Vacuolar and extracellular maturation of Saccharomyces cerevisiae proteinase A

The vacuolar aspartyl protease proteinase A (PrA) of Saccharomyces cerevisiae is encoded as a preproenzyme by the PEP4 gene and transported to the vacuole via the secretory route. Upon arrival of the proenzyme proPrA to the vacuole, active mature 42 kDa PrA is generated by specific proteolysis involving the vacuolar endoprotease proteinase B (PrB). Vacuolar activation of proPrA can also take place in mutants lacking PrB activity (prb1). Here an active 43 kDa species termed pseudoPrA is formed, probably by an autocatalytic process. When the PEP4 gene is overexpressed in wild-type cells, mature PrA can be found in the growth medium. We have found that prb1 strains overexpressing PEP4 can form pseudoPrA extracellularly. N-terminal amino acid sequence determination of extracellular, as well as vacuolar pseudoPrA showed that it contains nine amino acids of the propeptide, indicating a cleavage between Phe67 and Ser68 of the preproenzyme. This cleavage site is in accordance with the known substrate preference for PrA, supporting the notion that pseudoPrA is formed by autoactivation. When a multicopy PEP4 transformant of a prb1 mutant was grown in the presence of the aspartyl protease inhibitor pepstatin A, a significant level of proPrA was found in the growth medium. Our analyses show that overexpression of PEP4 leads to the secretion of proPrA to the growth medium where the zymogen is converted to pseudoPrA or mature PrA in a manner similar to the vacuolar processing reactions. Amino acid sequencing of secreted proPrA confirmed the predicted cleavage by signal peptidase between Ala22 and Lys23 of the preproenzyme.     

Determination of carotenoids in salmonoids

Summary A routine method for quantitative analysis of carotenoid pigments in salmonoids has been developed, based on an extraction with acetone and subsequent cleaning on silicagel prior to identification by thin-layer chromatography (TLC). Quantitative determinations of astaxanthin and canthaxanthin were performed by high-pressure liquid chromatography (HPLC) on a C-18 column eluted with 20% ethyl acetate and 80% methanol/water (9+1) followed by spectrophotometric detection in the visible region. Other combinations of carotenoids could be separated by this system using isocratic or gradient elution. Carotenoid contents of examined fish meat were 2.3–8.1 mg astaxanthin/kg and 0.3–1.0 mg canthaxanthin/kg. Purified colour extracts from fish could be stored at –18 °C for a least 1 month before a final carotenoid analysis was carried out.

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Bestimmung von Carotenoiden in Salmoniden

Zusammenfassung Es wurde eine Routinemethode für die quantitative Analyse von Carotenoidpigmenten in Salmoniden entwickelt, die auf einer Extraktion der Farbstoffe mit Aceton und einem Reinigungsschritt auf Kieselgel vor der Identifizierung durch TLC beruht. Eine quantitative Bestimmung von Astaxanthin und Canthaxanthin konnte durch HPLC auf einer C-18 Säule und Elution mit Essigester/Methanol Wasser (9 + 1) (20:80) gefolgt von photometrischer Detektion im sichtbaren Bereich erzielt werden. Auch andere Kombinationen von Carotenoiden können mit diesem System getrennt werden, wenn isokratische oder Gradientenelution verwendet wird. Die Gehalte an Carotenoiden betrugen in den untersuchten Fischen 2,3–8,1 mg Astaxanthin/Kg und 0,3–1,0 mg Canthaxanthin/kg. Die gereinigten Farbstoffextrakte sind bei –18 °C mindestens 1 Monat lang lagerstabil.

