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PubMed2002

Mode-doubling and tripling in reaction-diffusion patterns on growing domains: a piecewise linear model.

Reaction-diffusion equations are ubiquitous as models of biological pattern formation. In a recent paper we have shown that incorporation of domain growth in a reaction-diffusion model generates a sequence of quasi-steady patterns and can provide a mechanism for increased reliability of pattern selection. In this paper we analyse the model to examine the transitions between patterns in the sequence. Introducing a piecewise linear approximation we find closed form approximate solutions for steady-state patterns by exploiting a small parameter, the ratio of diffusivities, in a singular perturbation expansion. We consider the existence of these steady-state solutions as a parameter related to the domain length is varied and predict the point at which the solution ceases to exist, which we identify with the onset of transition between patterns for the sequence generated on the growing domain. Applying these results to the model in one spatial dimension we are able to predict the mechanism and timing of transitions between quasi-steady patterns in the sequence. We also highlight a novel sequence behaviour, mode-tripling, which is a consequence of a symmetry in the reaction term of the reaction-diffusion system.

Animals↗

PubMed2003

Sliding window dual gradient echo (SW-dGRE): T1 and proton resonance frequency (PRF) calibration for temperature imaging in polyacrylamide gel.

The aim of the work is to evaluate a magnetic resonance imaging (MRI) thermometry sequence suitable for targeting of focused ultrasound (FUS) when used in vascular occlusion studies. A sliding window dual gradient echo (SW-dGRE) sequence was used. This sequence has the capability of monitoring both T1 relaxation and phase changes, which vary with temperature. Preliminary work involved quantification of the changes in T1 relaxation time with temperature and obtaining the PRF shift coefficient in polyacrylamide gel as it underwent an exothermic reaction during polymerization (avoiding the use of an external heat source). Temperature changes were visualized using thermal maps acquired with the sequence. For FUS guidance a thermal imaging technique is required with a temporal resolution <5 s, a spatial resolution of approximately 1 mm and a temperature resolution of approximately 5 degrees C. The sequence was optimized to improve the CNR (contrast to noise ratio) and SNR (signal to noise ratio) in the phase and magnitude images respectively. The PRF coefficient obtained for the polyacrylamide gel was -9.98 +/- 0.24 ppb degrees C(-1), whilst deltaT1 and temperature change were related by a proportionality factor, the T1 temperature coefficient, of 102.3 +/- 2.9 ms degrees C(-1). The sequence produces an image at every 1.4 s interval. In both magnitude and phase data, the in-plane resolution is +/- 1.2 mm and the temperature resolution is approximately 2 degrees C. The advantage of this sequence is that the temperature obtained from the magnitude data can be confirmed independently using the phase data and vice versa. Thus the sequence can essentially be crosschecked.

Acrylic Resins↗

PubMed2004

Microsatellite and mitochondrial DNA analyses of Atlantic bluefin tuna (Thunnus thynnus thynnus) population structure in the Mediterranean Sea.

Genetic variation was surveyed at nine microsatellite loci and the mitochondrial control region (868 bp) to test for the presence of genetic stock structure in young-of-the-year Atlantic bluefin tuna (Thunnus thynnus thynnus) from the Mediterranean Sea. Bluefin tuna were sampled over a period of 5 years from the Balearic and Tyrrhenian seas in the western basin of the Mediterranean Sea, and from the southern Ionian Sea in the eastern basin of the Mediterranean Sea. Analyses of multilocus microsatellite genotypes and mitochondrial control region sequences revealed no significant heterogeneity among collections taken from the same location in different years; however, significant spatial genetic heterogeneity was observed across all samples for both microsatellite markers and mitochondrial control region sequences (FST=0.0023, P=0.038 and PhiST=0.0233, P=0.000, respectively). Significant genetic differentiation between the Tyrrhenian and Ionian collections was found for both microsatellite and mitochondrial markers (FST=0.0087, P=0.015 and PhiST=0.0367, P=0.030, respectively). These results suggest the possibility of a genetically discrete population in the eastern basin of the Mediterranean Sea.

Animals↗

PubMed1995

Role of RNA secondary structure of the iron-responsive element in translational regulation of ferritin synthesis.

