Enzyme inhibitors to increase poly-3-hydroxybutyrate production by transgenic tobacco

Chemical regulation of secondary-metabolite synthesis was investigated through the improvement of poly-3-hydroxybutyrate (PHB) production in transgenic tobacco plants by the use of enzyme inhibitors. Two tobacco lines, BC3 and rCAB8, that produce PHB in both the cytosol and plastids were used. An acetyl-CoA carboxylase inhibitor, D-(+)-Quizalofop-ethyl, increased PHB accumulation in both lines 2-fold. The accumulation rate of plastidial PHB in the rCAB8 line was 2.5-fold higher than that of cytosolic PHB in the BC3 line. A specific inhibitor of 3-hydroxy-3-methylglutaryl-CoA reductase, mevastatin, also increased PHB accumulation but only in the BC3 line. These results indicated that chemical regulation of the native metabolic flows by the specific enzyme inhibitors increased secondary-metabolite production in the transgenic tobacco plants we used.     

Telemetered cephalopod energetics: swimming, soaring, and blimping

Cephalopods are uniquely suited to field energetic studies.Their hollow mantles that pump water for respiration and jettingalso can accommodate differential transducer-transmitters. Thesetransmitters indicate pressure-flow power output, which canbe calibrated against oxygen consumption by swim-tunnel respirometry.Radio-acoustic positioning telemetry (RAPT) records pressure-flowpower and animal movements with meter accuracy in nature. Despiteinherent inefficiencies, jetting is the primary mode of locomotionfor both primitive nautilus and powerful, migratory oceanicsquids. In between, large-finned squid and cuttlefish mix jettingwith fin undulation in complex gaits that increase locomotorefficiency. Our studies show that the complex nervous systemscephalopods evolved to control mixed gaits are also sensitiveto flow and density fields in nature and that they use theseto further reduce locomotion costs. Buoyed up by evacuated shells,nautilus and cuttlefish live in boundary layers and navigatecheaply through them like balloonists. Large-finned, negativelybuoyant squid soar like eagles in rising currents, but losecontrol in currents above one body length per second. Many muscularsquids have life histories linked to current systems. Neutrallybuoyant ammoniacal cephalopods in the mesopelagic are a limitingcase in need of study. The small density differential betweenseawater and isotonic ammonium chloride trebles their volume,making them blimp-like with very low power densities. Some specieslive entirely in this restricted habitat, but most become ammoniacallate in ontogeny, as they approach semelparous reproduction.Ammonium retained for buoyancy as carbon is terminally mobilizedfrom muscle protein for gametes and energy, compensates forlost muscle power.     

Molecular scanner experiment with human plasma: improving protein identification by using intensity distributions of matching peptide masses

The development of high throughput utilities to identify proteins is a major challenge in present research in the field of proteomics. One such utility, the molecular scanner, uses proteins separated by two-dimensional polyacrylamide gel electrophoresis that are digested in the gel and during transfer onto a collecting membrane. After adding a matrix, the membrane is inserted into a matrix-assisted laser desorption/ionization-time of flight mass spectrometer and a peptide mass fingerprint (PMF) is measured for every scanned site. Since the spacing between scanned sites is much smaller than the size of the most abundant protein spots, there is a certain redundancy in the data that was used in an earlier experiment with Escherichia coli [1] to improve mass calibration and PMF identification results. It was observed that the signal intensity of a peptide mass as a function of the position on the membrane showed similar patterns if peptides stemmed from the same protein. Taking account of these similarities a clustering algorithm was used to find lists of experimental masses with similar intensity distributions, which provided clearer identification of the corresponding proteins. Here, these methods are applied to a human plasma scan, where proteins were highly modified and less separated. The presence of very abundant proteins like albumin and immunoglobulins added another difficulty. The calibration of the initial PMFs was not satisfactory and masses had to be recalibrated. After discarding chemical noise, the membrane was partitioned into regions and for each region protein identification was carried out separately. A new scoring method was used, where the PMF score was multiplied by a factor that measures the similarity of matching peptides. This method proved to be more robust than the method developed in [1] if the region where a protein was found had an extended, nonspherical shape and strong overlap with regions of other proteins. Many proteins annotated on the SWISS-2D PAGE human plasma master gel could be clearly identified and many interesting properties were observed.     

Different patterns of interleukin-1alpha and interleukin-1beta expression in organs of normal young and old mice

The different physiological roles of interleukin-1alpha (IL-1alpha) and interleukin-1beta (IL-1beta) are not well understood, especially when considering the apparent overlap and redundancy of the two IL-1 molecules. Characterization of IL-1alpha and IL-1beta expression was performed in this study in organs from young and old mice, using immunohistochemistry and ELISA (enzyme-linked immunosorbent assay). The results indicate that organ IL-1alpha and IL-1beta display different patterns of expression: IL-1alpha is manifested more prominently in lymphoreticular organs (lungs, small intestine, spleen, liver), while IL-1beta is more evident in highly specialized and more vulnerable organs, which do not play a leading role in defense against infections and intoxication (heart, brain, skeletal muscle, kidney). This differential expression is more accentuated in old mice, possibly pointing to the special relevance of these cytokines to organ homeostasis in old age. These findings may shed new light on the physiological functions of IL-1alpha and IL-1beta, and may also lead to the development of improved therapeutic approaches, based on the specific manipulation of these cytokines.     

ShiftDetector: detection of shift mutations

MOTIVATION: Sequencing of a bi-allelic PCR product, which contains an allele with a deletion/insertion mutation results in a superimposed tracefile following the site of this shift mutation. A trace file of this type hampers the use of current computer programs for base calling. ShiftDetector analyses a sequencing trace file in order to discover if it is a superimposed sequence of two molecules that differ in a shift mutation of 1 to 25 bases. The program calculates a probability score for the existence of such a shift and reconstructs the sequence of the original molecule. AVAILABILITY: ShiftDetector is available from http://cowry.agri.huji.ac.il     

Discovering statistically significant biclusters in gene expression data

In gene expression data, a bicluster is a subset of the genes exhibiting consistent patterns over a subset of the conditions. We propose a new method to detect significant biclusters in large expression datasets. Our approach is graph theoretic coupled with statistical modelling of the data. Under plausible assumptions, our algorithm is polynomial and is guaranteed to find the most significant biclusters. We tested our method on a collection of yeast expression profiles and on a human cancer dataset. Cross validation results show high specificity in assigning function to genes based on their biclusters, and we are able to annotate in this way 196 uncharacterized yeast genes. We also demonstrate how the biclusters lead to detecting new concrete biological associations. In cancer data we are able to detect and relate finer tissue types than was previously possible. We also show that the method outperforms the biclustering algorithm of Cheng and Church (2000).     

Mental competence and elderly care

Competence or decision-making capacity are concepts, which are far from clear-cut. The normative consequences of an incapacity-judgment in elderly care require a balanced approach, which takes into consideration the risk of misuse in the context of the justification of interventions that are applied without consent of the patient. In this article a number of central issues in the debate on decision-making capacity are discussed. The different criteria, which are proposed to assess capacity, are discussed as well as the development of instruments in order to objectify capacity. It is concluded that the common approach towards decision-making capacity, which is strongly cognitive-oriented, is inadequate. A number of relevant aspects remain underexposed within this approach. A broader approach is explored, which takes into account the role of emotions, identity, and dialogue and deliberation. The different approaches are illustrated by way of a case example.