The identification of cutin synthase: formation of the plant polyester cutin

A hydrophobic cuticle consisting of waxes and the polyester cutin covers the aerial epidermis of all land plants, providing essential protection from desiccation and other stresses. We have determined the enzymatic basis of cutin polymerization through characterization of a tomato extracellular acyltransferase, CD1, and its substrate, 2-mono(10,16-dihydroxyhexadecanoyl)glycerol. CD1 has in vitro polyester synthesis activity and is required for cutin accumulation in vivo, indicating that it is a cutin synthase.     

Discovering Relations Between Mind,Brain, and Mental Disorders Using Topic Mapping

Neuroimaging research has largely focused on the identification of associations between brain activation and specific mental functions. Here we show that data mining techniques applied to a large database of neuroimaging results can be used to identify the conceptual structure of mental functions and their mapping to brain systems. This analysis confirms many current ideas regarding the neural organization of cognition, but also provides some new insights into the roles of particular brain systems in mental function. We further show that the same methods can be used to identify the relations between mental disorders. Finally, we show that these two approaches can be combined to empirically identify novel relations between mental disorders and mental functions via their common involvement of particular brain networks. This approach has the potential to discover novel endophenotypes for neuropsychiatric disorders and to better characterize the structure of these disorders and the relations between them.     

Insect odorant receptors: channeling scent

Odorant detection in insects involves heterodimers between an odorant receptor (OR) and a conserved seven-transmembrane protein called Or83b, but the exact mechanism of OR signal transduction is unclear. Two recent studies in Nature (Sato et al., 2008; Wicher et al., 2008) now reveal that these OR-Or83b heterodimers form odorant-gated ion channels, revealing a surprising new mode of olfactory transduction.     

Reversible pegylation prolongs the hypotensive effect of atrial natriuretic peptide

Natriuretic peptides (NP), including atrial natriuretic peptide (ANP), induce potent natriuresis and vasodilation and thereby generate hypotension in vivo. Despite intensive efforts, clinical application of NP as an antihypertensive agent is limited because of their short biological half-life and poor bioavailability. Recently, we have developed a strategy that facilitates slow release of peptides from PEG-peptide inactive conjugates, based on reversible pegylation. Peptides prepared by this approach undergo slow, spontaneous chemical hydrolysis at physiological conditions, releasing the native active peptide/protein drug from the inactive conjugates over prolonged periods. A PEG chain of 30 kDa was linked covalently to the alpha-amino side chain of the hormone via a MAL-Fmoc-NHS spacer, yielding PEG 30-Fmoc-ANP, a prodrug that releases the native hormone upon incubation at physiological conditions. Bolus administration of native ANP to Wistar rats receiving adrenaline yields a short, transitory effect in lowering blood pressure (BP), reaching a maximum at 2 min, and then returning to control values after 12 to 25 min. In contrast, administration of PEG 30-Fmoc-ANP lowered BP following a lag period of 50 min, and maintained low BP for a period exceeding 60 min. Saline or PEG 30-Fmoc-Alanine were not effective in lowering BP in Wistar rats. These results show that the novel compound, PEG 30-Fmoc-ANP, is a reversible pegylated prodrug derivative that facilitates a prolonged BP lowering effect in rats and may be considered as a candidate for development into an antihypertensive drug.     

Patient-specific MRI-based 3D FSI RV/LV/patch models for pulmonary valve replacement surgery and patch optimization

A patient-specific right/left ventricle and patch (RV/LV/patch) combination model with fluid-structure interactions (FSIs) was introduced to evaluate and optimize human pulmonary valve replacement/insertion (PVR) surgical procedure and patch design. Cardiac magnetic resonance (CMR) imaging studies were performed to acquire ventricle geometry, flow velocity, and flow rate for healthy volunteers and patients needing RV remodeling and PVR before and after scheduled surgeries. CMR-based RV/LV/patch FSI models were constructed to perform mechanical analysis and assess RV cardiac functions. Both pre- and postoperation CMR data were used to adjust and validate the model so that predicted RV volumes reached good agreement with CMR measurements (error <3%). Two RV/LV/patch models were made based on preoperation data to evaluate and compare two PVR surgical procedures: (i) conventional patch with little or no scar tissue trimming, and (ii) small patch with aggressive scar trimming and RV volume reduction. Our modeling results indicated that (a) patient-specific CMR-based computational modeling can provide accurate assessment of RV cardiac functions, and (b) PVR with a smaller patch and more aggressive scar removal led to reduced stress/strain conditions in the patch area and may lead to improved recovery of RV functions. More patient studies are needed to validate our findings.     