Iron regulates synthesis of the iron storage protein ferritin at the translational level through interaction between a stem-loop structure, the iron-responsive element (IRE), located in the 5'-untranslated region (5'-UTR) of ferritin mRNAs, and a protein, the iron regulatory protein (IRP). The role of IRE secondary structure in translational regulation of ferritin synthesis was explored by introducing ferritin constructs containing mutations in the IRE into Rat-2 fibroblasts. Our in vivo studies demonstrate that size and sequence of the loop within the IRE and the distance and/or spatial relationship of this loop to the bulged nucleotide region closest to the loop must be preserved in order to observe iron-dependent translation of ferritin mRNA. In contrast, changes in nucleotide sequence of the upper stem can be introduced without affecting translational regulation in vivo, as long as a stem can be formed. Our in vivo results suggest that only a very small variation in the affinity of interaction of IRP with IRE can be tolerated in order to maintain iron-dependent regulation of translation.

Animals↗

PubMed1998

Molecular cloning of the chick Nau gene and analysis of its expression patterns during neurogenesis.

We have previously identified a novel gene, Nau (Neuron-specific gene, which has a number of AU-rich RNA instability motifs), from a quail spinal cord cDNA library. Nau expression was observed mainly in post-mitotic neuroblasts, especially in the stage of axonogenesis. In this study, we cloned its chick orthologue (cNau, 5069 bp), which encodes 369 amino acid residues. cNau's open reading frame has three highly conserved regions with rat STOP (stable tubule-only polypeptide). cNau may be an orthologue of STOP gene considering its uniqueness in zoo blot analysis and its sequence similarity. Both the genetic structure and temporo-spatial expression patterns indicate that the cNau gene may be related to axonogenesis. In the hindbrain, cNau mRNA was accumulated in rhombomere boundary regions and in the rhombomere 2, 4 and 6. These observations suggest a specific role of cNau in the hindbrain or the difference in development between the hindbrain and spinal cord.

Amino Acid Sequence↗

PubMed2003

Conserved residue clustering and protein structure prediction.

Protein residues that are critical for structure and function are expected to be conserved throughout evolution. Here, we investigate the extent to which these conserved residues are clustered in three-dimensional protein structures. In 92% of the proteins in a data set of 79 proteins, the most conserved positions in multiple sequence alignments are significantly more clustered than randomly selected sets of positions. The comparison to random subsets is not necessarily appropriate, however, because the signal could be the result of differences in the amino acid composition of sets of conserved residues compared to random subsets (hydrophobic residues tend to be close together in the protein core), or differences in sequence separation of the residues in the different sets. In order to overcome these limits, we compare the degree of clustering of the conserved positions on the native structure and on alternative conformations generated by the de novo structure prediction method Rosetta. For 65% of the 79 proteins, the conserved residues are significantly more clustered in the native structure than in the alternative conformations, indicating that the clustering of conserved residues in protein structures goes beyond that expected purely from sequence locality and composition effects. The differences in the spatial distribution of conserved residues can be utilized in de novo protein structure prediction: We find that for 79% of the proteins, selection of the Rosetta generated conformations with the greatest clustering of the conserved residues significantly enriches the fraction of close-to-native structures.

Acyl Coenzyme A↗

PubMedDate not supplied

Peptide/benzodiazepine hybrids as ligands of CCK(A) and CCK(B) receptors.

The (neuro)hormones gastrin and cholecystokinin (CCK) share a common C-terminal tetrapeptide amide sequence that has been recognized as the message portion while the N-terminal extensions are responsible for the CCK(A) and CCK(B) receptor subtype selectivity and avidity. 1,4-Benzodiazepine derivatives are potent and selective antagonists of these receptors, and according to comparative molecular field analysis, the structures of these nonpeptidic compounds could well mimic the message sequence of the peptide agonists at least in terms of spatial array of the aromatic residues. Docking of a larger series of low molecular weight nonpeptide antagonists to a homology modeling derived CCK(B) receptor structure revealed a consensus binding mode that is further validated by data from site-directed mutagenesis studies of the receptors. Whether this putative binding pocket of the nonpeptide antagonists is identical to that of the message portion of the peptide agonists, or whether it is distinct and spatially separated, or overlapping, but with distinct interaction sites, is still object of debate. Using a 1,4-benzodiazepine core amino-functionalized at the C3 position, related tryptophanyl derivatives were synthesized as mimics of the tetrapeptide and subsequently extended N-terminally with gastrin and CCK address sequences. All hybrid constructs were recognized as antagonists by the CCK(A) and CCK(B) receptors, but their address portions were incapable of enhancing in significant manner selectivity and avidity. Consequently, the binding of the peptide/benzodiazepine hybrids has to be dictated mainly by the benzodiazepine moiety, which apparently prevents optimal interactions of the address peptides with extracellular receptor subdomains. These findings would strongly support the view of distinct binding sites for the message portion of the peptide agonists and the benzodiazepine-based nonpeptide antagonists.

