Phosphorylation of a membrane curvature-sensing motif switches function of the HOPS subunit Vps41 in membrane tethering

Tethering factors are organelle-specific multisubunit protein complexes that identify, along with Rab guanosine triphosphatases, transport vesicles and trigger their SNARE-mediated fusion of specific transport vesicles with the target membranes. Little is known about how tethering factors discriminate between different trafficking pathways, which may converge at the same organelle. In this paper, we describe a phosphorylation-based switch mechanism, which allows the homotypic vacuole fusion protein sorting effector subunit Vps41 to operate in two distinct fusion events, namely endosome-vacuole and AP-3 vesicle-vacuole fusion. Vps41 contains an amphipathic lipid-packing sensor (ALPS) motif, which recognizes highly curved membranes. At endosomes, this motif is inserted into the lipid bilayer and masks the binding motif for the δ subunit of the AP-3 complex, Apl5, without affecting the Vps41 function in endosome-vacuole fusion. At the much less curved vacuole, the ALPS motif becomes available for phosphorylation by the resident casein kinase Yck3. As a result, the Apl5-binding site is exposed and allows AP-3 vesicles to bind to Vps41, followed by specific fusion with the vacuolar membrane. This multifunctional tethering factor thus discriminates between trafficking routes by switching from a curvature-sensing to a coat recognition mode upon phosphorylation.     

Proteome profile of human urine with two-dimensional liquid phase fractionation

Two-dimensional liquid chromatography separation (2-DL), based on chromatofocusing for first dimension and hydrophobicity for second, can be used as a complementary method to two-dimensional gel electrophoresis (2-DE). A platform now available, ProteomeLab PF 2D provided by Beckman Coulter, (Fullerton, CA, USA), assembles these methods in automation. This system was applied to resolve large numbers of urine proteins. Reproducibility and sensitivity in protein resolution were evaluated in this study using urines collected from male blood donors. About 1000 peaks were detected at a pH range of 4.0-8.5 by applying 1 mg of proteins. Furthermore, the same fractions showing peaks with high absorbance intensities in second dimension were collected and subjected to matrix-assisted laser desorption/ionization-time of flight/mass spectrometry analysis for identification. The results showed that the 2-DL provides high reproducibility of two-dimensional protein map, and lends fractions to subsequent mass spectrometry analysis without the further need for extraction or solubilization of samples as required for spots excised from 2-DE gels. In addition, this system also allows to separate particularly proteins with 40-9 kDa molecular weight.     

The influence of storage on the “chemical age” of red wines

Storage conditions and duration have a considerable influence on wine quality. Optimum temperature and humidity conditions may improve wine quality through ageing, while incorrect or excessively long storage leads to negative results. In order to evaluate the global effects of storage on red wine composition, 20 Sangiovese wines were stored in two different conditions (cellar or house) for a period of 2 years and analysed every 6 months. Untargeted LC–MS analysis showed various putative markers for the type and length of conservation (i.e. pigments, flavanols, pantothenic acid etc.), while targeted LC–MS confirmed and expanded these results within specific metabolic groups. The results of multivariate analysis showed that wines stored in the cellar changed little even after 2 years of storage, while wines stored in typical domestic conditions (house) developed approximately four times faster, reaching a composition similar to wines stored in the cellar for 2 years after just 6 months. The formation of several monosulfonated flavanols during domestic ageing provided the first evidence in wine of a reaction between wine tannins—both catechins and proanthocyanidins—and the exogenous antioxidant bisulfite. Moreover, ageing in domestic conditions appeared to induce an accelerated decrease in wine pigments, while specifically promoting the formation of pinotin A-like pigments and the hydrolysis of flavonol glycosides.     

Molecular Diagnostics of Fine Needle Aspiration for the Presurgical Screening of Thyroid Nodules

“The incidence of thyroid cancer, the most common endocrine malignancy, is rising. The two most common types of thyroid cancer are papillary and follicular” thyroid carcinomas. “Fine-needle aspiration (FNA) of thyroid nodules” can permit to detect many genetic mutations and other molecular alterations, including RAS and BRAF point mutations, PAX8/peroxisome proliferator-activated receptor (PPAR)γ and “RET/PTC rearrangements, occurring in thyroid papillary and follicular carcinomas” (more than 70% of cases), which can be used successfully to improve the diagnosis “and the management of patients with thyroid nodules”. The most extensive experience has been accumulated with “the diagnostic use of BRAF mutation”, which is highly specific for malignancy. “Testing FNA samples for a panel of mutations” that typically includes RAS, BRAF, PAX8/PPARγ and RET/PTC could permit to achieve the biggest diagnostic impact. “The accuracy of cancer diagnosis in thyroid nodules could be improved significantly using these and other emerging molecular markers”.     

