Translocon component Sec62 acts in endoplasmic reticulum turnover during stress recovery.

The endoplasmic reticulum (ER) is a site of protein biogenesis in eukaryotic cells. Perturbing ER homeostasis activates stress programs collectively called the unfolded protein response (UPR). The UPR enhances production of ER-resident chaperones and enzymes to reduce the burden of misfolded proteins.

Atg18 function in autophagy is regulated by specific sites within its β-propeller.

Autophagy is a conserved degradative transport pathway. It is characterized by the formation of double-membrane autophagosomes at the phagophore assembly site (PAS). Atg18 is essential for autophagy but also for vacuole homeostasis and probably endosomal functions.

Understanding phosphatidylinositol-3-phosphate dynamics during autophagosome biogenesis.

Autophagosomes, the hallmark of autophagy, are double-membrane vesicles sequestering cytoplasmic components. They are generated at the phagophore assembly site (PAS), the phagophore being the precursor structure of these carriers. According to the current model, autophagosomes result from the elongation and reorganization of membranes at the PAS/phagophore driven by the concerted action of the autophagy-related (Atg) proteins.

Guidelines for the use and interpretation of assays for monitoring autophagy.

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding.

Phosphatidylinositol-3-phosphate clearance plays a key role in autophagosome completion.

Background: The biogenesis of autophagosomes, the hallmark of autophagy, depends on the function of the autophagy-related (Atg) proteins and the generation of phosphatidylinositol-3-phosphate (PtdIns3P) at the phagophore assembly site (PAS), the location where autophagosomes arise. The current model is that PtdIns3P is involved primarily in the recruitment of Atg proteins to the PAS and that once an autophagosome is complete, the Atg machinery is released from its surface back into the cytoplasm and reused for the formation of new vesicles.

Results: We have identified a PtdIns3P phosphatase, Ymr1, that is essential for the normal progression of both bulk and selective types of autophagy.

Reticulophagy and ribophagy: regulated degradation of protein production factories.

During autophagy, cytosol, protein aggregates, and organelles are sequestered into double-membrane vesicles called autophagosomes and delivered to the lysosome/vacuole for breakdown and recycling of their basic components. In all eukaryotes this pathway is important for adaptation to stress conditions such as nutrient deprivation, as well as to regulate intracellular homeostasis by adjusting organelle number and clearing damaged structures. For a long time, starvation-induced autophagy has been viewed as a nonselective transport pathway; however, recent studies have revealed that autophagy is able to selectively engulf specific structures, ranging from proteins to entire organelles.

Multilocus sequence typing of oenological Saccharomyces cerevisiae strains.

This study describes the application of a multilocus sequence typing (MLST) analysis for molecular discrimination at the strain level of Spanish wine yeast strains. The discrimination power of MLST is compared to mitochondrial RFLP analysis. Fragments of the ADP1, ACC1, RPN2, GLN4, and ALA1 genes were amplified by PCR from chromosomal DNA of 18 wine Saccharomyces cerevisiae strains.

Construction of a recombinant autolytic wine yeast strain overexpressing the csc1-1 allele.

During the aging step of sparkling wines and wines aged on lees, yeast cells kept in contact with the wine finally die and undergo autolysis, releasing cellular compounds with a positive effect on the wine quality. In view of the interest of autolysis for wine properties, biotechnologists have tried to improve autolytic yield during winemaking. In this work we used genetic engineering techniques to construct an autolytic industrial strain by expressing the csc1-1 allele from the RDN1 locus.

Mitochondria: one of the origins for autophagosomal membranes?

Macroautophagy is a transport pathway to the lysosome/vacuole that contributes to the degradation of numerous intracellular components. Despite the recent advances achieved in the understanding of the molecular mechanism underlying macroautophagy, the membrane origin of autophagosomes, the hallmark of this process is still a mystery. It has been suggested that mitochondria may be one of the lipid sources for autophagosome formation and that possibly this organelle provides the phosphatidylethanolamine (PE) that covalently links to the members of the ubiquitin-like Atg8/microtubule-associated protein 1 light chain 3 (LC3) protein family.

