Analysis of covariance patterns in gene expression data and FT-IR spectra

The aim of this study was to detect and interpret correlation patterns in several large data matrices from the same biological system using Partial Least Squares Regression (PLSR) in order to get information on the system under investigation. To do this, DNA microarray data and Fourier Transform Infrared (FT-IR) spectra from a designed study where Campylobacter jejuni was exposed to environmental stress conditions, were used. The experimental design included variation in atmospheric conditions, temperature and time. PLSR was first used to analyse each of the two data types separately in order to explore the effect of the experimental parameters on the data. The results showed that both the gene expression and FT-IR spectra were affected by the variations in atmosphere, temperature and time, but that the effect was different for the two types of data. When the DNA microarray data and FT-IR spectra were linked together by PLSR, covariation due to temperature was seen. Both specific genes and ranges in the FT-IR spectra that were connected to the variation in temperature were detected. Some of these are possibly connected to properties of the cell wall of the bacteria. The results in this study show the potential of PLSR for investigation of covariance structures in biological data. By doing this, valuable information about the biological system can be detected and interpreted. It was also shown that the use of FT-IR spectroscopy provided important information about the stress responses in the bacteria, information that was not detected from the DNA microarray data.     

Integrating fluctuating nitrate uptake and assimilation to robust homeostasis

Nitrate is an important nitrogen source used by plants. Despite of the considerable variation in the amount of soil nitrate, plants keep cytosolic nitrate at a homeostatic controlled level. Here we describe a set of homeostatic controller motifs and their interaction that can maintain robust cytosolic nitrate homeostasis at fluctuating external nitrate concentrations and nitrate assimilation levels. The controller motifs are divided into two functional classes termed as inflow and outflow controllers. In the presence of high amounts of environmental nitrate, the function of outflow controllers is associated to efflux mechanisms removing excess of nitrate from the cytosol that is taken up by low-affinity transporter systems (LATS). Inflow controllers on the other hand maintain homeostasis in the presence of a high demand of nitrate by the cell relative to the amount of available environmental nitrate. This is achieved by either remobilizing nitrate from a vacuolar store, or by taking up nitrate by means of high-affinity transporter systems (HATS). By combining inflow and outflow controllers we demonstrate how nitrate uptake, assimilation, storage and efflux are integrated to a regulatory network that maintains cytosolic nitrate homeostasis at changing environmental conditions.     

Assessment of fraction of hypoxic cells in human tumor xenografts with necrotic regions by dynamic contrast-enhanced MRI

The potential usefulness of gadopentetate dimeglumine (Gd-DTPA)-based dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) for assessing hypoxia in tumors with significant necrosis was investigated. Small (100-350 mm(3)) and large (500-1000 mm(3)) D-12 and U-25 tumors were subjected to DCE-MRI, measurement of the fraction of necrotic tissue, and measurement of the fraction of radiobiologically hypoxic cells. Images of E.F (E is the initial extraction fraction of Gd-DTPA and F is perfusion) and lambda (lambda is proportional to extracellular volume fraction) were produced by subjecting the DCE-MRI data to Kety analysis. Necrotic tissue could be identified in lambda images but not in E.F images of the tumors. Most voxels in viable tissue showed lambda values of 0.15-0.70, whereas the lambda values of most voxels in necrotic tissue were either <0.15 or >0.70. The E.F and lambda frequency distributions of the viable tissue, but not the E.F and lambda frequency distributions of the whole tissue, were consistent with the observation that the four groups of tumors showed similar fractions of radiobiologically hypoxic cells. E.F and lambda images may thus provide useful information on the extent of hypoxia in tumors provided that voxels in necrotic tumor regions are identified and excluded from the images.     

Improved quantification of prostaglandins in biological samples by optimizing simultaneously the relationship eicosanoid/internal standard and using liquid chromatography tandem mass spectrometry

Although a wide variety of articles on quantification of eicosanoids by using internal standards are published every year, little has been done on how much internal standard should be added. This article demonstrates that the application of experimental design enables estimating the interaction eicosanoid/internal standard and to select confidently an optimal amount of internal standard and a response factor (RF) for the analysis of eicosanoids in a high number of samples, where the amount of sample is limited and the unknown levels of eicosanoids are spanned in a wide range of concentrations. The results revealed that the interaction eicosanoid/internal standard is an important factor that affects the validity of the RF and subsequently the accuracy of the analysis.     

Allele Interaction – Single Locus Genetics Meets Regulatory Biology

Background

Since the dawn of genetics, additive and dominant gene action in diploids have been defined by comparison of heterozygote and homozygote phenotypes. However, these definitions provide little insight into the underlying intralocus allelic functional dependency and thus cannot serve directly as a mediator between genetics theory and regulatory biology, a link that is sorely needed.

Methodology/Principal Findings

We provide such a link by distinguishing between positive, negative and zero allele interaction at the genotype level. First, these distinctions disclose that a biallelic locus can display 18 qualitatively different allele interaction sign motifs (triplets of +, – and 0). Second, we show that for a single locus, Mendelian dominance is not related to heterozygote allele interaction alone, but is actually a function of the degrees of allele interaction in all the three genotypes. Third, we demonstrate how the allele interaction in each genotype is directly quantifiable in gene regulatory models, and that there is a unique, one-to-one correspondence between the sign of autoregulatory feedback loops and the sign of the allele interactions.

Conclusion/Significance

The concept of allele interaction refines single locus genetics substantially, and it provides a direct link between classical models of gene action and gene regulatory biology. Together with available empirical data, our results indicate that allele interaction can be exploited experimentally to identify and explain intricate intra- and inter-locus feedback relationships in eukaryotes.     