     

Predictive Validity of an Empirical Approach for Selecting Promising Message Topics: A Randomized‐Controlled Study

Several message topic selection approaches propose that messages based on beliefs pretested and found to be more strongly associated with intentions will be more effective in changing population intentions and behaviors when used in a campaign. This study aimed to validate the underlying causal assumption of these approaches which rely on cross‐sectional belief–intention associations. We experimentally tested whether messages addressing promising themes as identified by the above criterion were more persuasive than messages addressing less promising themes. Contrary to expectations, all messages increased intentions. Interestingly, mediation analyses showed that while messages deemed promising affected intentions through changes in targeted promising beliefs, messages deemed less promising also achieved persuasion by influencing nontargeted promising beliefs. Implications for message topic selection are discussed.     

Electrically Addressable Hybrid Architectures of Zinc Oxide Nanowires Grown on Aligned Carbon Nanotubes

The fabrication and characterization of hybrid architectures of ZnO nanowires (ZNWs) grown on organized carbon nanotubes (CNTs), by a two‐step chemical vapor deposition (CVD) process involving CNT growth from a hydrocarbon source followed by ZNW growth using a Zn metal source, is reported. The ZNWs grow uniformly and radially from individual CNTs and CNT bundles, and the aligned morphology of the CNTs is not disturbed by the ZNW growth process. The nucleation and growth of ZnO crystals on CNTs are analyzed in relation to the classical vapor–solid mechanism. Importantly, the CNTs make uniform and distributed electrical contact to the ZNWs, with up to a 1000‐fold yield advantage over conventional ZNW growth on a flat substrate. Hybrid ZNW/CNT sheets are fabricated by scalable CVD, rolling, and printing methods; and their electrical properties, which are governed by transport through the anisotropic CNT network, are characterized. Functional interaction between the ZNWs and CNTs is demonstrated by photoconductive behavior and photocurrent generation of the hybrid material under UV illumination. There is significant future opportunity to extend these processing methods to fabricate other functional oxides on CNTs, and to build devices that harness the attractive properties of ZNWs and CNTs with high volumetric efficiency over large areas.     

Real Voices in the Technological Wilderness Using ethnographic methods to challenge global wireless paradigms

This paper explores the use of ethnographic research to help companies design, build, and implement products and services that meet the stated and unstated needs of consumers. Although many products enjoy an international presence in the global economy, the market is far from homogeneous. Cultural practices and beliefs strongly influence the meaning and, consequently, the usage of products. If a product is not culturally resonant, the product may not be adopted by consumers - no matter how technologically advanced or innovative the product may be. Specifically, this paper explores mobile phone, and advanced network I-mode adoption in Japan. Research methodologies used in the case study are described, including how these techniques elucidate the various social and cultural processes that influence adoption. In order to develop wireless and other mobile communication solutions that are culturally, emotionally, and technically satisfying for users, the current “ global paradigm” must be reconsidered. Incorporating anthropology into the design process is a crucial first step in helping telecommunication companies define their next generation of products and services in the mobile communications arena.     

A Model‐Based Analysis of Secure Video Transmission Based on IPSec and IPv4

A promising solution to protect wired Internet networks is to use the Secure Internet Protocol (IPSec); however, this has some drawbacks, particularly on the quality of service (QoS). This paper aims at evaluating the video traffic QoS in terms of end‐to‐end delay and packet loss rate. Based on some basic assumptions, our analysis shows that the performance with IPSec is rapidly inferior to the IPv4 performance. We thus suggest adding some QoS parameters into IPSec in order to achieve a compromise between QoS and security.     

Iron Oxide‐Labeled Collagen Scaffolds for Non‐Invasive MR Imaging in Tissue Engineering

