Genetic versus environmental influence on pigment variation in the ephippia of Daphnia pulicaria

1. The dormant eggs of Daphnia (Crustacea: cladocera) are encased in a protective, chitonous casing known as an ephippium. Ephippia are pigmented with melanin, and the degree of pigmentation ranges from transparent to opaque. Variation in pigmentation exists within and across populations and species, raising questions about the factors that influence the natural distribution of pigmentation. 2. We used image analysis to quantify pigmentation in ephippial casings of Daphnia pulicaria that were produced both in the field and in the laboratory. The percentage of the surface area of ephippia that is darkly pigmented ranged from 0.5 to 99.5%. The range, mean and variance of ephippial pigmentation varied across our 11 study lakes. 3. Laboratory experiments compared clonal variation (five clones/lake) and population variation (five lakes) at three temperatures (15 °C, 20 °C and 25 °C). The degree of pigment variation between clones was much greater than pigment variation between temperatures, indicating that there is a genetic component to pigmentation in the ephippia which is stronger than the effect of temperature. 4. Comparisons of pigment levels and lake characteristics were used to identify physical and biological factors associated with ephippial pigmentation. Investment in ephippial production was the strongest predictor with darker ephippia occurring more often in lakes with the highest ephippial production.     

The European Renal Genome Project: An Integrated Approach Towards Understanding the Genetics of Kidney Development and Disease

Rapid progress in genome research creates a wealth of information on the functional annotation of mammalian genome sequences. However, as we accumulate large amounts of scientific information we are facing problems of how to integrate and relate the data produced by various genomic approaches. Here, we propose the novel concept of an organ atlas where diverse data from expression maps to histological findings to mutant phenotypes can be queried, compared and visualized in the context of a three-dimensional reconstruction of the organ. We will seek proof of concept for the organ atlas by elucidating genetic pathways involved in development and pathophysiology of the kidney. Such a kidney atlas may provide a paradigm for a new systems-biology approach in functional genome research aimed at understanding the genetic bases of organ development, physiology and disease.Key Words: EuReGene, kidney, genome, development, pathophysiology, genetics     

Widespread and ancient distribution of a noncanonical genetic code in diplomonads

Recently, a group of diplomonads has been found to use a genetic code in which TAA and TAG encode glutamine rather than termination. To survey the distribution of this characteristic in diplomonads, we sought to identify TAA and TAG codons at positions where glutamine is expected in genes for alpha-tubulin, elongation factor-1 alpha, and the gamma subunit of eukaryotic translation initiation factor-2. These sequences show that the variant genetic code is utilized by almost all diplomonads, with the genus Giardia alone using the universal genetic code. Comparative phylogenetic analysis reveals that the switch to this genetic code took place very early in the evolution of diplomonads and was likely a single event. Termination signals and downstream untranslated regions were also cloned from three Hexamita genes. In all three of these genes, the predicted TGA termination codon was found at the expected position. Interestingly, the untranslated regions of these genes are high in AT. This is incongruent with the coding regions, which are comparatively GC-rich.      

Comparative Biochemistry of the Oxidative Burst Produced by Rose and French Bean Cells Reveals Two Distinct Mechanisms

Cultured cells of rose (Rosa damascena) treated with an elicitor derived from Phytophthora spp. and suspension-cultured cells of French bean (Phaseolus vulgaris) treated with an elicitor derived from the cell walls of Colletotrichum lindemuthianum both produced H₂O₂. It has been hypothesized that in rose cells H₂O₂ is produced by a plasma membrane NAD(P)H oxidase (superoxide synthase), whereas in bean cells H₂O₂ is derived directly from cell wall peroxidases following extracellular alkalinization and the appearance of a reductant. In the rose/Phytophthora spp. system treated with N,N-diethyldithiocarbamate, superoxide was detected by a N,N′-dimethyl-9,9′-biacridium dinitrate-dependent chemiluminescence; in contrast, in the bean/C. lindemuthianum system, no superoxide was detected, with or without N,N-diethyldithiocarbamate. When rose cells were washed free of medium (containing cell wall peroxidase) and then treated with Phytophthora spp. elicitor, they accumulated a higher maximum concentration of H₂O₂ than when treated without the washing procedure. In contrast, a washing treatment reduced the H₂O₂ accumulated by French bean cells treated with C. lindemuthianum elicitor. Rose cells produced reductant capable of stimulating horseradish (Armoracia lapathifolia) peroxidase to form H₂O₂ but did not have a peroxidase capable of forming H₂O₂ in the presence of reductant. Rose and French bean cells thus appear to be responding by different mechanisms to generate the oxidative burst.     

