Sequence, organization, and evolution of the A+T region of Drosophila melanogaster mitochondrial DNA

The long (4.6-kb) A+T region of Drosophila melanogaster mitochondrial DNA has been cloned and sequenced. The A+T region is organized in two large arrays of tandemly repeated DNA sequence elements, with nonrepetitive intervening and flanking sequences comprising only 22% of its length. The first repeat array consists of five repeats of 338-373 bp. The second consists of four intact 464-bp repeats and a fifth partial repeat of 137 bp. Three DNA sequence elements are found to be highly conserved in D. melanogaster and in several Drosophila species with short A+T regions. These include a 300-bp DNA sequence element that overlaps the DNA replication origin and two thymidylate stretches identified on opposite DNA strands. We conclude that the length heterogeneity observed in the A+T regulatory region in mitochondrial DNAs from the genus Drosophila results from the expansion (and contraction) of the number of repeated DNA sequence elements. We also propose that the 300-bp conserved DNA sequence element, in conjunction with another primary sequence determinant, perhaps the adjacent thymidylate stretch, functions in the regulation of mitochondrial DNA replication.      

Statistical discovery of site inter-dependencies in sub-molecular hierarchical protein structuring

Background

Much progress has been made in understanding the 3D structure of proteins using methods such as NMR and X-ray crystallography. The resulting 3D structures are extremely informative, but do not always reveal which sites and residues within the structure are of special importance. Recently, there are indications that multiple-residue, sub-domain structural relationships within the larger 3D consensus structure of a protein can be inferred from the analysis of the multiple sequence alignment data of a protein family. These intra-dependent clusters of associated sites are used to indicate hierarchical inter-residue relationships within the 3D structure. To reveal the patterns of associations among individual amino acids or sub-domain components within the structure, we apply a k-modes attribute (aligned site) clustering algorithm to the ubiquitin and transthyretin families in order to discover associations among groups of sites within the multiple sequence alignment. We then observe what these associations imply within the 3D structure of these two protein families.

Results

The k-modes site clustering algorithm we developed maximizes the intra-group interdependencies based on a normalized mutual information measure. The clusters formed correspond to sub-structural components or binding and interface locations. Applying this data-directed method to the ubiquitin and transthyretin protein family multiple sequence alignments as a test bed, we located numerous interesting associations of interdependent sites. These clusters were then arranged into cluster tree diagrams which revealed four structural sub-domains within the single domain structure of ubiquitin and a single large sub-domain within transthyretin associated with the interface among transthyretin monomers. In addition, several clusters of mutually interdependent sites were discovered for each protein family, each of which appear to play an important role in the molecular structure and/or function.

Conclusions

Our results demonstrate that the method we present here using a k- modes site clustering algorithm based on interdependency evaluation among sites obtained from a sequence alignment of homologous proteins can provide significant insights into the complex, hierarchical inter-residue structural relationships within the 3D structure of a protein family.

     

Overexpression of alcohol oxidase in Pichia pastoris

The protein import capacity of peroxisomes in methylotrophic yeasts was studied using Pichia pastoris containing one or two extra copies of the gene encoding the peroxisomal protein alcohol oxidase. The alcohol oxidase overproduced in this strain was only partially imported and assembled into the active, octameric form of the protein. The excess remained in the cytosol as protein aggregates composed of monomers. These results are discussed in view of the possible application of peroxisomes as storage compartments for heterologous proteins.     

Accumulation rates of soil organic matter in wet dune slacks on the Dutch Wadden Sea islands

Background and aims

A long-term monitoring program (ranging from 16 to 77 years) on the Dutch Wadden Sea Islands provided well-documented examples of vegetation succession in wet dune slacks. We used this opportunity to study soil organic matter (SOM) accumulation in relation to vegetation succession. The aim of this paper is to identify the factors which regulate accumulation rates of SOM in wet dune slacks.

Methods

We used several soil chronosequences using data from the monitoring program together with data from a long-term research activity and more recent measurements. We used several soil chronosequences using data from the monitoring program together with data from a long-term research (up to 150 years) and more recent measurements. Field measurements included pH, soil organic matter, above ground standing crop and water levels. Water level regimes (inundation duration and mean minimum water level), were simulated using a hydrological model. Capable of simulating inundation duration and water-level fluctuations, this model used field measurements collected over more than 5 years, as well as precipitation and evapotranspiration data collected over a period of 25 years.

Results

Sampling two synchronic chronosequences showed that SOM accumulations increased linearly during the first 50–60 years and then levelled off. Sampling various diachronic chronosequences over time showed a wide variation in accumulation rates. Slacks with low productive species, such as Littorella uniflora, showed low accumulation rates (0.02–0.08 kg/m²/year), and persisted even over a period of more than 90 years. In contrast, slacks dominated by high productive species, such as Phragmites australis, showed ten times higher accumulation rates (0.17–0.26 kg/m²/year) over a similar time period and comparable annual inundation periods (176–240 days). A multiple linear regression showed that variation in SOM accumulation rates was best explained by above-ground biomass of the vegetation.

Conclusions

We conclude that the rate of SOM accumulation in wet dune slacks is primarily controlled by plant above-ground biomass. Both above-ground biomass and SOM accumulation can remain very low over a long period of time when dune slacks are flooded during most of the year and plants with adaptive traits are able to maintain vegetation succession at a pioneer stage.     