Benzodiazepines↗

PubMed1997

Measures of spatial memory and routes of learning.

Spatial memory was investigated in two experiments by direct methods. Methods included scaling of distances, estimation of bearings, and positioning of objects. Participants learned small-scale configurations under different orders of presentation. In Exp. 1, routes included a shortest path, i.e., a traveling salesman solution, a random sequence, and a path that maximized distances. In Exp. 2, spatial and temporal distances varied independently. It was analyzed whether the different methods yielded the same information. For bearing estimates a new scaling procedure was developed. Computations resulted in two-dimensional Euclidean solutions in close correspondence with the stimulus configuration. In addition, solutions showed an effect of temporal conditions of learning.

Attention↗

PubMed2005

High-resolution real-time spiral MRI for guiding vascular interventions in a rabbit model at 1.5 T.

PURPOSE: To study the feasibility of a combined high spatial and temporal resolution real-time spiral MRI sequence for guiding coronary-sized vascular interventions. MATERIALS AND METHODS: Eight New Zealand White rabbits (four normal and four with a surgically-created stenosis in the abdominal aorta) were studied. A real-time interactive spiral MRI sequence combining 1.1 x 1.1 mm(2) in-plane resolution and 189-msec total image acquisition time was used to image all phases of an interventional procedure (i.e., guidewire placement, balloon angioplasty, and stenting) in the rabbit aorta using coronary-sized devices on a 1.5 T MRI system. RESULTS: Real-time spiral MRI identified all rabbit aortic stenoses and provided high-temporal-resolution visualization of guide-wires crossing the stenoses in all animals. Angioplasty balloon dilatation and deployment of coronary-sized copper stents in the rabbit aorta were also successfully imaged by real-time spiral MRI. CONCLUSION: Combining high spatial and temporal resolution with spiral MRI allows real-time MR-guided vascular intervention using coronary-sized devices in a rabbit model. This is a promising approach for guiding coronary interventions.

Animals↗

PubMed2000

Chondrocyte-specific enhancer regions in the COMP gene.

The molecular events governing the differentiation of mesenchymal cells into chondrocytes and the expression of cartilage marker genes are poorly understood. Cartilage oligomeric matrix protein is a noncollagenous extracellular matrix protein with a relatively cartilage-specific spatial and temporal expression pattern. To understand the mechanisms controlling chondrocyte-specific expression of cartilage oligomeric matrix protein, we cloned 1.9 kb of the 5' flanking promoter sequence of the murine cartilage oligomeric matrix protein gene and identified two spatially distant cartilage-specific enhancer regions by deletion analysis. One element is situated proximally (proximal positive element: -125 to -75) and a second region is located distally (distal positive region: -1925 to -592) relative to the transcription start site. Interestingly, nucleotides within the proximal positive element are conserved between the mouse and human promoters and resemble consensus sites for the binding of members of the high mobility group class of transcription factors. Defining cartilage-specific regulatory elements in the cartilage oligomeric matrix protein promoter may provide useful molecular probes for identifying transcription factors that control acquisition of the chondrocytic phenotype.

Animals↗

PubMed2002

Extraction of functional binding sites from unique regulatory regions: the Drosophila early developmental enhancers.

The early developmental enhancers of Drosophila melanogaster comprise one of the most sophisticated regulatory systems in higher eukaryotes. An elaborate code in their DNA sequence translates both maternal and early embryonic regulatory signals into spatial distribution of transcription factors. One of the most striking features of this code is the redundancy of binding sites for these transcription factors (BSTF). Using this redundancy, we explored the possibility of predicting functional binding sites in a single enhancer region without any prior consensus/matrix description or evolutionary sequence comparisons. We developed a conceptually simple algorithm, Scanseq, that employs an original statistical evaluation for identifying the most redundant motifs and locates the position of potential BSTF in a given regulatory region. To estimate the biological relevance of our predictions, we built thorough literature-based annotations for the best-known Drosophila developmental enhancers and we generated detailed distribution maps for the most robust binding sites. The high statistical correlation between the location of BSTF in these experiment-based maps and the location predicted in silico by Scanseq confirmed the relevance of our approach. We also discuss the definition of true binding sites and the possible biological principles that govern patterning of regulatory regions and the distribution of transcriptional signals.

