Abstract
Macroautophagy is a highly conserved intracellular bulk degradation system of all eukaryotic cells. It is governed by a large number of autophagy proteins (ATGs) and is crucial for many cellular processes. Here, we describe the phenotypes of Dictyostelium discoideum ATG16− and ATG9−/16− cells and compare them to the previously reported ATG9− mutant. ATG16 deficiency caused an increase in the expression of several core autophagy genes, among them atg9 and the two atg8 paralogues. The single and double ATG9 and ATG16 knock-out mutants had complex phenotypes and displayed severe and comparable defects in pinocytosis and phagocytosis. Uptake of Legionella pneumophila was reduced. In addition, ATG9− and ATG16− cells had dramatic defects in autophagy, development and proteasomal activity which were much more severe in the ATG9−/16− double mutant. Mutant cells showed an increase in poly-ubiquitinated proteins and contained large ubiquitin-positive protein aggregates which partially co-localized with ATG16-GFP in ATG9−/16− cells. The more severe autophagic, developmental and proteasomal phenotypes of ATG9−/16− cells imply that ATG9 and ATG16 probably function in parallel in autophagy and have in addition autophagy-independent functions in further cellular processes.
Highlights
Macroautophagy is the major lysosomal route for the turnover of cytoplasmic components
We find that the cell survival after starvation, and the developmental and proteasomal phenotypes of the ATG9/16 double mutant were much more severe than for either of the single mutants, while pinocytosis and phagocytosis defects were similar in all three mutant strains, suggesting that ATG9 and ATG16 probably have cellular functions in addition to their role in autophagy
Successful targeted integration was confirmed by PCR of genomic DNA with gene-specific primers in combination with bsrspecific primers for two independent ATG16 knock-out strains in the AX2 background and for the atg9/atg16 double knock-out strain
Summary
Macroautophagy (hereafter autophagy) is the major lysosomal route for the turnover of cytoplasmic components It serves as a housekeeping mechanism and as a cellular response to different stresses such as starvation, the presence of protein aggregates or intracellular bacteria and is essential for temporary cell survival [1]. It plays a key role in a range of normal developmental processes such as sporulation in Saccharomyces cerevisiae, fruiting body formation in Dictyostelium discoideum, dauer development of Caenorhabditis elegans and pupa formation in Drosophila melanogaster [2,3].
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
