[en] The yeast Saccharomyces cerevisiae is a standard model system to study endocytosis. Here we describe the examination of a representative subset of deletion mutants to identify and locate steps in endocytic transport, endosomal/lysosomal acidification and in intracellular transport of hydrolases in non-viral transfection processes. When transport in late endocytosis is inhibited, transfection efficiency is significantly enhanced. Similarly, transfection efficiency is enhanced when the pH-value of the endosomal/vacuolar system is modified. Transfection efficiency is furthermore elevated when the N+/K+ transport in the endosomal system is disturbed. Finally, we observe enhanced transfection efficiency in mutants disturbed in the CVT/autophagy pathway and in hydrolase transport to the vacuole. In summary, non-viral transfection efficiency can be significantly increased by either (i) inhibiting the transport of endocytosed material before it enters the vacuole, or (ii) inducing a non-natural pH-value of the endosomal/vacuolar system, or (iii) slowing down degradative processes by inhibiting vacuolar hydrolases or the transport between Golgi and late endosome/vacuole.
Biochemistry, biophysics & molecular biology
Author, co-author :
Riechers, Sean-Patrick Hermann ; Technische Universität Berlin, Department of Microbiology and Genetics, Institute for Biotechnology, Gustav-Meyer-Allee 25, D-13355 Berlin, Germany.
External co-authors :
Defects in intracellular trafficking and endocytic/vacuolar acidification determine the efficiency of endocytotic DNA uptake in yeast.
Publication date :
Journal title :
Journal of Cellular Biochemistry
John Wiley & Sons, Hoboken, United States - New Jersey