Alternate AChE-R variants facilitate cellular metabolic activity and resistance to genotoxic stress through enolase and RACK1 interactions

Tumorogenic transformation is a multifaceted cellular process involving combinatorial protein-protein interactions that modulate different cellular functions. Here, we report apparent involvement in two independent tumorogenic processes by distinct partner protein interactions of the stress-induced acetylcholinesterase AChE-R and N-AChE-R variants. Human testicular tumors showed elevated levels of N-terminally extended N-AChE-R compared with healthy tissue, indicating alternate promoter usage in the transformed cells. Two-hybrid screens demonstrate that the C-terminus common to both N-AChE-R and AChE-R interacts either with the glycolytic enzyme enolase or with the scaffold protein RACK1. In vitro, the AChE-R C-terminal peptide ARP elevated enolase's activity by 12%, suggesting physiological relevance for this interaction. Correspondingly, CHO cells expressing either human AChE-R or N-AChE-R but not AChE-S showed a 25% increase in cellular ATP levels, indicating metabolic significance for this upregulation of enolase activity. ATP levels could be reduced by AChE-targeted siRNA in CHO cells expressing AChE-R but not AChE-S, attributing this elevation to the AChE-R C-terminus. Additionally, transfected CHO cells expressing AChE-R but not N-AChE-R showed resistance to up to 60muM of the common chemotherapeutic agent, cis-platinum, indicating AChE-R involvement in another molecular pathway. cis-Platinum elevates the expression of the apoptosis-regulator p53-like protein, p73, which is inactivated by interaction with the scaffold protein RACK1. In co-transfected cells, AChE-R competed with endogenous RACK1 for p73 interaction. Moreover, AChE-R-transfected CHO cells presented higher levels than control cells of the pro-apoptotic TAp73 as well as the anti-apoptotic dominant negative DeltaNp73 protein, leading to an overall decrease in the proportion of pro-apoptotic p73. Together, these findings are compatible with the hypothesis that in cancer cells, both AChE-R and N-AChE-R elevate cellular ATP levels and that AChE-R modifies p73 gene expression by facilitating two independent cellular pathways, thus conferring both a selective metabolic advantage and a genotoxic resistance.     

Using fluorescence correlation spectroscopy to study conformational changes in denatured proteins

Fluorescence correlation spectroscopy (FCS) is a sensitive analytical tool that allows dynamics and hydrodynamics of biomolecules to be studied under a broad range of experimental conditions. One application of FCS of current interest is the determination of the size of protein molecules in the various states they sample along their folding reaction coordinate, which can be accessed through the measurement of diffusion coefficients. It has been pointed out that the analysis of FCS curves is prone to artifacts that may lead to erroneous size determination. To set the stage for FCS studies of unfolded proteins, we first show that the diffusion coefficients of small molecules as well as proteins can be determined accurately even in the presence of high concentrations of co-solutes that change the solution refractive index significantly. Indeed, it is found that the Stokes-Einstein relation between the measured diffusion coefficient and solution viscosity holds even in highly concentrated glycerol or guanidinium hydrochloride (GuHCl) solutions. These measurements form the basis for an investigation of the structure of the denatured state of two proteins, the small protein L and the larger, three-domain protein adenylate kinase (AK). FCS is found useful for probing expansion in the denatured state beyond the unfolding transition. It is shown that the denatured state of protein L expands as the denaturant concentration increases, in a process akin to the transition from a globule to a coil in polymers. This process continues at least up to 5 M GuHCl. On the other hand, the denatured state of AK does not seem to expand much beyond 2 M GuHCl, a result that is in qualitative accord with single-molecule fluorescence histograms. Because both the unfolding transition and the coil-globule transition of AK occur at a much lower denaturant concentration than those of protein L, a possible correlation between the two phenomena is suggested.     

