neither Rab27a nor JFC1/Slp1 was integrated into the phagolysosome membrane during phagocytosis. Neutrophils from Rab27a-deficient mice efficiently phagocytose zymosan opsonized particles and deliver MPO to the phagosome. We conclude that Rab27a and JFC1/Slp1 permit MPO release into the surrounding milieu and constitute key components of the secretory machinery of azurophilic granules in granulocytes. Our results suggest that the granules implicated in cargo release towards the surrounding milieu are molecularly and mechanistically different from those involved in their release towards the phagolysosome.     

Phylogeny and biogeography of the sandalwoods (Santalum, Santalaceae): repeated dispersals throughout the Pacific

Results of the first genus-wide phylogenetic analysis for Santalum (Santalaceae), using a combination of 18S-26S nuclear ribosomal (ITS, ETS) and chloroplast (3' trnK intron) DNA sequences, provide new perspectives on relationships and biogeographic patterns among the widespread and economically important sandalwoods. Congruent trees based on maximum parsimony, maximum likelihood, and Bayesian methods support an origin of Santalum in Australia and at least five putatively bird-mediated, long-distance dispersal events out of Australia, with two colonizations of Melanesia, two of the Hawaiian Islands, and one of the Juan Fernandez Islands. The phylogenetic data also provide the best available evidence for plant dispersal out of the Hawaiian Islands to the Bonin Islands and eastern Polynesia. Inability to reject rate constancy of Santalum ITS evolution and use of fossil-based calibrations yielded estimates for timing of speciation and colonization events in the Pacific, with dates of 1.0-1.5 million yr ago (Ma) and 0.4-0.6 Ma for onset of diversification of the two Hawaiian lineages. The results indicate that the previously recognized sections Polynesica, Santalum, and Solenantha, the widespread Australian species S. lanceolatum, and the Hawaiian species S. freycinetianum are not monophyletic and need taxonomic revision, which is currently being pursued.     

Why migrate? A test of the evolutionary precursor hypothesis

The question of why birds migrate is still poorly understood despite decades of debate. Previous studies have suggested that use of edge habitats and a frugivorous diet are precursors to the evolution of migration in Neotropical birds. However, these studies did not explore other ecological correlates of migration and did not control for phylogeny at the species level. We tested the evolutionary precursor hypothesis by examining the extent to which habitat and diet are associated with migratory behavior, using a species-level comparative analysis of the Tyranni. We used both migratory distance and sedentary versus migratory behavior as response variables. We also examined the influences of foraging group size, membership in mixed-species flocks, elevational range, and body mass on migratory behavior. Raw species analyses corroborated some results from studies that put forth the evolutionary precursor hypothesis, but phylogenetically independent contrast analyses highlighted an important interaction between habitat and diet and their roles as precursors to migration. Foraging group size was consistently associated with migratory behavior in both raw species and independent contrast analyses. Our results lead to a resource variability hypothesis that refines the evolutionary precursor hypothesis and reconciles the results of several studies examining precursors to migration in birds.     

Synergistic mutations produce blue-shifted bioluminescence in firefly luciferase

Light emission from the North American firefly Photinus pyralis, which emits yellow-green (557 nm) light, is widely believed to be the most efficient bioluminescence system known, making this luciferase an excellent tool for monitoring gene expression. In a previous study designed to produce luciferases for simultaneously monitoring two gene expression events, we identified a very promising blue-shifted emitter (548 nm) that contained the mutations Val241Ile, Gly246Ala, and Phe250Ser [Branchini, B. R., Southworth, T. L., Khattak, N. F., Michelini, E., and Roda, A. (2005) Red- and green-emitting firefly luciferase mutants for bioluminescent reporter applications, Anal. Biochem. 345, 140-148]. To establish the basis of the unusual blue-shifted emission, we determined that a simple additive effect of the three individual mutations did not account for the spectral properties of the triple mutant. Instead, the bioluminescence emission spectra of two double mutants containing Phe250Ser and either Val241Ile or Gly246Ala very closely resembled that of the triple mutant. Additional mutagenesis results confirmed that the blue-shifted emission of the double mutants was determined by the synergistic behavior of active site residues. Molecular modeling studies of the Gly246Ala and Phe250Ser double mutant supported the notion that the blue-shifted emission was due to localized changes that increased the hydrophobicity at the emitter site as a result of the addition of a single methyl group at position 246. Moreover, the modeling data suggested that the Ala246 side chain remained close to the emitter through an additional H-bond between Ala246 and the hydroxyl group of Phe250, providing a possible structural basis for the synergistic behavior.     

Raised salivary testosterone in women is associated with increased attraction to masculine faces

Women's preferences for masculinity in men's faces, voices and behavioral displays change during the menstrual cycle and are strongest around ovulation. While previous findings suggest that change in progesterone level is an important hormonal mechanism for such variation, it is likely that changes in the levels of other hormones will also contribute to cyclic variation in masculinity preferences. Here we compared women's preferences for masculine faces at two points in the menstrual cycle where women differed in salivary testosterone, but not in salivary progesterone or estrogen. Preferences for masculinity were strongest when women's testosterone levels were relatively high. Our findings complement those from previous studies that show systematic variation in masculinity preferences during the menstrual cycle and suggest that change in testosterone level may play an important role in cyclic shifts in women's preferences for masculine traits.