X-Ray crystallographic, NMR and antimicrobial activity studies of magnesium complexes of fluoroquinolones - racemic ofloxacin and its S-form, levofloxacin

The magnesium complexes of racemic ofloxacin (oflo) and its pure S-form levofloxacin (S-oflo) have been studied by X-ray crystallography and NMR spectroscopy. Two compounds, [Mg(R-oflo)(S-oflo)(H(2)O)(2)].2H(2)O (1) and [Mg(S-oflo)(2)(H(2)O)(2)].2H(2)O (2), respectively, have been prepared by hydrothermal reactions and their crystal structures have been determined. In both structures the anionic fluoroquinolone ligands are coordinated through the keto and carboxylate oxygens forming 1:2 Mg:oflo complexes. The two structures are practically identical except for the orientation of one of the oxazine methyl groups at the chiral center of 2 which was found in equatorial position, the other oxazine methyl groups in 1 and 2 being axial. This difference affects the stacking pattern of quinolone molecules in the cell. (1)H NMR chemical shift data and Mn(II) paramagnetic line broadening measurements on the free ofloxacin suggest that the coordination of the ligands in solution involves the keto and carboxylate oxygens. However, it is not possible to decide whether the complexes in aqueous solution have 1:1 or 1:2 stoichiometry. The methylated piperazine nitrogen does not interact with the metal ion. Magnesium-quinolone interaction is discussed in relation to the biological activity of quinolones. The antimicrobial activity of the complexes against various microorganisms was tested and it was established that their activity is similar to that of free quinolone drugs.     

Estimation of hooded seal (<Emphasis Type="Italic">Cystophora cristata</Emphasis>) pup production in the Greenland Sea pack ice during the 2005 whelping season

In March 2005 aerial surveys were carried out to estimate pup production of Greenland Sea hooded seals, Cystophora cristata in three whelping patches using photographic strip transects. The majority of whelping occurred 17–23 March and surveys were carried out on 24 March. The temporal distribution of births and estimated proportion of solitary bluebacks present within the survey area were used to correct the abundance estimates. The total estimated pup production (15,250 pups, s.e. = 3,470; CV = 22.8%) was lower than an estimate from a comparable study in 1997. The current estimate is considered to be negatively biased, as it is uncorrected for pups born outside the whelping concentrations. Population modeling suggests a substantial decline with subsequent stabilization at a lower level for Greenland Sea hooded seals in recent decades. It is suggested that future research on the population explore possible factors (e.g. reduced fertility, predation, food shortage, disease, reductions in ice cover) that may have contributed to the apparent prevailing low abundance.     

Geosmin production in the cyanobacterium Oscillatoria brevis

The cyanobacterium Oscillatoria brevis (Kütz.) Gom., strain NIVA CYA 7, was used to investigate how geosmin production is related to the synthesis of chlorophyll a, phycobiliproteins and β-carotene under nitrogen (NH ₄ ⁺ ) and light limiting conditions. Chemostat samples were used to inoculate batch cultures that were treated with inhibitors of isoprenoid synthesis, norflurazon and dimethazone, and gabaculine that inhibits tetrapyrrole synthesis. Dimethazone decreased and norflurazon increased geosmin production under light limited conditions, as was expected due to their sites of action in the isoprenoid pathway. This effect was not so pronounced in nitrogen limited cultures due to the additional effect of increasing nitrogen deficiency during the experimental period. Norflurazon was the only inhibitor that uncoupled geosmin production completely from β-carotene formation which indicates a strikt coupling between geosmin and β-carotene biosynthesis. From the observed increase of geosmin production relative to pigment synthesis after norflurazon treatment it was suggested that isoprenoid precursors are directed to geosmin synthesis when the demand for pigment precursors is very low. Within the framework of this study the data strongly support the hypothesis of geosmin formation via the isoprenoid pathway in Oscillatoria brevis as was found for actinomycetes. This research was performed at the Department of Microbiology, University of Amsterdam, with financial support provided by the royal Norwegian Ministry of Foreign Affairs and the Royal Norwegian Council for Scientific and Industrial Research     

Casein SNP in Norwegian goats: additive and dominance effects on milk composition and quality

Background

The four casein proteins in goat milk are encoded by four closely linked casein loci (CSN1S1, CSN2, CSN1S2 and CSN3) within 250 kb on caprine chromosome 6. A deletion in exon 12 of CSN1S1, so far reported only in Norwegian goats, has been found at high frequency (0.73). Such a high frequency is difficult to explain because the national breeding goal selects against the variant''s effect.

Methods

In this study, 575 goats were genotyped for 38 Single Nucleotide Polymorphisms (SNP) located within the four casein genes. Milk production records of these goats were obtained from the Norwegian Dairy Goat Control. Test-day mixed models with additive and dominance fixed effects of single SNP were fitted in a model including polygenic effects.

Results

Significant additive effects of single SNP within CSN1S1 and CSN3 were found for fat % and protein %, milk yield and milk taste. The allele with the deletion showed additive and dominance effects on protein % and fat %, and overdominance effects on milk quantity (kg) and lactose %. At its current frequency, the observed dominance (overdominance) effects of the deletion allele reduced its substitution effect (and additive genetic variance available for selection) in the population substantially.

Conclusions

The selection pressure of conventional breeding on the allele with the deletion is limited due to the observed dominance (overdominance) effects. Inclusion of molecular information in the national breeding scheme will reduce the frequency of this deletion in the population.