Algorithms↗

PubMed1996

Spectroscopic probe for the detection of local DNA bending at an AAA triplet.

Spectroscopic evidence of a DNA bend in solution is presented by analyzing model 15-mer duplexes spin-labeled with the five-atom-tethered nitroxide DUAP located in the major groove. Three 15-mers containing AATT with DUAP enzymatically incorporated into three different positions yielded nearly identical line shapes while a fourth 15-mer containing AAATT produced an EPR spectrum with significant additional line broadening. These results are interpreted according to the dynamic cylinder model where the DNA dynamics are decoupled into overall and internal contributions. It is shown that the AAATT sequence induces a change in the internal dynamics characterized by local ordering of DUAP. The increase in ordering evident in 15-mers containing AAATT rather than AATT suggests that the former sequence gives rise to a bend toward the major groove resulting in spatial restriction of the probe.

Base Sequence↗

PubMed1986

Representation of DNA sequences by use of helical wheels to identify pattern formation with protein binding potential.

Similar to the protein segments with helical potential the three-dimensional structures of DNA sequences can be represented by a two-dimensional projection along the axis of the double helix. We will call this projection according to Schiffer and Edmundson "helical wheel". In the first place the wheels are employed here as graphical restatements of known sequences which represent binding sites of restriction-modification enzymes. Specific recognition of these sites has to be based on sequence related multi contact interactions of weak bonds. Because of the spatial and directional characteristics of H-bonds we will utilize the pattern formation of H-bond donor--and acceptor groups protruding into the major groove of the helix to identify the particular sequences. The characteristics of these patterns can be readily visualized and compared by examination of these wheels.

Amino Acid Sequence↗

PubMed2003

Spatial analysis of archaeal community structure in grassland soil.

The complex structure of soil and the heterogeneity of resources available to microorganisms have implications for sampling regimens when the structure and diversity of microbial communities are analyzed. To assess the heterogeneity in community structure, archaeal communities, which typically contain sequences belonging to the nonthermophilic Crenarchaeota, were examined at two contrasting spatial scales by using PCR-denaturing gradient gel electrophoresis (DGGE) analysis followed by unweighted pair group method with arithmetic mean analysis of 16S rRNA- and ribosomal DNA-derived profiles. A macroscale analysis was carried out with soil cores taken at 2-m intervals along triplicate 8-m transects from both managed (improved) and natural (unimproved) grassland rhizosphere soils. A microscale analysis was carried out with a single soil core by assessing the effects of both sample size (10, 1, and 0.1 g) and distance between samples. The much reduced complexity of archaeal profiles compared to the complexity typical of the bacterial community facilitated visual comparison of profiles based on band presence and revealed different levels of heterogeneity between sets of samples. At the macroscale level, heterogeneity over the transect could not be related to grassland type. Substantial heterogeneity was observed across both improved and unimproved transects, except for one improved transect that exhibited substantial homogeneity, so that profiles for a single core were largely representative of the entire transect. At the smaller scale, the heterogeneity of the archaeal community structure varied with sample size within a single 8- by 8-cm core. The archaeal DGGE profiles for replicate 10-g soil samples were similar, while those for 1-g samples and 0.1-g samples showed greater heterogeneity. In addition, there was no relationship between the archaeal profiles and the distance between 1- or 0.1-g samples, although relationships between community structure and distance of separation may occur at a smaller scale. Our findings demonstrate the care required when workers attempt to obtain a representative picture of microbial community structure in the soil environment.

Crenarchaeota↗

PubMed2004

Bilateral lesion of the cerebellar-dentate nucleus impairs egocentric sequential learning but not egocentric navigation in the rat.

The involvement of the cerebellum in procedural learning is demonstrated in visuomotor-sequence tasks, as lesion of this area impedes the acquisition of new sequences. Likewise, the lateral cerebellum appears to be involved in the acquisition of new sequences, but not in the execution of learned sequences. In contrast, the dentate nucleus participates only in the execution of learned visuomotor sequences. In previous studies, disruption of the procedural elements of spatial navigation following cerebellar or dentate lesions has been reported. However, as praxic strategies (egocentric learning) are included in the procedural elements of the navigation, the participation of the cerebellar-dentate nucleus in egocentric procedural learning processes has not been evaluated. Therefore, using colchicine, bilateral lesions were made in the cerebellar-dentate nucleus of Sprague-Dawley rats, and these rats were given two tasks: egocentric-based motor sequence learning in the radial maze and egocentric navigation in the Morris water maze. The lesioned rats were unable to use the sequential information in the short term and showed delayed long-term acquisition, which was probably due to the inability to detect the sequence. No effects on the egocentric navigation task were observed. Our results indicate that the cerebellar-dentate nucleus is involved in the detection of egocentric sequential information but not in the use of this information in the navigation process. Further, they show differential involvement of the cerebellar-dentate nucleus in the execution of learned visuomotor sequences, as the dentate lesion disrupted the acquisition of new egocentric-motor-based sequences.