Non‐invasive imaging holds significant potential for implementation in tissue engineering. It can be used to monitor the localization and function of tissue‐engineered implants, as well as their resorption and remodelling. Thus far, however, the vast majority of effort in this area of research have focused on the use of ultrasmall super‐paramagnetic iron oxide (USPIO) nanoparticle‐labeled cells, colonizing the scaffolds, to indirectly image the implant material. Reasoning that directly labeling scaffold materials might be more beneficial (enabling imaging also in the case of non‐cellularized implants), more informative (enabling the non‐invasive visualization and quantification of scaffold degradation), and easier to translate into the clinic (cell‐free materials are less complex from a regulatory point‐of‐view), three different types of USPIO nanoparticles are prepared and incorporated both passively and actively (via chemical conjugation; during collagen crosslinking) into collagen‐based scaffold materials. The amount of USPIO incorporated into the scaffolds is optimized, and correlated with MR signal intensity, showing that the labeled scaffolds are highly biocompatible, and that scaffold degradation can be visualized using MRI. This provides an initial proof‐of‐principle for the in vivo visualization of the scaffolds. Consequently, USPIO‐labeled scaffold materials seem to be highly suitable for image‐guided tissue engineering applications.     

Micrometer‐Scale Porous Buckling Shell Actuators Based on Liquid Crystal Networks

Micrometer‐scale liquid crystal network (LCN) actuators have potential for application areas like biomedical systems, soft robotics, and microfluidics. To fully harness their power, a diversification in production methods is called for, targeting unconventional shapes and complex actuation modes. Crucial for controlling LCN actuation is the combination of macroscopic shape and molecular‐scale alignment in the ground state, the latter becoming particularly challenging when the desired shape is more complex than a flat sheet. Here, one‐step processing of an LCN precursor material in a glass capillary microfluidic set‐up to mold it into thin shells is used, which are stretched by osmosis to reach a diameter of a few hundred micrometers and thickness on the order of a micrometer, before they are UV crosslinked into an LCN. The shells exhibit radial alignment of the director field and the surface is porous, with pore size that is tunable via the osmosis time. The LCN shells actuate reversibly upon heating and cooling. The decrease in order parameter upon heating induces a reduction in thickness and expansion of surface area of the shells that triggers continuous buckling in multiple locations. Such buckling porous shells are interesting as soft cargo carriers with capacity for autonomous cargo release.     

Long‐Term Electrical and Mechanical Function Monitoring of a Human‐on‐a‐Chip System

The goal of human‐on‐a‐chip systems is to capture multiorgan complexity and predict the human response to compounds within physiologically relevant platforms. The generation and characterization of such systems is currently a focal point of research given the long‐standing inadequacies of conventional techniques for predicting human outcome. Functional systems can measure and quantify key cellular mechanisms that correlate with the physiological status of a tissue, and can be used to evaluate therapeutic challenges utilizing many of the same endpoints used in animal experiments or clinical trials. Culturing multiple organ compartments in a platform creates a more physiologic environment (organ–organ communication). Here is reported a human 4‐organ system composed of heart, liver, skeletal muscle, and nervous system modules that maintains cellular viability and function over 28 days in serum‐free conditions using a pumpless system. The integration of noninvasive electrical evaluation of neurons and cardiac cells and mechanical determination of cardiac and skeletal muscle contraction allows the monitoring of cellular function, especially for chronic toxicity studies in vitro. The 28‐day period is the minimum timeframe for animal studies to evaluate repeat dose toxicity. This technology can be a relevant alternative to animal testing by monitoring multiorgan function upon long‐term chemical exposure.     

Photoresponsive Sponge‐Like Coating for On‐Demand Liquid Release

Many publications report on stimuli responsive coatings, but only a few on the controlled release of species in order to change the coating surface properties. A sponge‐like coating that is able to release and absorb a liquid upon exposure to light has been developed. The morphology of the porous coating is controlled by the smectic liquid crystal properties of the monomer mixture prior to its polymerization, and homeotropic order is found to give the largest contraction. The fast release of the liquid can be induced by a macroscopic contraction of the coating caused by a trans to cis conversion of a copolymerized azobenzene moiety. The liquid secretion can be localized by local light exposure or by creating a surface relief. The uptake of liquid proceeds by stimulating the back reaction of the azo compound by exposure at higher wavelength or by thermal relaxation. The surface forces of the sponge‐like coating in contact with an opposing surface can be controlled by light‐induced capillary bridging revealing that the controlled release of liquid gives access to tunable adhesion.