Trophic ecology and persistence of invasive silver carp Hypophthalmichthys molitrix in an oligotrophic South African impoundment

The alien invasive silver carp Hypophthalmichthys molitrix established a self-sustaining feral population in an oligotrophic impoundment, Flag Boshielo Dam, in South Africa. The ability of this population to persist in a dam with low algal biomass (median annual suspended chlorophyll a = 0.08 µg l^?1), and limited access to rivers considered large enough for successful spawning, has implications for their invasive potential in other systems. Stomach content and stable isotope analysis were used to assess the trophic ecology of H. molitrix, which was then compared with indigenous Mozambique tilapia Oreochromis mossambicus, on a seasonal basis during 2011. Hypophthalmichthys molitrix are generalist filter feeders, with a diet consisting primarily of sediment, vegetative detritus, dinoflagellates and diatoms. The dominance of sediments in their stomachs suggests occasional benthic scavenging. However, H. molitrix occupied a higher trophic level (TL = 2.8) than expected, suggesting that this population subsidised their diet with an unidentified dietary constituent, characterised by enriched nitrogen values. Although the stomach contents indicated dietary overlap between H. molitrix and O. mossambicus, stable isotopes revealed fine-scale resource partitioning, despite both species occupying the same trophic level. Nonetheless, the persistence of this feral H. molitrix population in an oligotrophic impoundment highlights their phenotypic plasticity.     

Extracting information from imaging cytometry: a review

The extraction of statistically meaningful quantitative information from microscopy images is increasingly important for modern biological research. Obtaining accurate, quantitative information from biological specimens, however, is a complex process that requires optimization of several parameters. One must consider the number of probes, fluorescent channels required, type of plate to be used, number of fields to be acquired and optimal resolution for image acquisition. The extraction of information from images is dependent on and can be aided greatly by careful consideration of the factors involved in the image acquisition process. I summarize here the general principles behind the imaging and software technology that is used to quantify images and highlight particular issues of concern for critically applying image quantitation techniques for research.     

Sequence analysis of photoaffinity-labelled peptides derived by proteolysis of photosystem-2 reaction centres from thylakoid membranes treated with [14C]azidoatrazine.

Photosystem-2 reaction centres were prepared from pea thylakoid membranes that had been photoaffinity labelled with [14C]-azidoatrazine (2-azido-4-ethylamino-6-isopropylamino-s-triazine), a derivative of the herbicide atrazine which binds to the secondary plastoquinone electron-acceptor site of photosystem 2. SDS/PAGE of the 14C-labelled reaction centres followed by fluorography revealed photoaffinity-labelled proteins of apparent molecular masses 30 kDa and 55 kDa, which corresponded to the D1 polypeptide and to an SDS-stable heterodimer of the D1 and D2 polypeptides, respectively. To obtain sequence information on the site of photoaffinity labelling, an 8-kDa photoaffinity-labelled peptide, generated by proteolysis of the reaction-centre material with trypsin, was isolated and purified to apparent homogeneity using reverse-phase and size-exclusion HPLC techniques. The amino terminus of the photoaffinity-labelled peptide was determined to be Leu-Gly-Met-Arg-Pro-Xaa-Ile-Ala-Val-Ala-Tyr by Edman sequencing. This corresponds to the amino terminus of a predicted tryptic peptide of D1 and confirms that azidoatrazine photolabels the D1 polypeptide of photosystem 2 in the region Leu137-Arg225. Chymotrypsin/trypsin digestion of photoaffinity-labelled reaction centres followed by reverse-phase HPLC was used to isolate a smaller photoaffinity-labelled peptide. On Edman sequencing, Ser-Ala were identified as the first two residues and 14C was released on the third cycle, after which further degradation was blocked. The two potential peptide fragments with Ser-Ala at the amino terminus in the region Leu137-Arg225 are Ser148-Ala-Pro and Ser212-Ala-Met. Proline is an unlikely target for reaction with the nitrene of the photoactivated azidoatrazine, and the data are thus consistent with Met214 as the site of photoaffinity labelling on D1 when thylakoid membranes are illuminated with ultraviolet irradiation in the presence of [14C]azidoatrazine.     

Phosphorylation of phenylalanine ammonia-lyase: evidence for a novel protein kinase and identification of the phosphorylated residue.

The site of phosphorylation of phenylalanine ammonia-lyase (PAL) has been identified as a threonine residue. A Ca(2+)-stimulated protein kinase of approximately 55 kDa has been partially purified from elicited cells. The kinase can phosphorylate a synthetic peptide derived from PAL and a recombinant poplar PAL. PAL phosphorylation was associated with a decrease in Vmax in agreement with the suggestion that protein phosphorylation is involved in marking PAL subunits for turnover. The phosphorylation site in French bean PAL is most likely Thr545 in the sequence VAKRTLTT (539-546). Conservation of the phosphorylation site in PAL from diverse species suggests that phosphorylation of PAL may be a ubiquitous regulatory mechanism in higher plants.     

Fitness effects of fixed beneficial mutations in microbial populations

Beneficial mutations are intuitively relevant to understanding adaptation, yet not all beneficial mutations are of consequence to the long-term evolutionary outcome of adaptation. Many beneficial mutations-mostly those of small effect-are lost due either to (1) genetic drift or to (2) competition among clones carrying different beneficial mutations, a phenomenon called the "Hill-Robertson effect" for sexual populations and "clonal interference" for asexual populations. Competition among clones becomes more prevalent with increasing genetic linkage and increasing population size, and it is thus generally characteristic of microbial populations. Together, these two phenomena suggest that only those beneficial mutations of large fitness effect should achieve fixation, despite the fact that most beneficial mutations produced are predicted to have very small fitness effects. Here, we confirm this prediction-both empirically and theoretically-by showing that fitness effects of fixed beneficial mutations follow a distribution whose mode is positive.