Analysis of Variance↗

PubMed2006

A specific helical orientation underlies the functional contribution of the activin responsive unit to transcriptional activity of the murine gonadotropin-releasing hormone receptor gene promoter.

Activin responsiveness of the murine GnRH receptor (GnRHR) gene promoter requires two spatially distinct regulatory elements termed the GnRH receptor activating sequence or GRAS and the downstream activin regulatory element or DARE. While GRAS interacts with multiple transcription factors, DARE activity requires tandem homeodomain binding motifs (TAAT) and displays specific binding to the LIM homeodomain protein LHX3. Herein, we find that both the murine GnRHR gene promoter and DARE fused to a minimal heterologous promoter are responsive to LHX3 overexpression. A dominant-repressor of LHX3 attenuates transcriptional activity of the murine GnRHR gene promoter but had no impact on activin responsiveness. Thus, LHX3 would not appear to be the protein mediating activin responsiveness of this promoter. Within DARE itself, the tandem TAAT motifs are separated by 4 bp. Although this arrangement differs from the prototypical P2 or P3 binding sites characterized for paired-like homeodomain proteins and from the directly abutting TAAT motifs found for LHX3, a LIM-class homeodomain protein, we find that separation of the TAAT sites by 5 and 10 bp decreases GnRHR promoter activity to a level similar to promoters containing loss of function mutations in either the proximal or distal TAAT motif. Thus, the juxtaposition of the TAAT sites is critical for the functional activity of DARE. That activin responsiveness of the GnRHR promoter requires both GRAS and DARE suggests that these elements may be both functionally and structurally coupled. As to the latter, GRAS and DARE are separated by 20 bp, thus placing the elements on the same side of the helical backbone. To determine if this spatial organization is functionally relevant, multiples of 5 bp were inserted or deleted between GRAS and DARE. Any insertion or deletion that resulted in a half-turn alteration in the helical positioning between the two elements reduced promoter activity. Thus, an important spatial relationship underlies functional cooperation between GRAS and DARE and the emergence of a complex activin responsive unit (ARU) within the mouse GnRHR promoter.

Activins↗

PubMed1995

A novel regulatory region is required for trophoblast-specific transcription in transgenic mice.

The trophectoderm epithelium of the blastocyst is the first tissue to differentiate in the mammalian embryo and gives rise to all cells of the trophoblast lineage. These cells form the majority of the fetal component of the placenta, which is critical for intrauterine development. Our interest in the molecular determinants of trophoblast lineage development has led us to undertake an analysis of the regulatory region of a trophoblast-specific gene termed 4311. The transcription start site was mapped by both primer extension and RNase protection analyses. We show that cloned 4311 upstream sequences are able to confer the correct temporal and spatial expression pattern of 4311 on a reporter gene in transgenic mice. In addition, we have characterized a 340-bp region, 3.7 kb upstream of the transcription start, that is sufficient for the specific expression of lacZ in cells of the developing spongiotrophoblast in transgenic mice. Delineation of a diploid trophoblast-specific regulatory region will facilitate the identification of factors that may regulate the development of this lineage.

Animals↗

PubMed1983

The spatial distribution of fixed mutations within genes coding for proteins.

We have examined the extensive amino acid sequence data now available for five protein families - the alpha crystallin A chain, myoglobin, alpha and beta hemoglobin, and the cytochromes c - with the goal of estimating the true spatial distribution of base substitutions within genes that code for proteins. In every case the commonly used Poisson density failed to even approximate the experimental pattern of base substitution. For the 87 species of beta hemoglobin examined, for example, the probability that the observed results were from a Poisson process was the minuscule 10(-44). Analogous results were obtained for the other functional families. All the data were reasonably, but not perfectly, described by the negative binomial density. In particular, most of the data were described by one of the very simple limiting forms of this density, the geometric density. The implications of this for evolutionary inference are discussed. It is evident that most estimates of total base substitutions between genes are badly in need of revision.

Amino Acid Sequence↗