Sortin2 enhances endocytic trafficking towards the vacuole in Saccharomyces cerevisiae

BACKGROUND: A highly regulated trafficking of cargo vesicles in eukaryotes performs protein delivery to a variety of cellular compartments of endomembrane system. The two main routes, the secretory and the endocytic pathways have pivotal functions in uni- and multi-cellular organisms. Protein delivery and targeting includes cargo recognition, vesicle formation and fusion. Developing new tools to modulate protein trafficking allows better understanding the endomembrane system mechanisms and their regulation. The compound Sortin2 has been described as a protein trafficking modulator affecting targeting of the vacuolar protein carboxypeptidase Y (CPY), triggering its secretion in Saccharomyces cerevisiae. RESULTS: A reverse chemical-genetics approach was used to identify key proteins for Sortin2 bioactivity. A genome-wide Sortin2 resistance screen revealed six yeast deletion mutants that do not secrete CPY when grown at Sortin2 condition where the parental strain does: met18, sla1, clc1, dfg10, dpl1 and yjl175w. Integrating mutant phenotype and gene ontology annotation of the corresponding genes and their interactome pointed towards a high representation of genes involved in the endocytic process. In wild type yeast endocytosis towards the vacuole was faster in presence of Sortin2, which further validates the data of the genome-wide screen. This effect of Sortin2 depends on structural features of the molecule, suggesting compound specificity. Sortin2 did not affect endocytic trafficking in Sortin2-resistant mutants, strongly suggesting that the Sortin2 effects on the secretory and endocytic pathways are linked. CONCLUSIONS: Overall, the results reveal that Sortin2 enhances the endocytic transport pathway in Saccharomyces cerevisiae. This cellular effect is most likely at the level where secretory and endocytic pathways are merged. Them Sortin2 specificity over the endomembrane system places it as a powerful biological modulator for cell biology.

Signal transduction networks and the biology of plant cells

The development of plant transformation in the mid-1980s and of many new tools for cell biology, molecular genetics, and biochemistry has resulted in enormous progress in plant biology in the past decade. With the completion of the genome sequence of Arabidopsis thaliana just around the corner, we can expect even faster progress in the next decade. The interface between cell biology and signal transduction is emerging as a new and important field of research. In the past we thought of cell biology strictly in terms of organelles and their biogenesis and function, adn researches focused on questions such as, how do proteins enter chloroplasts? or, what is the structure of the macromolecules of the cell wall and how are the se molecules secreted? Signal transduction dealt primarily with the perception of light (photomorphogenesis) or hormones and with the effect such signals have on enhancing the activity of specific genes. Now we see that the fields of cell biology and signal transduction pathway usually involves multiple organelles of cellular structures Here are some examples to illustrate this new paradigm. How does abscisic acid (ABA) regulate stomatal closure? This pathway involves not only ABA receptors whose location is not yet known, but cation and anion channels in the plasma membrane, changes in the cytoskeleton, movement of water through water channels in the tonoplast and the plasma membrane, proteins with a farnesyl tail that can be located either in the cytosol or attached to a membrane, and probably unidentified ion channels in the tonoplast. In addition there are highly localized calcium oscillations in the cytoplasm resulting from the release of calcium stored in various compartments. The activities of all these cellular structures need to be coordinated during ABA-induced stomatal closure. For another example of the interplay between the proteins of signal transduction pathways and cytoplasmic structures, consider how plants mount defense response against pathogens. Elicitors produced by pathogens bind to receptors on the plant plasma membrane or in the cytosol and eventually activate a large number of genes. This results in the coordination of activities at the plasma membrane (production of reactive oxygen species), in the cytoskeleton, localized calcium oscillations, and the modulation of protein kinases and protein phosphatases whose locations remain to be determined. The movement of ...