RESUMEN
The vacuole of the yeast Saccharomyces cerevisiae plays an important role in nutrient storage. Arginine, in particular, accumulates in the vacuole of nitrogen-replete cells and is mobilized to the cytosol under nitrogen starvation. The arginine import and export systems involved remain poorly characterized, however. Furthermore, how their activity is coordinated by nitrogen remains unknown. Here we characterize Vsb1 as a novel vacuolar membrane protein of the APC (amino acid-polyamine-organocation) transporter superfamily which, in nitrogen-replete cells, is essential to active uptake and storage of arginine into the vacuole. A shift to nitrogen starvation causes apparent inhibition of Vsb1-dependent activity and mobilization of stored vacuolar arginine to the cytosol. We further show that this arginine export involves Ypq2, a vacuolar protein homologous to the human lysosomal cationic amino acid exporter PQLC2 and whose activity is detected only in nitrogen-starved cells. Our study unravels the main arginine import and export systems of the yeast vacuole and suggests that they are inversely regulated by nitrogen.
Asunto(s)
Arginina/metabolismo , Nitrógeno/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Aminoácidos/genética , Transporte Biológico/genética , Humanos , Membranas Intracelulares/metabolismo , Lisosomas/genética , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Saccharomyces cerevisiae/metabolismo , Vacuolas/genética , Vacuolas/metabolismoRESUMEN
In this review article, yeast model-based research advances regarding the role of Amyloid-β (Aβ), Tau and frameshift Ubiquitin UBB+1 in Alzheimer's disease (AD) are discussed. Despite having limitations with regard to intercellular and cognitive AD aspects, these models have clearly shown their added value as complementary models for the study of the molecular aspects of these proteins, including their interplay with AD-related cellular processes such as mitochondrial dysfunction and altered proteostasis. Moreover, these yeast models have also shown their importance in translational research, e.g., in compound screenings and for AD diagnostics development. In addition to well-established Saccharomyces cerevisiae models, new upcoming Schizosaccharomyces pombe, Candida glabrata and Kluyveromyces lactis yeast models for Aß and Tau are briefly described. Finally, traditional and more innovative research methodologies, e.g., for studying protein oligomerization/aggregation, are highlighted.
Asunto(s)
Enfermedad de Alzheimer/metabolismo , Modelos Biológicos , Enfermedad de Alzheimer/fisiopatología , Péptidos beta-Amiloides/metabolismo , Animales , Humanos , Kluyveromyces/metabolismo , Saccharomyces cerevisiae/metabolismo , Schizosaccharomyces/metabolismo , Ubiquitina/metabolismo , Proteínas tau/metabolismoRESUMEN
The yeast proteome includes about 300 polytopic membrane proteins known or predicted to function as transporters. Such proteins ensure active or passive transport of small ions or metabolites across the plasma or internal membranes. Despite decades of research on yeast transporters, many of these remain uncharacterized in terms of substrate selectivity range, subcellular localization, and biological function. Assaying the uptake of radiolabeled compounds into whole cells or isolated organelles remains a powerful method for characterizing the function and biochemical properties of these proteins. Here we describe established protocols for measuring transporter activity in whole cells, intact vacuoles, or reconstituted vacuolar vesicles. These methods have proved particularly useful in the context of our work on yeast amino acid transporters, and can in principle be applied to assaying the uptake of other categories of compounds.