Intraspecific nuclear DNA variation in Drosophila

We have summarized and analyzed all available nuclear DNA sequence polymorphism studies for three species of Drosophila, D. melanogaster (24 loci), D. simulans (12 loci), and D. pseudoobscura (5 loci). Our major findings are: (1) The average nucleotide heterozygosity ranges from about 0.4% to 2% depending upon species and function of the region, i.e., coding or noncoding. (2) Compared to D. simulans and D. pseudoobscura (which are about equally variable), D. melanogaster displays a low degree of DNA polymorphism. (3) Noncoding introns and 3' and 5' flanking DNA shows less polymorphism than silent sites within coding DNA. (4) X-linked genes are less variable than autosomal genes. (5) Transition (Ts) and transversion (Tv) polymorphisms are about equally frequent in non-coding DNA and at fourfold degenerate sites in coding DNA while Ts polymorphisms outnumber Tv polymorphisms by about 2:1 in total coding DNA. The increased Ts polymorphism in coding regions is likely due to the structure of the genetic code: silent changes are more often Ts's than are replacement substitutions. (6) The proportion of replacement polymorphisms is significantly higher in D. melanogaster than in D. simulans. (7) The level of variation in coding DNA and the adjacent noncoding DNA is significantly correlated indicating regional effects, most notably recombination. (8) Surprisingly, the level of polymorphism at silent coding sites in D. melanogaster is positively correlated with degree of codon usage bias. (9) Three proposed tests of the neutral theory of DNA polymorphisms have been performed on the data: Tajima's test, the HKA test, and the McDonald-Kreitman test. About half of the loci fail to conform to the expectations of neutral theory by one of the tests. We conclude that many variables are affecting levels of DNA polymorphism in Drosophila, from properties of nucleotides to population history and, perhaps, mating structure. No simple, all encompassing explanation satisfactorily accounts for the data.      

A molecular phylogeny for the Drosophila melanogaster subgroup and the problem of polymorphism data

Drosophila melanogaster belongs to a closely related group of eight species collectively known as the melanogaster subgroup; all are native to sub-Saharan Africa and islands off the east coast of Africa. The phylogenetic relationships of most species in this subgroup have been well documented; however, the three most closely related species, D. simulans, D. sechellia, and D. mauritiana, have remained problematic from a phylogenetic standpoint as no data set has unambiguously resolved them. We present new DNA sequence data on the nullo and Serendipity-alpha genes and combine them with all available nuclear DNA sequence data; the total data encompass 12 genes and the ITS of rDNA. A methodological problem arose because nine of the genes had information on intraspecific polymorphisms in at least one species. We explored the effect of inclusion/exclusion of polymorphic sites and found that it had very little effect on phylogenetic inferences, due largely to the fact that 82% of polymorphisms are autapomorphies (unique to one species). We have also reanalyzed our previous DNA-DNA hybridization data with a bootstrap procedure. The combined sequence data set and the DNA-DNA hybridization data strongly support the sister status of the two island species, D. sechellia and D. mauritiana. This at least partially resolves what had been a paradox of parallel evolution in these two species.      

Element fluxes in watershed-lake ecosystems recovering from acidification: Čertovo Lake,the Bohemian Forest, 2001–2005

Fluxes of major ions and nutrients were measured in the watershed-lake ecosystem of a strongly acidified lake, ?ertovo jezero (?ertovo Lake), in the 2001 through 2005 hydrological years. Water balance was estimated from precipitation and throughfall amounts, and measured outflow from the lake. The average water input into and outflow from the watershed-lake ecosystem was 1461 mm and 1271 mm (40 L km^?2 s^?1), respectively, and the water residence time in the lake averaged 662 days. The ecosystem has been recovering from acidification since the late 1980s. Still, however, ?ertovo watershed was an average net source of 23 mmol m^?2 yr^?1 of SO ₄ ^2? . Nitrogen saturation of the watershed caused low retention of the deposited inorganic N (23% on average). After a dry summer in 2003 and a cold winter in 2004, the watershed became a net source of inorganic N (19 mmol m^?2 yr^?1). Nitrogen transformations and SO ₄ ^2? release were the dominant terrestrial sources of H⁺ (81 and 47 mmol m^?2 yr^?1, respectively) and the watershed was a net source of 42 mmol H⁺ m^?2 yr^?1. Ionic composition of tributaries showed seasonal variations with the most pronounced changes in NO ₃ ^? , base cations, DOC, and ionic Al (Al_i) concentrations. The in-lake biogeochemical processes reduced the incoming H⁺ by ～50% (i.e., neutralized on average 222 mmol H⁺ m^?2 yr^?1, on a lake-area basis). Denitrification, SO ₄ ^2? reduction, and photochemical and microbial decomposition of allochthonous organic matter were the most important in-lake H⁺ consuming processes (215, 85, and 122 mmol H⁺ m^?2 yr^?1, respectively), while hydrolysis of Al_i was the dominant H⁺ generating process (96 mmol H⁺ m^?2 yr^?1) in ?ertovo Lake. Photochemical liberation from organic complexes was an additional in-lake source of Al_i. The net in-lake retention or removal of nutrients (carbon, phosphorus, nitrogen, and silica) varied between 18% and 34% of their inputs.     