Accelerated Biodegradation of Cement by Sulfur-Oxidizing Bacteria as a Bioassay for Evaluating Immobilization of Low-Level Radioactive Waste

Disposal of low-level radioactive waste by immobilization in cement is being evaluated worldwide. The stability of cement in the environment may be impaired by sulfur-oxidizing bacteria that corrode the cement by producing sulfuric acid. Since this process is so slow that it is not possible to perform studies of the degradation kinetics and to test cement mixtures with increased durability, procedures that accelerate the biodegradation are required. Semicontinuous cultures of Halothiobacillus neapolitanus and Thiomonas intermedia containing thiosulfate as the sole energy source were employed to accelerate the biodegradation of cement samples. This resulted in a weight loss of up to 16% after 39 days, compared with a weight loss of 0.8% in noninoculated controls. Scanning electron microscopy of the degraded cement samples revealed deep cracks, which could be associated with the formation of low-density corrosion products in the interior of the cement. Accelerated biodegradation was also evident from the leaching rates of Ca²⁺ and Si²⁺, the major constituents of the cement matrix, and Ca exhibited the highest rate (up to 20 times greater than the control rate) due to the reaction between free lime and the biogenic sulfuric acid. Leaching of Sr²⁺ and Cs⁺, which were added to the cement to simulate immobilization of the corresponding radioisotopes, was also monitored. In contrast to the linear leaching kinetics of calcium, silicon, and strontium, the leaching pattern of cesium produced a saturation curve similar to the control curve. Presumably, the leaching of cesium is governed by the diffusion process, whereas the leaching kinetics of the other three ions seems to governed by dissolution of the cement.     

Response of Spinach Leaves (Spinacia oleracea L.) to Ozone Measured by Gas Exchange,Chlorophyll a Fluorescence,Antioxidant Systems,and Lipid Peroxidation

Spinach (Spinacia oleracea L. cv. Clermont) leaves grown in open-top chambers and exposed to three different concentrations of ozone were measured for gas exchange, chlorophyll a fluorescence, antioxidant systems, and lipid peroxidation at the end of growing season. High O₃ concentration reduced F_v/F_m, indicating that the efficiency in the energy conversion of photosystem 2 (PS2) was altered. The rate of non-cyclic electron transport rate and the capacity to reduce the quinone pool were also affected. The development of non-photochemical quenching was not high enough to decrease the photon excess in the PS2. The limitation of photosynthetic activity was probably correlated with stomata closure and with an increase in intercellular CO₂ concentration. Under oxidative stress, superoxide dismutase (SOD) activity was stimulated in parallel with lipid peroxidation. We did not find any differences in the ascorbate (AsA) pool and ascorbate peroxidase (APX) or glutathione reductase (GR) activities between air qualities. Small, but similar responses were observed in spinach leaves exposed to ambient ozone concentration.     

Dynamics of Inclusion and Exclusion: Comparing Mental Illness Narratives of Haredi Male Patients and their Rabbis

By comparing versions of mental illness narratives – told by Haredi (Utraorthodox Jews) male patients of a mental health clinic in Israel and by their rabbis – this paper relates to two distinct, yet interrelated, theoretical questions: the place and agency of narrators, and the tension between experience and representation. A pair of narratives exemplifies a pattern in which the patients (Talmudic students) tell a narrative of a sudden breakdown related to a dramatic meeting with a non-human figure (often, a woman) or force. Their rabbis, by contrast, tell a narrative that emphasizes their students' mundane symptoms, ``abnormal' and ``immoral' behavior, and use a local adaptation of a Western psychological explanatory model. A dynamic of inclusion and exclusion emerges as students are seeking legitimization and avoidance of stigma, while their rabbis are silencing themes that challenge social and cultural orders. The different narratives are further interpreted in the context of the micropolitics of the interviews and of identity politics between the Haredim and secular Israelis. This social dynamics shows how differently placed social actors-narrators-interpreters construct differently contested and diverse cultural narratives of a seemingly shared reality.