Regulation of autophagy in yeast Saccharomyces cerevisiae.

Autophagy is a conserved catabolic process that initially involves the bulk or the selective engulfment of cytosolic components into double-membrane vesicles and successively the transport of the sequestered cargo material into the lysosome/vacuole for degradation. This pathway allows counteracting internal and external stresses, including changes in the nutrient availability, that alter the cell metabolic equilibrium. Consequently, the regulation of autophagy is crucial for maintaining important cellular functions under various conditions and ultimately it is essential for survival.

Autophagy in wine making.

Aging that involves contact with dying yeast cells is one of the differential processes between sparkling and still wine production. The release of the products of autolysis during this aging step is fundamental for the quality of sparkling wines made by the traditional method. These cells undergo an autolysis process characterized by self-digestion of yeast intracellular and cell-wall macromolecules, and the release of the degradation products to the wine.

A recombinant Saccharomyces cerevisiae strain overproducing mannoproteins stabilizes wine against protein haze.

Stabilization against protein haze was one of the first positive properties attributed to yeast mannoproteins in winemaking. In previous work we demonstrated that deletion of KNR4 leads to increased mannoprotein release in laboratory Saccharomyces cerevisiae strains. We have now constructed strains with KNR4 deleted in two different industrial wine yeast backgrounds.

Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes.

Research in autophagy continues to accelerate,(1) and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.

Autophagy: from basic research to its application in food biotechnology.

Autophagy is a catabolic process by which the cytoplasm is sequestered into double-membrane vesicles and delivered to the lysosome/vacuole for breaking down and recycling of the low molecular weight degradation products. The isolation in the yeast Saccharomyces cerevisiae of many of the genes involved in autophagy constituted a milestone in understanding the molecular bases of this pathway. The identification of ortholog genes in other eukaryotic models revealed that the mechanism of autophagy is conserved among all eukaryotes.

Transgenic wine yeast technology comes of age: is it time for transgenic wine?

Saccharomyces cerevisiae is the main yeast responsible for alcoholic fermentation of grape juice during wine making. This makes wine strains of this species perfect targets for the improvement of wine technology and quality. Progress in winemaking has been achieved through the use of selected yeast strains, as well as genetic improvement of wine yeast strains through the sexual and pararexual cycles, random mutagenesis and genetic engineering.

Induction of autophagy by second-fermentation yeasts during elaboration of sparkling wines.

Autophagy is a transport system mediated by vesicles, ubiquitous in eukaryotic cells, by which bulk cytoplasm is targeted to a lysosome or vacuole for degradation. In the yeast Saccharomyces cerevisiae, autophagy is triggered by nutritional stress conditions (e.g.

Overexpression of csc1-1. A plausible strategy to obtain wine yeast strains undergoing accelerated autolysis.

The potential of several alternative genetic engineering based strategies in order to accelerate Saccharomyces cerevisiae autolysis for wine production has been studied. Both constitutively autophagic and defective in autophagy strains have been studied. Although both alternatives lead to impaired survival under starvation conditions, only constitutively autophagic strains, carrying a multicopy plasmid with the csc1-1 allele under the control of the TDH3 promoter, undergo accelerated autolysis in the experimental conditions tested.

Evidence for yeast autophagy during simulation of sparkling wine aging: a reappraisal of the mechanism of yeast autolysis in wine.

Yeast autolysis is the source of several molecules responsible for the quality of wines aged in contact with yeast cells. However, the mechanisms of yeast autolysis during wine aging are not completely understood. All descriptions of yeast autolysis in enological conditions emphasize the disturbance of cell organization as the starting event in the internal digestion of the cell, while no reference to autophagy is found in wine-related literature.

Comparison of two alternative dominant selectable markers for wine yeast transformation.

Genetic improvement of industrial yeast strains is restricted by the availability of selectable transformation markers. Antibiotic resistance markers have to be avoided for public health reasons, while auxotrophy markers are generally not useful for wine yeast strain transformation because most industrial Saccharomyces cerevisiae strains are prototrophic. For this work, we performed a comparative study of the usefulness of two alternative dominant selectable markers in both episomic and centromeric plasmids.