The tick plasma lectin, Dorin M, is a fibrinogen-related molecule

A lectin, named Dorin M, previously isolated and characterized from the hemolymph plasma of the soft tick, Ornithodoros moubata, was cloned and sequenced. The immunofluorescence using confocal microscopy revealed that Dorin M is produced in the tick hemocytes. A tryptic cleavage of Dorin M was performed and the resulting peptide fragments were sequenced by Edman degradation and/or mass spectrometry. Two of three internal peptide sequences displayed a significant similarity to the family of fibrinogen-related molecules. Degenerate primers were designed and used for PCR with hemocyte cDNA as a template. The sequence of the whole Dorin M cDNA was completed by the method of RACE. The tissue-specific expression investigated by RT-PCR revealed that Dorin M, in addition to hemocytes, is significantly expressed in salivary glands. The derived amino-acid sequence clearly shows that Dorin M has a fibrinogen-like domain, and exhibited the most significant similarity with tachylectins 5A and 5B from a horseshoe crab, Tachypleus tridentatus. In addition, other protein and binding characteristics suggest that Dorin M is closely related to tachylectins-5. Since these lectins have been reported to function as non-self recognizing molecules, we believe that Dorin M may play a similar role in an innate immunity of the tick and, possibly, also in pathogen transmission by this vector.     

Influence of the cystic fibrosis transmembrane conductance regulator on expression of lipid metabolism-related genes in dendritic cells

Background

Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Infections of the respiratory tract are a hallmark in CF. The host immune responses in CF are not adequate to eradicate pathogens, such as P. aeruginosa. Dendritic cells (DC) are crucial in initiation and regulation of immune responses. Changes in DC function could contribute to abnormal immune responses on multiple levels. The role of DC in CF lung disease remains unknown.

Methods

This study investigated the expression of CFTR gene in bone marrow-derived DC. We compared the differentiation and maturation profile of DC from CF and wild type (WT) mice. We analyzed the gene expression levels in DC from naive CF and WT mice or following P. aeruginosa infection.

Results

CFTR is expressed in DC with lower level compared to lung tissue. DC from CF mice showed a delayed in the early phase of differentiation. Gene expression analysis in DC generated from naive CF and WT mice revealed decreased expression of Caveolin-1 (Cav1), a membrane lipid raft protein, in the CF DC compared to WT DC. Consistently, protein and activity levels of the sterol regulatory element binding protein (SREBP), a negative regulator of Cav1 expression, were increased in CF DC. Following exposure to P. aeruginosa, expression of 3β-hydroxysterol-Δ7 reductase (Dhcr7) and stearoyl-CoA desaturase 2 (Scd2), two enzymes involved in the lipid metabolism that are also regulated by SREBP, was less decreased in the CF DC compared to WT DC.

Conclusion

These results suggest that CFTR dysfunction in DC affects factors involved in membrane structure and lipid-metabolism, which may contribute to the abnormal inflammatory and immune response characteristic of CF.     

The EGF-TM7 family: a postgenomic view

With the human and mouse genome projects now completed, the receptor repertoire of mammalian cells has finally been elucidated. The EGF-TM7 receptors are a family of class B seven-span transmembrane (TM7) receptors predominantly expressed by cells of the immune system. Within the large TM7 superfamily, the molecular structure and ligand-binding properties of EGF-TM7 receptors are unique. Derived from the processing of a single polypeptide, they are expressed at the cell surface as heterodimers consisting of a large extracellular region associated with a TM7 moiety. Through a variable number of N-terminal epidermal growth factor (EGF)-like domains, EGF-TM7 receptors interact with cellular ligands such as CD55 and chondroitin sulfate. Recent in vivo studies demonstrate a role of the EGF-TM7 receptor CD97 in leukocyte migration. The different number of EGF-TM7 genes in man compared with mice, the chimeric nature of EMR2 and the inactivation of human EMR4 point toward a still-evolving receptor family. Here we discuss the currently available information on this intriguing receptor family.     

Determination of selected xenobiotics with ferrofluid-modified trypsin

Ferrofluid-modified trypsin has been used for the detection and determination of selected xenobiotics that inhibit trypsin activity. The procedure is useful especially when colored samples or samples containing suspended solid impurities are to be assayed. Ferrofluid-modified trypsin was inhibited by Ag⁺ and Pb²⁺, selected dyes (safranin, thionin), bacitracin and 4-aminobenzamidine. Enzymes immobilized on magnetic particles can form a basis of new automated assay procedures for the determination of xenobiotics.     

Ammonium uptake in alpine streams in the High Tatra Mountains (Slovakia)

Uptake of NH _inf4 ^sup+ -N by streambed biota of mountain brooks was studied in the alpine zone of the High Tatra Mountains. Experiments were performed involving in situ dosing of ammonium directly to the acidified stream and incubations of ammonium and streambed bryophytes in enclosures within a range of pH from 4.45 to 8.10.NH _inf4 ^sup+ -N uptake length decreased with decreasing stream discharge, while comparable values of discharge-normalized uptake lengths were found during two in situ experiments.Maximum uptake rates of NH _inf4 ^sup+ -N obtained during the incubation of bryophytes (6 to 11 mg m^–2 h^–1) were comparable with results of two in situ experiments (8 and 12 mg m^–2 h^–1). The average NH _inf4 ^sup+ -N uptake rates observed during incubations lasting 3 to 5 hours (4.3 mg m^–2 h^–1) were not related to the pH of stream water. Nitrification of about 50% of the NH _inf4 ^sup+ -N added was observed in non-acidified streams, but was negligible in acidified streams. Significant photoinhibition of nitrification was observed in non-acidified streams during enclosure experiments.