Obstructive lung diseases and inhaler treatment: results from a national public pragmatic survey

Background

The opinions held by the general population on obstructive lung disease and inhaler devices could influence asthma and chronic obstructive pulmonary disorder (COPD) management and treatment adherence.The aim of the present public pragmatic survey was to evaluate the opinions, beliefs and perceptions of Italian people with respect to respiratory diseases as well as their perspectives on the use of inhaler devices.

Methods

This survey was conducted on a group of 2,008 individuals forming a representative sample of the Italian population aged 15 years and over. It was based on personal interviews that were administered in the homes of the respondents using a structured questionnaire that took approximately 30 minutes.

Results

Awareness of obstructive lung diseases is poor. Asthma, but not COPD, was perceived as a common and increasingly prevalent disease by the majority of the interviewees. Allergy, pollution and smoking were considered to be responsible for both of these diseases. The rates at which the respondents claimed to be suffering from asthma and COPD were lower than expected (4% and 2%, respectively). Inhaled drugs were recognised as the main treatment by 65% of the respondents. The great majority of respondents attributed positive characteristics to the inhaler device (e.g., safety, reliability, effectiveness, ease of use and practicality). Compared to people who have never used inhaler devices, individuals who suffer from asthma or COPD were more confident in their use and showed a greater belief in their safety, reliability and trustworthiness. People older than 64 years showed less attention to the properties of these devices.

Conclusions

The present results highlight the need for public interventions aimed at improving awareness of obstructive lung disease and reveal various potentialities and critical issues for inhaler device usage. Switching of devices was considered feasible by most of the interviewees, as long as the choice is carefully explained by their physician.     

Genetic aberrations in macroautophagy genes leading to diseases

The catabolic process of macroautophagy, through the rapid degradation of unwanted cellular components, is involved in a multitude of cellular and organismal functions that are essential to maintain homeostasis. Those functions include adaptation to starvation, cell development and differentiation, innate and adaptive immunity, tumor suppression, autophagic cell death, and maintenance of stem cell stemness. Not surprisingly, an impairment or block of macroautophagy can lead to severe pathologies. A still increasing number of reports, in particular, have revealed that mutations in the autophagy-related (ATG) genes, encoding the key players of macroautophagy, are either the cause or represent a risk factor for the development of several illnesses. The aim of this review is to provide a comprehensive overview of the diseases and disorders currently known that are or could be caused by mutations in core ATG proteins but also in the so-called autophagy receptors, which provide specificity to the process of macroautophagy. Our compendium underlines the medical relevance of this pathway and underscores the importance of the eventual development of therapeutic approaches aimed at modulating macroautophagy.     

Correction to: The evolution of life cycle assessment in european policies over three decades

The International Journal of Life Cycle Assessment -     

The actin cytoskeleton is required for selective types of autophagy, but not nonspecific autophagy, in the yeast Saccharomyces cerevisiae

Autophagy is a catabolic multitask transport route that takes place in all eukaryotic cells. During starvation, cytoplasmic components are randomly sequestered into huge double-membrane vesicles called autophagosomes and delivered into the lysosome/vacuole where they are destroyed. Cells are able to modulate autophagy in response to their needs, and under certain circumstances, cargoes such as aberrant protein aggregates, organelles and bacteria can be selectively and exclusively incorporated into autophagosomes. In the yeast Saccharomyces cerevisiae, for example, double-membrane vesicles are used to transport the Ape1 protease into the vacuole, or for the elimination of superfluous peroxisomes. In the present study we reveal that in this organism, actin plays a role in these two types of selective autophagy but not in the nonselective, bulk process. In particular, we show that precursor Ape1 is not correctly recruited to the PAS, the putative site of double-membrane vesicle biogenesis, and superfluous peroxisomes are not degraded in a conditional actin mutant. These phenomena correlate with a defect in Atg9 trafficking from the mitochondria to the PAS.     

Allometric scaling of lung volume and its consequences for marine turtle diving performance

Marine turtle lungs have multiple functions including respiration, oxygen storage and buoyancy regulation, so lung size is an important indicator of dive performance. We determined maximum lung volumes (V(L)) for 30 individuals from three species (Caretta caretta n=13; Eretmochelys imbricata n=12; Natator depressus n=5) across a range of body masses (M(b)): 0.9 to 46 kg. V(L) was 114 ml kg(-1) and increased with M(b) with a scaling factor of 0.92. Based on these values for V(L) we demonstrated that diving capacities (assessed via aerobic dive limits) of marine turtles were potentially over-estimated when the V(L)-body mass effect was not considered (by 10 to 20% for 5 to 25 kg turtles and by >20% for turtles > or =25 kg). While aerobic dive limits scale with an exponent of 0.6, an analysis of average dive durations in free-ranging chelonian marine turtles revealed that dive duration increases with a mass exponent of 0.51, although there was considerable scatter around the regression line. While this highlights the need to determine more parameters that affect the duration-body mass relationship, our results provide a reference point for calculating oxygen storage capacities and air volumes available for buoyancy control.