Ecdysteroids and meiotic reinitiation in Palaemon serratus (Crustacea Decapoda Natantia) and in Locusta migratoria (Insecta orthoptera). A comparative study

Summary

Meiotic reinitiation has been studied in Locusta migratoria and Palaemon serratus in relation to the titre of free ecdysteroids present in the maturing oocyte. In both species meiotic reinitiation is characterized by two meiotic arrests, in prophase I and in metaphase I, and the first meiotic resumption which leads to germinal vesicle breakdown (GVBD) is correlated with increasing titres of ecdysteroids in the oocyte. Meiotic reinitiation has been successfully triggered in the oocytes of both species by incubation with physiological doses of ecdysteroids.     

Evidence against the use of surrogates for biomonitoring of Neotropical floodplains

1. Community concordance measures the level of association between the compositional patterns shown by two groups of organisms. If strong community concordance occurs, one group could be used as a surrogate for another in conservation planning and biodiversity monitoring. In this study, we evaluated the variability in the strength of community concordance, the likely mechanisms underlying community concordance and the degree to which one community can predict another in a set of Neotropical floodplain lakes (Upper Paraná River floodplain, Brazil). 2. We used a data set including six aquatic communities: fish, macrophytes, benthic macroinvertebrates, zooplankton, phytoplankton and periphyton. We used Mantel and PROTEST approaches to evaluate the levels of community concordance in up to four sampling periods. Also, we used partial Mantel test and information about biotic interactions to investigate reasons for observed patterns of concordance. Finally, we used co‐correspondence analysis to evaluate the performance of one taxonomic group in predicting the structures of other communities. 3. The levels of community concordance varied over time for almost all cross‐taxa comparisons. Concordance between phytoplankton and periphyton probably resulted from similar responses to environmental gradients, whereas other patterns of concordance were likely generated by interactions among groups. However, the levels of predictability were low, and no particular taxonomic group significantly predicted all other groups. 4. The low and temporally variable levels of community concordance cast doubts on the use of surrogate groups for biodiversity management in Neotropical floodplains.     

Monitoring nitrogen leaching for the evaluation of the Dutch minerals policy for agriculture in clay regions

This paper presents the results of the Dutch monitoring program for agriculture in the clay regions for the period 1996-2000 and evaluates the monitoring strategy. A wide range of farms (25 to 85%) had a NO3--N concentration in tile drainwater higher than the EU standard of 11.3 mg/l. The low figure is related to wet winters; the high, to dry winters. Arable farms are more prone to NO3- leaching than dairy farms. On arable farms, about 25% of the N surplus leached to groundwater and tile drainwater, on dairy farms this was about 15%. N in tile drainwater has shown to be the best indicator for monitoring the effects of farming practice changes in the clay regions. The average NO3--N concentration in tile drainwater was 18.8 and 3.2 mg/l in borehole water on farms where both were monitored. It is known that N use has a relationship with NO3- in tile drainwater and not with NH4+ and organic N. The presented results indicate that crop rotation and precipitation strongly influence NO3- concentration in tile drainwater.     

Entanglement is a costly life‐history stage in large whales

Individuals store energy to balance deficits in natural cycles; however, unnatural events can also lead to unbalanced energy budgets. Entanglement in fishing gear is one example of an unnatural but relatively common circumstance that imposes energetic demands of a similar order of magnitude and duration of life‐history events such as migration and pregnancy in large whales. We present two complementary bioenergetic approaches to estimate the energy associated with entanglement in North Atlantic right whales, and compare these estimates to the natural energetic life history of individual whales. Differences in measured blubber thicknesses and estimated blubber volumes between normal and entangled, emaciated whales indicate between 7.4 × 10¹⁰ J and 1.2 × 10¹¹ J of energy are consumed during the course to death of a lethal entanglement. Increased thrust power requirements to overcome drag forces suggest that when entangled, whales require 3.95 × 10⁹ to 4.08 × 10¹⁰ J more energy to swim. Individuals who died from their entanglements performed significantly more work (energy expenditure × time) than those that survived; entanglement duration is therefore critical in determining whales’ survival. Significant sublethal energetic impacts also occur, especially in reproductive females. Drag from fishing gear contributes up to 8% of the 4‐year female reproductive energy budget, delaying time of energetic equilibrium (to restore energy lost by a particular entanglement) for reproduction by months to years. In certain populations, chronic entanglement in fishing gear can be viewed as a costly unnatural life‐history stage, rather than a rare or short‐term incident.     

Induction kinetics and cell surface distribution of Escherichia coli lipoprotein under lac promoter control.

The induction kinetics and surface accessibility of the outer membrane lipoprotein were studied in an Escherichia coli strain with the lpp gene under control of the lac promoter. Free lipoprotein appeared rapidly after induction with isopropyl-beta-D-thiogalactopyranoside and reached a steady-state level after 30 min. The newly induced lipoprotein was slowly bound to the peptidoglycan layer. Immunological methods were developed to detect lipoprotein accessible at the cell surface after various pretreatments as well as peptidoglycan-bound lipoprotein at the surface of isolated peptidoglycan sacculi with specific antibodies in combination with 125I-protein A. With these methods an increase in lipoprotein molecules at the cell surface and bound to the peptidoglycan sacculus could be detected following induction. The topology of newly synthesized lipoprotein was examined in thin sections as well as at the cell surface and the surface of the peptidoglycan sacculus with immunoelectron microscopy. Ultrathin cell sections, whole cells, and isolated peptidoglycan sacculi showed lipoprotein distributed homogeneously over the entire surface.